Received: 23 January 2020 Revised: 18 March 2020 Accepted: 9 April 2020

DOI: 10.1111/obr.13040

HUMAN OVERFEEDING/BIOLOGY

The biology of human overfeeding: A systematic review

George A. Bray | Claude Bouchard

Pennington Biomedical Research Center,

Louisiana State University System, Baton
Rouge, Louisiana, USA
Correspondence
George A. Bray, MD and Claude Bouchard,
PhD, Pennington Biomedical Research Center,
Louisiana State University System, Baton
Rouge, LA, USA.
Email: brayga@pbrc.edu; Claude.Bouchard@
pbrc.edu
Funding information
National Institute of General Medical Sciences
(NIGMS), Grant/Award Number: 8-P30-GM-
118430-01
Summary
This systematic review has examined more than 300 original papers dealing with
the biology of overfeeding. Studies have varied from 1 day to 6 months. Over-
feeding produced weight gain in adolescents, adult men and women and in older
men. In longer term studies, there was a clear and highly significant relationship
between energy ingested and weight gain and fat storage with limited individual
differences. There is some evidence for a contribution of a genetic component to
this response variability. The response to overfeeding was affected by the base-
line state of the groups being compared: those with insulin resistance versus
insulin sensitivity; those prone to obesity versus those resistant to obesity; and
those with metabolically abnormal obesity versus those with metabolically normal
obesity. Dietary components, such as total fat, polyunsaturated fat and carbohy-
drate influenced the patterns of adipose tissue distribution as did the history of
low or normal birth weight. Overfeeding affected the endocrine system with
increased circulating concentrations of insulin and triiodothyronine frequently pre-
sent. Growth hormone, in contrast, was rapidly suppressed. Changes in plasma
lipids were influenced by diet, exercise and the magnitude of weight gain. Adi-
pose tissue and skeletal muscle morphology and metabolism are substantially
altered by chronic overfeeding.

KEYWORDS

adipose tissue and muscle metabolism, endocrine and metabolic changes, energy intake and
expenditure, diet composition and mediators of response

1 | I N T RO D UC TI O N 2 | AIMS OF THIS SYSTEMATIC REVIEW

Regulation of body weight is complex and multifactorial. In the
early 20th century, two reports appeared on the empirical mea-
surement of body weight changes under the influence of various
levels of caloric intake.1,2 The paradigm of overfeeding has been
used to probe the complex regulation of human body weight. This
important subject has provided the basis for several reviews.3–27 In
this paper, we present a detailed systematic review of the biology
of human overfeeding against the backdrop of the growing world-
wide obesity epidemic.28
This systematic review summarizes the findings of the past 70 years
of research on the biology of human adaptation to experimental over-
feeding. It brings together the findings from early studies and the
more recent experimental interventions. It covers the changes in body
weight and body composition, adipose tissue topography and biology,
the biology of skeletal muscle, physiology and metabolism at the
organ and tissue levels, hormonal and cytokine profiles, energy metab-
olism and behavioural traits that occur when human beings are
exposed to experimental overfeeding. We also address the

Obesity Reviews. 2020;1–78. wileyonlinelibrary.com/journal/obr © 2020 World Obesity Federation 1

2 BRAY AND BOUCHARD
controversial topic of whether there is overfeeding-induced energy
wasting and the mechanisms that might be involved. Finally, we
summarize evidence regarding recovery from experimental overfeeding
in human volunteers. For the purpose of this review, we have grouped
the available human overfeeding studies into three classes defined by
the duration of the overfeeding protocols. Consideration of biological
and behavioural responses across duration of studies should shed light
on the early and late changes as well as their underlying mechanisms.
3 | STUDIES PRIOR TO 1950
The first two studies reported early in the 20th century were based
on changes in body weight when the investigator was the experimen-
tal subject and exposed himself to increasing or decreasing amounts
of ingested calories (see Bray5 for a full review). One example of these
efforts was that of Rudolf Otto Neumann who undertook several
experiments on himself. In one of them, he consumed variable
amounts of protein (94 g, 89 g or 71 g of protein per day) and calories
(1760 kcal, 2199 kcal or 2403 kcal/day) during three extended periods
and monitored his body weight.2 He concluded that body weight sta-
bility could be achieved over a range of protein and caloric intake.
Neumann introduced the term “Lukus Konsumption” to describe his
observation that he did not gain as much weight as he predicted from
the excess of calories that he had ingested. Similarly, Addison Gulick,
some 20 years later, varied his daily energy intake from a low of
1974 kcal to a high of 4113 kcal over 1 year while recording his body
weight and physical activity.1 He gained less weight than expected on
the high energy intake diet but concluded that the search for the
luxusconsumption should not focus on the resting metabolic rate. In a
re-examination of the energy intake and body weight changes in these
two studies, Forbes showed that their body weight matched quite
closely, albeit not perfectly, to their energy intake in contrast to the
claims of the original reports.29 This is illustrated in Figure 1 drawn by
2 1
Forbes from the data points reported by Neumann and Gulick.
In 1931, Wiley and Newburgh reported on a single subject who
claimed that he could overeat and not gain weight. 30
They “vigorously”
overfed this very lean subject, who was weight stable on 3000 kcal/day,
by providing him with 5000 kcal/day for 15 days. This individual gained
4.5 kg of body weight, with no evidence of luxusconsumption.
These important single-subject studies generated useful observa-
tions that set the stage for experiments with multiple volunteers
which began to appear in the 1950s. However, the debate over the
existence of energy wasting mechanisms in the presence of chronic
overfeeding continued unabated at the onset of the new period char-
acterized by controlled experimental overfeeding.
4 | MATERIAL, METHODS AND
ASSUMPTIONS
Over the last 70 years, more than 110 human overfeeding studies
encompassing more than 150 overfed experimental groups have
F I G U R E 1 Plots of weight change in g/day in relation to daily
energy intake calculated separately from the data of Neumann2
(r = 0.969) and of Gulick1 (r = 0.902). Reproduced from Forbes.29
generated more than 300 scientific papers plus reviews and commen-
taries. These studies have relied on a variety of complex study designs
and broad panels of biological and behavioural phenotypes, such as
doubly labelled water estimates of energy expenditure, 24-h meta-
bolic chamber measurements of metabolic rates, adipose tissue and
skeletal muscle biopsies, euglycaemic-hyperinsulinaemic glucose
clamp, gene expression profiling and imaging of tissues and organs to
highlight the most commonly used technologies.
Parallel arm designs incorporating contrasting overfeeding proto-
cols (e.g., differences in the macronutrient composition of the over-
feeding diet) with or without a control group, and with or without
randomization, have been reported. A number of crossover designs
have been used whereby subjects were exposed to one specific over-
feeding protocol and then subsequently to another protocol in ran-
domized order.31,32 Fully randomized controlled trials on the effects
of overfeeding have also been conducted in a limited number of
cases.33 A recent focus has been on the quantification of the variabil-
ity in adaptation to a given dose of overfeeding with the aim of identi-
fying potential biomarkers or predictors of the response level.34
4.1 | Methodology of the review
This systematic review was registered at Prospero, an international
prospective register of systematic reviews (www.crd.york.ac.uk/
PROSPERO). Guidelines from the Preferred Reporting Items for Sys-
tematic Reviews and Meta-Analyses (PRISMA) group were followed
see.35 Using overfeeding as the primary search term, articles in
English, German and French were identified in several databases.
PubMed yielded 749 publications (www.ncbi.nlm.nih.gov/pubmed),
the Web of Science identified 1500 articles (www.isiknowledge.com)
and the Cochrane Database identified 10 unique papers. All titles
BRAY AND BOUCHARD 3

were reviewed by one author (GAB) and those that appeared to be at least 3% of their body weight or 4 kg. Table 2 includes studies that
inappropriate were noted for concurrence by the other author. A total lasted from 8 to 28 days. Table 3 includes studies where overfeeding
of about 350 original papers were identified in this process and con- lasted 24 h or more, and up to and including Day 7.
stitute the basis of this review. Table 1 includes 25 studies lasting 29 days or more. The number
of subjects ranged from a low of six (see Bray71) to more than 40 indi-
viduals.43,50,55,61,62,98 The longest study was by Sims et al and lasted
4.2 | Nomenclature, calculations and assumptions up to 6 months.63,145 Erdmann et al conducted a study averaging
139 days48 and Bouchard et al reported a study lasting 100 days.146
In this review, we are using the kilocalorie (kcal) nomenclature and Nine studies lasted 56 days. Women were included in 10 of these
have converted joules (1 cal = 4.18 J) and kilojoules (1 kcal = 4.18 kJ) 25 studies. Several studies overfed participants until they gained a
where needed. Our focus is on the metabolizable energy available to prespecified amount of weight.45,48,49,55,61–63,65,147,148 The amount of
support basal metabolism, growth, repair of tissues, pregnancy, lacta- energy that was overfed was expressed in two ways: (1) as a percent-
tion, sedentary activities, physical activity of daily living and exercise age above the energy estimated to maintain body weight as deter-
pursuits. About 5% of the ingested energy is not metabolically avail- mined for each individual at baseline33,44,47,50,52,54,56,66,69,87,88,98; or
able and appears in the feces with traces (<1%) in the urine resulting (2) as a fixed calorie surplus.44,46,58–60,64 The precision in calculating
36
from incomplete oxidation of proteins. The gain in weight associ- the amount of extra energy is important for interpreting energy bal-
ated with chronic overfeeding can be accounted for by increases in ance data and the efficiency of weight gain. Individuals who
adipose tissue and lean tissues assuming that the body water reten- were obese or overweight were included in several
47,49,50,55,66,98,148
tion has not changed from baseline levels. Whereas the gains in adi- studies, but body weight and BMI were within the
pose mass are almost entirely composed of newly accumulated normal range for most participants. The studies by Sims et al,63
triglycerides plus traces of structural elements, the gains in lean tis- Kolaczymski et al,53 and Pasquet149 reported weight gains of >15 kg,
sues are composed of proteins, minerals and water, with the latter but for most of the other studies weight gains ranged between
representing about 70% of the increment in lean mass. For our calcu- 3 and 8 kg. Seven of the studies were conducted with participants liv-
lations, we assumed that the energy content of 1 kg of body fat is ing as inpatients.33,44,46,47,55,58,63,148 The effect of age differences on
9300 kcal whereas that of 1 kg of lean mass is 1020 kcal. the response to overfeeding in men was evaluated by Roberts
Total daily EE in humans can be decomposed into several compo- et al.150 Other reports compared high fat and low fat diets,85 satu-
nents in free-living people and experimental subjects confined to a rated fatty acids versus polyunsaturated fatty acids,62 and low versus
metabolic ward. These components include sleeping metabolic rate, normal- or high-protein diets in two studies.33,57 Another study exam-
basal metabolic rate, resting metabolic rate, thermic effect of food, ined overfeeding in individuals with obesity who were either metabol-
fidgeting and physical activity. Total daily EE is best estimated using ically normal or metabolically abnormal.49 Individuals who are
37
the doubly labelled water technique. The method is based on the constitutionally lean versus normal weight were examined in one
total production of carbon dioxide taking into consideration the differ- study.76,151 The effects of cardiorespiratory fitness51 and of resis-
ential rates of elimination of doses of isotope tracers in the blood, tance training64 on the response to overfeeding were also evaluated.
urine or saliva (2H and 18
O). Total daily EE can also be approximated Table S1 provides a detailed listing of these studies and publications
by the number of calories consumed when individuals are weight sta- derived from them.
ble for a substantial period of time. Resting metabolic rates and ther- It is important to appreciate that overfeeding studies lasting
mic effect of food (standardized meal with single macronutrient or 29 days and more are particularly useful for the evaluation of the joint
mixed composition) are measured using indirect calorimetry either effects of excess calories consumed, specific nutrient overload
with the ventilated hood technique or in a room calorimeter. Fidgeting depending on the overfeeding protocol, and weight gain and body
activities and EE of physical activities are assessed by a variety of composition changes resulting from exposure to the overfeeding
approaches including activity counts, radar detection of movement, treatment.
step-counting, indirect calorimetry and various types of accelerometer Table 2 summarizes the 34 studies of overfeeding lasting from
and movement-detection devices. More information on concepts, 8 through 28 days. Several studies had less than
methods, assumptions and calculations can be found 10 subjects67,71,77,79,80,83,84,86,89,90,93,95–97,99,105,110,137 and two stud-
elsewhere.36,38–42 ies had more than 30.68,136 Women were included in 16 of the
34 studies. The amount of overfed energy was expressed in several
ways: (1) as a percentage of energy above that estimated to main-
4.3 | Overview of studies by duration of overfeeding tain body weight as determined for each individual at
protocol baseline31,68,78,79,81,82,90–93,96,99,105,110,113,118,119,136,137,139,152,153;
(2) as a fixed amount above baseline resting metabolic
70,83,95,154 71,73,75,84,85,97,152
Original studies of overfeeding have been grouped in three tables rate ; (3) as an absolute figure ; or (4) as
according to the duration of overfeeding. Table 1 includes those stud- a fixed amount per unit body weight.72 Individuals with
ies with 29 or more days of overfeeding or where individuals gained obesity were included in several studies,31,70,71,82,93,129,136 but
 4

TABLE 1 Summary of major overfeeding studies lasting 29 or more days or gaining at least 3% or 4 kg of body weight

Initial weight Weight gain

Author Year Days OF Design and diet Overfed % or kcal M/F Age and/or BMI or loss Methods, comments
Alligier, M43 2012 56 days Before & After 70 g lipid (20 g 720 kcal/day 44/0 33 ± 1 79.1 ± 1.8 kg +2.5 kg Fat Bx –Gene Expression; Dietary
butter; 100 g cheese; 40 g 18–30 Records twice;
almonds)
Bouchard, C44 1990 100 days 12 pairs Identical Twins before 1000 kcal/day for 24/0 21 ± 2 60.3 ± 8.0 kg +8.1 kg Inpatient Study in 12 pairs of
and after CHO50;FAT30; 84 days identical twins; Food Intake
PROT15% estimated over 12–14 days
Brands, M45 2013 28–43 days ~7–14 days Eucaloric run-in 40% 9/0 27–43 73.1–95.5 kg +4.3 kg Insulin secretion; insulin
~35 days Hypercaloric 20.6–25.6 response; GLP-1 meassured at
~31 days Hypocaloric Diet end of each period
Bray, GA46 1974 28–42 days Before and After; Supplemental 4000 kcal/day 6/0 24.5 ± 1.9 62.7 ± 1.6 kg +3.0–10.5 kg Cycle ergometry: Exercise
food mostly FAT and CHO efficiency on cycle NOT
affected
Bray, GA33 2012 56 days 8 weeks 5% PROT Diet or; 40% 5/3 6/3 22.9 ± 2.8 69.1 ± 11.6 kg +3.16 kg Parallel arm Inpatient study;
8 weeks 15% PROT Diet or; (945 kcal/day) 5/3 22.9 ± 5.5 77.6 ± 13.0 kg +6.05 kg PROT varied;
8 weeks 25% PROT Diet 26.8 ± 2.0 76.0 ± 15.4 kg +6.51 kg
Diaz, EO47 1992 42 days 3 weeks Bsln 50% 7/0 26.3 ± 4.5 68.9 ± 5.0 kg +7.6 ± 1.5 kg 7-month study; 5 periods; Only
6 weeks OF lean 37.3 ± 9.0 21.7 ± 1.3 Periods 1, 2 & 3 reported here;
6 weeks Free Diet 3/0 84.0 ± 5.0 kg Whole body calorimeter; DLW
6 weeks Underfeeding OW 26.8 ± 1.0 En cost of deposition 26.7 kcal;
6 weeks Free Diet No luxuskonsumption; Post-OF
Diet 55% (range 42–86%) excess
CHO46;FAT42;PROT12 Wt lost
Erdmann, J48 2008 139 ± 26 days Before & after 300–500 kcal/day 10/0 26.2 71.1 ± 3.0 kg +6.2 kg Studied before and at 4-week
+2 BMI CHO40–50; FAT30–40; ± 1.26 21.8 ± 0.7 stable BMI after gain of 2 BMI
PROT15–20% Extra cal Energy units; O-GTT, iv GTT,
Dense
Fabbrini, E49 2015 6% Weight MNO 1170 kcal/day 2/10 43 ± 10 95.8 ± 13.7 kg +5.9 kg MNO defined by IHTG resist Met
Gain MAO 1325 kcal/day Ob 52 ± 7 34.0 ± 3.0 +6.0 kg Comp NOT MAO; Gene
4/4 Ob 103.0 ± 11.0 kg Expression;
35.7 ± 3.9 DXA; CT; CLAMP
Franck, N50 2011 28 days 16 weeks VLCD 450 kcal/day Twice Bsln 18/6 28–61 30.7–48.8 +7.3 ± 3.9 VLCD 52 Adipocyte genes
then 2 weeks refeeding Ob Not Avail Not Avail regulated by caloric intake;
8 weeks VLCD II 450 kcal/day 9/15 21–28 18.7–24 PEDF (Pigment
4 weeks OF Fast Food Diet Ob epithelium-derived factor)
4/2 secreted by fat cell & related to
lean EI
Gentile CL51 2007 56 days High CRF 1000 kcal/day 6/0 21.0 ± 1.0 75.5 ± 5.3 kg +5 ± 0.2 kg High & Low CRF by median split
Low CRF to gain 5 kg 6/0 26.0 ± 2.0 23.6 ± 1.2 +5 ± 0.2 kg High CRF less BF and VAT and
1000 kcal/day 72.8 ± 4.0 kg less rise in BP; MNSA in 2nd
to gain 5 kg 24.2 ± 1.0 study
BRAY AND BOUCHARD
TABLE 1 (Continued)

Initial weight Weight gain

Author Year Days OF Design and diet Overfed % or kcal M/F Age and/or BMI or loss Methods, comments
52
Johanssen, D 2014 56 days Before and After Wt Gain Diet 40% 29/0 26.8 ± 5.4 81.9 ± 10.3 kg +7.6 ± 2.1 kg MRI for hepatic fat; Insulin clamp
EAT Study CHO41;FAT44;PROT15 (10 A-A) 25.5 ± 2.3 4.2 ± 1.4 kg DLW for EE; Fat and muscle Bx
fat
BRAY AND BOUCHARD

Kolaczynski, 1996 1 day & 24 h OF Fast Food 120 kcal/kg 120 kcal/kg 6/2 31.8 ± 3.2 28.1 ± 1.9 72.2 No gain Leptin was focus of study. Acute
JW53 10% gain ingested 5 weeks Sustacal 10% gain 6/0 27.8 ± 0.8 ± 4.6 kg 24.1 16.8 kg (24 h) and chronic study. In
suppl 22.5–27.5 kcal/day + ± 0.9 Chronic study all gained >10%
Bsln in 5 weeks
Koopman, KE54 2013 42 days 6 weeks HF-HS-Large meal size 40% 8/0 22.6 ± 2.7 78.0 ± 5.6 kg +2.4 kg Parallel-arm study; MRI for brain
6 weeks HF-HS-Frequent meals 40% 5/0 21.0 ± 1.4 22.2 ± 0.9 +2.8 kg serotonin; and in
6 weeks HS-Large meals 40% 5/0 21.8 ± 1.6 80.6 ± 9.6 kg +3.3 kg accompanying analysis MRI for
6 weeks HS-Frequent meal 40% 7/0 21.7 ± 2.9 22.6 ± 1.7 +1.9 kg liver fat; 39 began; 30 finished
Control 0% 5/0 23.0 ± 3.1 79.8 ± 7.8 kg −0.5 kg
22.3 ± 0.8
82.5 ± 8.4 kg
23.0 ± 1.3
76.6 ± 7.7 kg
22.3 ± 2.1
Leibel, R55 1995 28–42 days Non-Ob +10% Wt Gain 5–8000 kcal/day 16/7 27 ± 7 66.5 ± 11.8 kg +6.7 kg Formula diet with CHO45;
42–70 days Non-Ob to Initial Wt 5–8000 kcal/day 11 25 ± 7 70.5 ± 11.7 kg −6.8 kg FAT40; PROT15 during Wt
Non-Ob Wt red 10% 11/7 28 ± 8 131.2 ± 25.3 kg +11.9 kg stability, but not during Wt
Ob +10% Wt 9 32 ± 8 132.1 ± 26.9 kg −17.8 kg gain; composition; REE;
Ob return to initial Wt 10 31 ± 8 124.8 ± 29.6 kg −19.2 kg Overfed calories are actual
Ob −10% Wt Loss intake range
Ob −20% Wt Loss
Levine, JA56 1999 56 days CHO40%; FAT40%; PROT20% 1000 kcal/day 12/4 25–36 65.8 ± 10.3 kg 4.7 kg REE, NREE and Leptin in separate
FFM 2.3 kg papers
Fat 2/3
Miller, D57 1967 42 days Expt 1 8 weeks 2.8% PROT 480, 890 kcal 1/1 38, 40 72.5 & 58 kg + 0.3; −0.1 Low PROT groups gained less Wt
28 days Expt 2a 4 weeks 2.6% PROT 1330 kcal/day 2/0 21.5 ± 0.5 64.8 ± 4.8 kg +0.9 than expected based on the
21 days Expt 2b 15.5% PROT 1330 kcal/day 0/6 23.1 ± 1.8 60.5 ± 1.9 kg +3.7 increased calorie load.
Expt 3a 3 weeks 2.8% PROT 1530 kcal/day 3/0 20.6 ± 0.3 67.9 ± 1.8 kg +1.0
Expt 3b 3 weeks 14.9% PROT 1530 kcal/day 0/3 20.6 ± 0.6 64.2 ± 3.1 kg +3.8
Norgan, NG58 1980 44 days 14 days Control Diet: 1482 kcal/day 6/0 21.6 63.6 ± 2.4 kg +6.03 kg Inpatient; Prepared & weighed
CHO46;FAT33;PROT14;A7 ± 0.49 20.9 ± 1.1 FFM 2.3 meals; Alcohol 7% in both
7 weeks OF Diet: CHO38;FAT43; Fat 3.7 periods; Density & lung vol;
PROT12;A7% Water 1.8 L Energy Loss in urine & feces
measured. EE with walking,
sitting
Pasquet, P59 1992 61 ± 65 days Guru Walla Cameroonian 228 000 kcal 9/0 23–35 60.9–79.6 kg +17.0 ± 4 kg 10 days Bsln; REE; TDEE; No
OF festival Cumulative evidence for
Luxuskonsumption
5

(Continues)
 6

TABLE 1 (Continued)

Initial weight Weight gain

Author Year Days OF Design and diet Overfed % or kcal M/F Age and/or BMI or loss Methods, comments
Riumallo, JA60 1989 56 days 4 weeks Customary diet 0 6/0 26.8 ± 4.4 55.0 ± 3.9 kg +1.8 ± 1.2 kg 4 weeks Bsln; underweight
12 weeks Wt Stable Con diet 720 kcal/day 19.6 ± 0.2 subjects REE; TDEE;
8 weeks Wt Gain diet
Romero-Corral, 2010 56 days 8 weeks OF 1000 kcal/day to 22/13 26 ± 3.5 70.8 ± 13.0 kg +3.3 kg 2’Groups – one gained then lost
A61 CHO40;FAT40;PROT20 gain 3–4 kg 5/3 29.6 ± 7.1 22.3 ± 1.9 +1.3 kg Wt; other had no weight
16 weeks return to Bsln Wt 66.6 ± 13.0 kg 0.0 kg change over 8 weeks.
8 weeks Wt maintainers 23.1 ± 3.9 Flow-mediated dilatation;
Related to VAT gain
Rosqvist, F62 2014 49 days (3% 49 days OF PUFA Sunflower 3% Wt Gain goal 13/5 26.7 ± 4.6 67.4 ± 8.2 kg 1.6 kg 41 SS Randomized; PUFA & SFA
gain) Suppl 13/6 27.1 ± 3.6 20.8 1.6 kg provided in muffins; 4 days
49 days OF SFA Palm Oil 63.3 ± 6.8 kg food record; MRI-liver fat;
Suppl 19.9 gene expression;
Sims, EAH63 1968 ~170 days Group I 3000 kcal/70 kg 6/0 21–28 61 kg 21–31% Vermont study Groups I & II; See
Group II 3/0 20–33 70 kg 21–31% Goldman51 for additional
groups
Spillane, M64 2016 56 days High CHO 312 g/day CHO Suppl 1248 kcal/day 10/0 19.4 ± 1.2 86.1 ± 13.5 kg 1.4 kg Resistance trained athletes;
High PROT/CHO Suppl 1248 kcal/day 11/0 21.4 ± 4.1 26.9 3.8 kg Resistance Training &
196 g CHO; 94 g PROT; 22 g 84.3 ± 12.0 kg Overfeeding
FAT/day 26.3
Tchoukalova, 2010 56 days Bsln To Gain 4 kg 15/13 29 ± 2 22.1 ± 0.5 4.6 ± 2.2 kg Fat Cell Bx; DXA; CT; Fat cell size
Y65 (+4 kg) CHO50;FAT35;PROT15 Fat 3.8
OF with “snacks”
Webb, P66 1983 30 days Average American Diet 1000 kcal/day 2/2 40–55 59–108 (2L-2Ob) 2.68 kg Should have gained 5 kg if
60% CHO En 2/2 46–72 (3L-1Ob) 2.73 kg adipose tissue has 6 kcal/g; Ob
70% FAT/PROT En 2/2 63–109 (2L-2Ob) 1.75 kg & Lean same gain, fecal and
urine En & thermogenesis
measured

Abbreviations: A, Alcohol; A-A, African-American; Bsln, baseline; BF, body fat; BMI, body mass index (kg/m2); BP, blood pressure; BW, Body weight; Bx, biopsy; CHO, carbohydrate; CLAMP, euglycaemic hyper-
insulinaemic clamp; CT, computed tomography; CRF, cardiorespiratory fitness; DLW, doubly labelled water; En, energy; EE, energy expenditure; EI, energy intake; Expt, experiment; FFM, fat free mass; GLP-1,
glucagon like protein-1; HF, high fat; HS, high sugar; IHTG, intrahepatic triglyceride; iv GTT, intravenous glucose tolerance test; MAO, metabolically abnormal obesity; MNO, Metabolically normal obesity; MRI,
magnetic resonance imaging; MSNA, muscular sympathetic nerve activity; NREE, nonrusting energy expenditure; Ob, obese; OF, overfeeding; O-GTT, oral glucose tolerance test; OW, overweight; PROT, protein;
PUFA, polyunsaturated fatty acid; REE, resting energy expenditure; SFA, saturated fatty acid; SS, Subjects; TDEE, total energy expenditure; VAT, visceral adipose tissue; VLCD, very low calorie diet; Wt, weight
BRAY AND BOUCHARD
TABLE 2 Summary of major overfeeding studies lasting 8 to 28 days [Colour table can be viewed at wileyonlinelibrary.com]

Initial BMI or Wt gain or

Author Year Days OF Design and diet Overfed % or kcal # M/F Age weight loss Methods, comments
Acheson KJ67 1988 7 days Admission 3642 ± 263 to 4930 3/0 21–22 68.2 ± 5.5 −0.8 kg CHO depletion and then repleated in
3 days glycogen depletion ± 311 kc/d 67.4 ± 6.2 +4.6 kg stepwise fashion.
7 days High CHO 71.9 ± 5.9 −4.4 kg
BRAY AND BOUCHARD

3 days PROT Sparing Mod 67.4 ± 5.4

Fast
Apfelbaum, M68 1971 15 days Grp A Normal Diet 15 days +1500 kcal/day 9/0 Lean N/A N/A — BMR in Chamber decreased
Grp B OF diet Restricted 8/0 Lean N/A N/A +2.6% 0.9%/day with 55 g/day casein diet;
Grp C PSMF 220 kcal/day 41/0 Ob N/A 90.9 kg −8.0% Cycling at 2 rates + walking EE.
casein
Baba, N69 1999 28 days HC CHO60/FAT25/ 40% 4/4 18–30 56.0 +2.5 Body Comp comparing HF & HC diets
PROT15 40% 4/4 ± 2.49 kg ± 0.38 kg
HF CHO35/FAT50/ 19.2 ± 0.41 +2.1
PROT15 ± 0.23 kg
Bandini, L70 1989 13–15 days 6 nonobese adolescents 2.45 × REE 4/2 15.5 ± 1.0 62.1 ± 7.8 kg +2.61 ± 3.8 REE, TDEE (DLW) T3. Insulin, urinary
7 adolescent with obesity 5/2 14.9 ± 0.7 90.4 +2.60 ± 2.6 VMA, plasma NE
HC OF 49% Bsln to 66% OF ± 12.1 kg
Bray, G71 1972 14–21 days 3 M at Peak Wt Nibbling 5000 kcal/day 3/0 Ob 25–42 140–245 kg +1.1–2.7 kg F overate after Wt loss Adipose tissue
Gorging 0/3 Ob 21–22 98–120 kg +1.1–2.4 kg biopsies for lipogenesis.
3 F After Wt Loss +3.6–13.8 kg
Nibbling Gorging +3.5–9.9 kg
50–75% CHO Formula Diet
Claesson, AL72 2009 14 days Candy Supplement 20 kcal/kg/day 5/7 23.2 ± 3.5 67.3 ± 7.6 kg +0.7 kg Insulin; Claims nuts less detrimental than
Peanut Supplement 1346 kcal/day 6/7 23.6 ± 1.8 22.2 ± 1.4 +0.3 kg candy
20 kcal/kg/day 68.7 ± 6.1 kg
1374 kcal/day 22.2 ± 2.0
Cornford, AS73 2012 14 days OF placebo treatment ~2000 kcal/day 7/2 24 ± 1 75.0 ± 2.6 kg +2.4 ± 0.6 Inpatients during 14 days OF; Body
OF + hGH 0.3 mg/day ~2000 kcal/day 7/1 75.3 ± 3.3 kg +3.6 ± 0.6 Comp; Insulin dynamics. Muscle Metab
OF + hGH 1.0 mg/d ~2000 kcal/day 4/1 23.5 ± 0.3
Forbes, GB74 1986 17–21 days 7 days EI = 1.5 × BMR 717 kcal/day 2/13 18–41 44–93 kg +4.4 kg Inpatient; food had 6% PROT 45–50%
OF 2 days at 1195–1793 FFM 2.2 CHO & FAT; fecal losses 5% energy.
OF 15–19 days at kcal kcal/day Fat 2.2 Light PA
range
French, SJ75 1995 14 days 2 weeks dietary inventory 2200 kcal/day 12/0 21–40 74.2 ± 2.4 kg 1.9 kg CCK; Food Intake and food records; VAS
2 weeks of OF HF (58%)
2 weeks control diet
Germain, N76 2014 28 days Constitutional thin 630 kcal/day 0/8 0/8 21.6 ± 1.9 44.6 ± 2.3 kg +0.2 kg Outpatient; Urine metabolomics PYY,
BMI < 17.5 22.1 ± 0.8 17.1 ± 0.3 +0.6 kg ghrelin, GLP1 obestatin REE; DXA; BIA
Normal Wt BMI 18–25 59.2 ± 2.1 kg MRS; Study days were Bsln d 5 OF d
Usual Diet + olive oil, 22.1 ± 0.3 34 after OF Day 62
cheese
Peanuts, Parallel Arm study
7

(Continues)
 (Continued)
8

TABLE 2

Initial BMI or Wt gain or

Author Year Days OF Design and diet Overfed % or kcal # M/F Age weight loss Methods, comments
Goldrick RB77 1972 13 days 13 days Normal Wt 33% 2/0 21–22 55 kg 1.6 & 1.8 kg hGH, Ins, Lipids were main focus
19 days CHO45/FAT40/PPROT15 22 or 35% 2/0 19 50 & 60 kg 3.8 & 4.2 kg
22 days 19 days Nor Wt Fruc or 96% 0/1 24 30 kg 11 kg
Gluc
Anorexia Nervosa
Goran, M78 1994 10 days 19 had control diet 10 days 50% 19/0 23 ± 2 66 ± 4 kg Data only for TDEE by DLW Positive and negative
then 50% + PA 5/0 26 ± 2 75 ± 4 kg FFM energy flux
EI & PA unchanged 10 days Neg E Bal 6/0 25 ± 2 74 ± 4 kg
EI up 50% PA unchanged 6/0 26 ± 3 67 ± 6 kg
10 days 2/0
EI up 50% PA up 50%
10 days
EI Unchanged PA up 50%
10 days
Horton, TJ31 1995 14 days Lean HF Diet 1 week later 50% 9/0 28.6 ± 5.4 68.4 ± 9.9 kg 2.34 kg Caltrac; Chamb 1,7,14 1 meal eaten on
HC Diet 50% 7/0 37.6 ± 5.4 21.3 2.47 kg site/day others were take out.
Obese HF Diet 1 week later 50% 103.9 2.98 kg
HC Diet 50% ± 10.9 kg 3.47 kg
Food given to subjects 32.4
Jebb, SA79 1996 12 days HC 540 g; FAT 120 g 33% 3/0 OF 21–42 60–80 kg +2.90 kg In chamber 12 days for both studies
Restricted: CHO83g; 3/0 UF 19–42 67–77 kg –3.13 kg
FAT20g/d
Jebb, SA80 2006 21 days Dietary fat 43% at low OF 20% 6/0 43.3 ± 10.6 69.0 ± 8.8 kg BF + 13% 21 days at each level with 1 week “free”
Dietary fat 46% at medium 40% intake to test FI Compensation; Leptin,
OF 60% ghrelin
Dietary fat 48% at high OF
Joosen, AM81 2005 14 days OF Diet: CHO 53% 50% 0/14 25 ± 4 64.8 ± 7.0 kg 1.45 kg Gene Expression in subsequent paper
FAT 40% 22.1 ± 2.3 Fat 1.05 2006
PROT 7%
Katzeff, HL82 1986 18 days Lean OF 1000 kcal/day Lean 6/0 23 ± 1 67.5 ± 2.8 kg +2.5 ± 0.5 kg NE Infusion; Exercise thermogenesis see
Ob OF 1000 kcal/day Ob 1/5 25 ± 1 98.9 ± 8.6 kg +1.6 ± 0.3 kg also 1989
After OF Ob UF by −5.5
593 kcal/day ± 0.9 kg
Klein S83 1993 8 days 8 days Hypocaloric diet 1.32× RMR 4/0 30 ± 2 69 ± 2 kg −0.8 Inpatient CRC; TDEE by DLW;
8 days Hypercaloric diet 2.26× RMR 22.9 ± 0.4 ± 0.3 kg RMR
+0.7
± 0.54 kg
Lammert, O84 1982 14 days 14 days Norm diet 2857 kcal/day 8/1 30.1 ± 1.71 71.4 ± 2.4 kg +2.51 ± 0.31 Wt, TBW, VO2 VCO2 measured;
2857 kcal/day Cycle Ergometry; Wt returned to normal
14 days OF at 5714 by 5 months
kcal/day
BRAY AND BOUCHARD

7 days UF at 500 kcal/day

TABLE 2 (Continued)

Initial BMI or Wt gain or

Author Year Days OF Design and diet Overfed % or kcal # M/F Age weight loss Methods, comments
Lammert, O85 2000 21 days Phase 1: 10–14 days Record 1195 kcal/day 10/0 22.4 ± 1.9 73.4 ± 6.7 kg +1.58 ± 0.41 De novo lipogenesis; Inpatient;
of food intake 1195 kcal/day 10/0 22.3 ± 1.7 76.4 ± 8.8 kg +1.35 ± 0.42 Pairs of men were housed together
Phase 2: 2–3 days Bsln data eating either HF or HC diet
BRAY AND BOUCHARD

Phase 3: OF HF
OR OF HC
Mann GV86 1955 28 days Period A Ad lib E bal 7 days 2793–3030 4/0 All were 70.4–90.9 kg 0 Fat Intake kept constant; kcal & Exercise
21 days 2788–3093 kcal/day 2773–3030 24 years 23.5 ± 1.5 +0.9–2.5 kg varied. 3 SS provided Wt change. Main
Period B OF + Ex +5.4–6.8 kg focus lipoprotein and phospholipid
Period C OF No Ex −6.5 kg changes
Period D 14 days Ad lib diet
2014–2274 kcal/day
McLaughlin, 2016 28 days 7 days Wt Stabilization 880 kcal/day 8/7 54 ± 8 86.2 +3.4 kg SSPG; DXA; CT; MRS; Fat Bx with gene
TL87 OW Insulin Sensitive 880 kcal/day 8/8 57 ± 6 ± 10.1 kg +2.7 kg expression
OW Insulin Resistant 29.3 ± 2.4
89.4
± 11.2 kg
30.5 ± 2.6
Meugnier, E88 2007 28 days Fat added to diet including 760 kcal/day 8/0 Lean 23 ± 1 64.6 ± 2.3 kg +1.0 kg Gene expression in muscle;
20 g butter; 100 g cheese; Or 30% 20.7 ± 0.7 DXA, MRS; Dietary rec – 25% excess
40 g Almonds energy (close to expected 30%)
Out-Patient
O’Dea, K89 1982 10 days Hypocaloric 10 days 1000 kcal/day 5/1 18–40 20.0–24.3 −2.92 NE appearance and clearanace related to
400 kcal/day ± 0.24 kg intake. T3 rose, rT3 fell
Eucaloric 10 days Wt Main —
Hypercaloric 10 days +2.58
± 0.53 kg
Olefsky, J90 1975 21 days 7 days Isocaloric Diet 2000 kcal/day 6/2 21–51 50.2–82.7 kg +4.4 kg Inpatient study Insulin resistance;
21 days Hypercaloric Mixed Formula diet with 35kca/kg
Diet + Formula
Passmore, R91,92 1955 a 10–14 days Balanced diet 1700 kcal/day 3/0 21–24 57.0–61.8 kg +1.5 to Food bombed; feces bombed; urine too;
&b +3.5 kg Inpatient trial
Passmore, R93 1963 9 days 9 days energy balance 1185 kcal/day 0/2 25, 35 91.9, 92.8 kg +2.58 & Detailed study of 2 women with obesity
3390 kcal/day restricted 2.86 kg who were below their maximal weight
Diet: CHO39;FAT44; of 112 & 118 kg; gain >2 M per kcal
PROT17 eaten
9 days OF Diet:
CHO64;FAT20;PROT16
5 days Wt loss Diet:
300 kcal/day

(Continues)
9
TABLE 2 (Continued)
10

Initial BMI or Wt gain or

Author Year Days OF Design and diet Overfed % or kcal # M/F Age weight loss Methods, comments
Poehlman, E94 1986 22 days 6 pairs of monozygotic 1000 kcal/day 12/0 19.2 ± 2.3 64.7 ± 8.4 kg +2.5 kg Inpatient; Density with residual volume;
twins 21.3 ± 0.74 Glucose Lipids;
Diet CHO50;FAT35;
PROT15%
Ravussin, E95 1985 9 days 13 days Wt Stable +60% 5/0 22–27 71.3 ± 7.1 kg +3.2 ± 0.2 kg Chamber; density with food eaten in
CHO45/FAT:40;PROT15 21.9 FFM 1.4 front of dietitian; BMR Increased;
9 days OF Mixed diet Fat 1.8
Roberts, SB96 1990 21 days 10 days Wt stable 1000 kcal/day 7/0 23.7 ± 0.4 76.3 ± 4.7 kg +2.48 Outpatient except DLW day; Caltrac for
CHO45;FAT35;PROT20 24.0 ± 1.3 ± 0.44 kg activity Assumed digestibility
11–21 days OF Hi Energy
Foods
21–31 days ad libitum
Robertson, 2004 21 days 7 days Wt stable 932 kcal/day 6/0 lean 21–34 69.3 ± 3.3 kg +2.03 Ghrelin; leptin; gastric emptying oral fat
MD97 21 days HF supplement: 22.2 ± 0.42 ± 0.3 kg tolerance test
peanuts
and cream
Samocha-Bonet, 2010 28 days No Fam Hx – for T2D 1243 kcal/day 12/12 37 ± 12 73.5 +2.2 kg 3 days Bsln than OF Fam Hx Diabetes
D98 Fam Hx + for T2D 9/8 38 ± 12 ± 13.0 kg +3.4 kg gained more; See also Sato for SHBG;
25.1 ± 3.1 See also S-B 2012; Heilmann 2013.
77.3
± 10.0 kg
26.4 ± 4.1
Welle, S99 1983 20 days 6 days Wt Stable CHO45; 1833 kcal/day 7/0 19–36 69 (58–89) +4.8% Inpatient; T3 up 32%; REE up 11.5%;
FAT40; kg FFM 1.4 thermic effect of food no change; NE
20 days OF 70% CHO Fat 1.9 Infusion
Welle, S100 1989 10 days 10 days Wt Stable 1600 kcal/day 6/0 18–40 75.8 ± 5.0 kg +2.59 ± 0.39 Inpatient; REE change and the
45CHO;40FAT;15PROT 1600 kcal/day 5/0 25.3 ± 0.8 +2.81 ± 0.53 beta-blocker propranolol begun on d 4
10 days OF 400 g CHO/day 3/0 80.3 ± 3.3 kg of control and continued through
10 days Weight 10 days OF
Stable-Propranolol
10 days OF 400 g CHO/day
20 days Wt
Stable-propranolol

Abbreviations: Bsln, baseline; BIA, bioelectric analysis; BMR, basal metabolic rate; BMI, body mass index (kg/m2); Bx, biopsy; CCK, cholecystokinin; CHO, carbohydrate; CRC, clinical research unit inpatient facil-
ity; CT, computed tomography; DLW, doubly labelled water; DXA, dual X-ray analysis; EI, energy intake; Ex, exercise; F, Female; Fam Hx, family history; FI, food intake; Fruc, fructose; Gluc, glucose; HC, high car-
bohydrate; HF, high fat; hGH, human growth hormone; M, Male; MRS, magnetic resonance spectroscopy; NE, norepinephrine; Ob, obese; OF, overfeeding; OW, overweight; PA, physical activity; PROT, protein;
PSMF, protein-sparing modified fast; PYY, Protein YY; REE, resting energy expenditure; RMR, resting metabolic rate; rT3, reverse triiodothyronine; SS, subjects; SSPG, steady state plasma glucose method of
insulin clamp; T3, triiodothyronine; TDEE, total energy expenditure (DLW); TBW, total body water; UF, underfeeding; VAS, variable analogue scale; VMA, vanillylmandelic acid; VO2, oxygen uptake; VCO2, car-
bon dioxide expiration; Wt, weight
BRAY AND BOUCHARD
TABLE 3 Summary of overfeeding studies lasting 1 to 7 days [Colour table can be viewed at wileyonlinelibrary.com]

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
Aarsland, A101 1997 4 days Bsln study Day 0 14 476 kca/day (200 5/0 31 ± 4 73 ± 3 kg 2 kg Inpatient-IV & NG infusions;
4 days fed by NG tube kJ/kg/d) (2W & 3B) 23.2 ± 0.36 On Day 4 liver made 3 g/day
(201 kJ/kg/day fat whole body 170 g/d
88% C) and IV (glucose at
BRAY AND BOUCHARD

2.0 mg/kg//min)
Adochio, RL102 2009 5 days 5 days Eu Diet 40% 11/10 27.8 ± 0.9 66.6 ± 1.5 kg Clamp – No Change insulin
CHO50%;FAT30%; 40% 21.8 ± 0.4 sensitivity; IRS-1 up with
PROT20 CHO diet (Cornier group)
5 days HC Diet: Then counterbalanced 5
CHO60%FAT20%; days
PPROT20%
& 5 days HF Diet:
CHO30%;FAT50%;
PROT20%
Apolzan, J103 2014 2 days HF-Low Energy Density 40% 15/5 34 ± 9 92.9 ± 6.2 kg Counter-balanced
HF-High Energy Density 40% 30.7 ± 4.6 cross-over; Outcome was
HC-Low Energy Density 40% the food intake chosen
over 3 post-OF days
Brons, C104 2010 5 days Normal Birth Wt; Control 50% 26/0 24.6 ± 1.0 78.3 ± 9.1 kg Insulin resistance and
HF OF 50% 20/0 24.2 ± 0.5 23.4 ± 2.4 PPARGC1A methylation
Low Birth Wt; Control 77.7 ± 10.9 kg and OXPHOS gene
HF OF 24.8 ± 3.7 expression.
Casper, K105 1990 7 days Tube fed: Isosenergetic 100% 8/0 37.1 ± 80.2 ± 17.7 kg 3.26 kg 2 phases found; Days 1–4
Tube fed 100% excess 13.6 2.58 ± 4.9 Days 4–7; In-patient NG
Tube fed: Isoenergetic feeding
Chen, M106 2014 3 days Natural Conception 1250 kcal/day 6/14 21.5 ± 0.6 64.4 ± 2.8 kg Bsln Diet CHO55%;
IVF Conception 1250 kcal/day 4/10 20.6 ± 0.6 22.4 ± 0.7 FAT30%; PROT15% OF
69.0 ± 4.2 kg Diet CHO40%; FAT45%;
23.2 ± 1.5 PROT15%
Clore JN107 1995 4 days Study I (Glucose Cycle) 1000 kcal/day 6/0 23.8 ± 0.5 75.7 ± 4.1 Progressive OF; Inpatient;
Study II (FFA and 1000 kcal/day 5/4 23.9 ± 0.5 23.2 ± 0.9 REE; Non-diabetics
Gluconeogenesis) 70.6 ± 4.3 Neoglucogenesis; HGO;
Diet: CHO67;FAT22; 21.8 ± 0.7 Overwhelms hepatic
PROT11
Cooper JA108 2009 3 days Days 1–3 Eu (N = 8) 30% Rest or 30% 4/0 (OF 27 ± 7 28 104 ± 11 kg 33.7 ± 12 days Cross-over &
Days 4–6 OF (N = 4) UF UF) 4/0 ±8 3.6 102 ± 13 kg counter-balanced; Leptin
(N = 4) (UF OF) 32.5 ± 3.1 Respond to low Energy
Days 7–9 Eu (N = 8) but not high Energy
Days 10–12 Cross-over
OF & UF
Cornier, MA109 2004 3 days Days 1–7 Thin E Bal Diet 50% 6/0 29.3 ± 7.6 70.8 72 kg Hunger and Satiety in this
Days 8–10 OF 50% 0/7 30.6 ± 8.0 21.3 ± 3.0 paper; FI measured for 1
11

(Continues)
 (Continued)
12

TABLE 3

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
Days 1–7 Reduced Ob 4/0 36.5 ± 7.0 54.96.1 kg day after OF. Only
EB 0/5 38.2 ± 8.3 20.6 ± 1.8 females reduced intake.
Days 8–10 OF 92.47.0 kg Metabolic data in a later
27.5 ± 1.8 (2006) paper
85.39.9 kg
30.4 ± 2.6
Dallosso, H110 1984 7 days 7 days Bsln Diet 50% 8/0 22.9 ± 2.0 69.7 ± 4.9 kg 1.22 kg Inpatient; Two 36H Chamb
CHO57;FAT30;PROT17 21.9 each week, 1 lo & 1 Hi
7 days OF Bsln Diet Cycle ergometry; Fecal &
suppl with fat to Urine E Loss; TEF
bring fat from 30 to 50%
Dirlewanger, M111 2000 3 days Eu (CHO50;FAT35: 40% 0/10 22.4 60.9 ± 2.4 kg Crosssover Counter-bal;
PROT15%) 40% (20–26) 21.9 ± 2.2 REE; Samples on d 4;
OF HC Leptin; 24 Clamp;
(CHO64;FAT25; Subcutaneous
PROT11%) mIcrodialysis for lactate &
OF HF glucose
(CHO35;FAT55:
PROT11%)
Faeh, D112 2005 6 days 7 days Con diet 800–1000 kcal/day 6/0 22.2 25.4 71.5 ± 4.0 kg 1.1 kg Metabolic study Cross-over
(CHO50;FAT35: 800–1000 kcal/day 20.2–25.4 0.6 kg with 12 weeks washout
PROT15%) 800–1000 kcal/day 1.6 kg after fish oil; DNL and
28 days 7.2 g/day Fish insulin sensitivity
Oil + Con Diet
6 days Fruc 3 g/kg
(CHO63;FAT26;
PROT11%)
28 days Fish Oil last 6
days + fruc
Fagerberg, B113 1984 7 days 7 days ad lib Con All 25% 5/0 51.2 ± 5.6 83.6 ± 1.5 kg 0.6 kg Ob Outpatients; 2 of 12
Subjects 25% 5/0 91.6 ± 3.1 kg 0.7 kg excluded; Na22 efflux
OF HF con + butter before and after Wt gain;
OF HC con + sugar Catechols and Thyroid did
not change
Freymond, D114 1989 3 days Con Diet: D1 150% 5/3 12.4 ± 1.5 47.9 ± 5.7 kg Pima 10–15 year old
CHO50;FAT30;PROT20 D2–3 200% 2/6 12.3 ± 1.5 21.3 ± 3.0 children with both lean or
Offspring of obese 64.0 ± 15.4 kg both obese parents;
parent 26.9 ± 4.6 Urinary NE; energy
Diet: CHO50;FAT30; expenditure; sleeping EE;
PROT20 REE.
Offspring lean parents
Diet: CHO50;FAT30;
PROT20
BRAY AND BOUCHARD
TABLE 3 (Continued)

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
Glick, Z115 1977 5 days Period 1: 5 days ad lib ~2300 kcal/day 0/4 NW 20.5 ± 1.8 54.7 ± 3.5 kg Thermogenesis. Wt loss
diet 0/4 OW 21.8 ± 1.7 83.3 ± 4.7 kg during home dieting was 2
Period 2: OF 5 days to 5 kg.
BRAY AND BOUCHARD

Period 3: Home – 4
weeks
Period 4: 2 days
Hagobian, TA116 2006 3 days EB 2 days CHO50–60; 25% 6/0 30 ± 8 78.6 ± 6.1 kg o-GTT; VO2 max; REE
FAT25–30 50% 0/3 23 ± 2 67.6 ± 22.8 kg ×1.5–1.8; Exercise
OF 3 days no Ex equalled extra energy
OF 1 day + Ex = 732
kcal/day
He, J117 2012 3 days 4 days con run-in Wt 50% 15/6 42 ± 9 33.2 ± 6.5 0.67 kg Core T; Accelerometers;
Main Diet Compared ad lib after OF;
3 days Wt Main or OF Gut peptides
3 days ad lib FI Vending
machines
3 days wash-out –
crossed-over to
3 days OF or Wt Main
3 days ad lib FI vending
machines
Hill, JO118 1989 7 days LCTG CHO45;FAT40; 50% 10/0 31 ± 2 79.4 ± 3.8 kg 0.9 kg MCTG 61% C8:0; LCTG
PROT15 50% 26 ± 1 0.8 kg 51% C18:2; 32% C18:1;
MCTG CHO45;FAT40; TEF; REE; Nitrogen
PROT15 balance; Stool & urine
(Cross-over) bombed
Hulston, CJ119 2015 7 days 7 days 65% HF Diet No 50% 7/2 24 ± 2 72.1 ± 4.8 0.6 ± 0.2 kg o-GTT; Probiotics affect
probiotic 50% 7/1 25 ± 2 24.2 ± 1.2 0.3 ± 0.2 kg insulin sensitivity
7 days 65% HF Diet + 73.4 ± 2/3
probiotic 23.5 ± 0.6
Johnstone, AM120 1996 1 day 5 days protocol done 3 40% 6/0 25.3 ± 4.3 71.4 ± 7.0 HP diet more satiating that
times HF or HC. Large EB on 1
Days 1–2 CHO47; day poorly compensated
FAT40; by changes in energy In
PROT13 at 1.6 × RMR the next day
Day 3 OF CHO47;
FAT40;
PROT13 + 550 kcal as
PROT or
FAT or CHO
Days 4–5 ad lib
CHO47;FAT40;PROT13
diet
13

(Continues)
 (Continued)
14

TABLE 3

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
121
Kosmiski, LA 2007 3 days 3 days Eu CHO55; 50% 9/0 10/0 48.0 ± 5/1 22.7 ± 3.7 Study of HIV-lipodystrophy;
FAT30;PROT15; 9/0 38.4 ± 9.3 24.0 ± 2.8 REE measured by hood;
3 days OF 30.1 ± 9.5 24.9 ± 2.3 OF was 50% above 1.3 ×
HIV Lipodystrophy REE; TEF
HIV-Infected No
Lipodystrophy
Healthy Controls
Kovacs, EM32 2006 7 days Ex to Glycogen depl then D4–6 30%; D7–8 10/0 lean 24 ± 5 71.1 ± 5 kg 2.9 ± 0.2 kg Cross-over design of
Days 1–3 HF LC 45%; D10 75 21.8 ± 2.1 hydorxycitrate vs. placibo
CHO25;FAT69;PROT15 with 4 weeks between the
Days 4–10 two 10-day trials; DNL
CHO85;FAT5;PROT10
with
either Hydroxycitrate or
placebo
Lagerpusch, M122 2013 7 days 7 days ad lib diet then 50% 8/0 74.1 ± 8.2 23.1 ± 1.8 −4.3 ± 0.8 Glydemic index; SS OF, then
7 days OF then 50% 8/0 26.0 ± 4.1 22.8 ± 1.6 kg calorically restricted and
21 days Cal Restricted by 50% 77.2 ± 4.8 2.5 ± 1.1 kg then refed with either
50% 27.6 ± 4.2 3.4 ± 1.1 kg Low GI-high fibre or High
14 days either HI glyc-Lo GI low fibre diets.
Fibre diet
or LO glycaemic HI Fibre
diet
Lecoultre, V123 2013 6–7 days Day 2 in chamber; 32 ± 1% 37 ± 1% 7/0 17/0 22.5 ± 1.6 22.4 ± 1.6 Intra-hepatic triglycride
Day 3 ad lib food 30% 10/0 69.3 ± 7.5 kg measured
Fructose 1.5 g/kg/day 11/0 70.7 ± 6.4 kg
Fructose 3.0 g/kg/day 10/0 73.2 ± 6.8 kg
Fructose 4.5 g/kd/day 71.9 ± 5.7 kg
Glucose 3.0 g/kg/day 73.8 ± 6.8 kg
OF 30% Fat
Lundsgaard, AM124 2017 3 days 3 days Con diet 75% 9/0 23 ± 3 23.7 ± 1.7 Randomized 3 period
(CHO62;FAT24; 75% cross-over; CHO diet
PROT14%) increased FGF-21; VO2
3 days HC diet max; Adiponectin; hepatic
(CHO80;FAT9; glucose output; Part of
PROT11%) larger unpublished study
3 days HF diet
(CHO10;FAT78;
PROT12%)
Magkos, F125 2014 1 day 1 day Eu 30% 8/0 38 ± 3 105 ± 7 kg Cross-over; 4 weeks
(CHO55;FAT35; 34 ± 2 between Hourly blood
PROT15%) sampling No effect on
1 day Hypercaloric daytime FFA, glu or Ins-
BRAY AND BOUCHARD
TABLE 3 (Continued)

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
(CHO55;FAT30; Glu & Ins increased FFA
PROT15%) fell at night with OF
McDevitt, RM126 2000 3 days Cross-over – all subjects 50% 0/8 Lean 53.1 ± 0.3 65.6 ± 6.0 kg Each woman had 5 Chamber
BRAY AND BOUCHARD

ate all 50% 0/5 Obese 52.4 ± 4.8 25 ± 1.0 session; OF fructose,
supplements over 5 visits 50% 81.0 ± 4.5 kg glucose, sucrose and fat;
5 days Eu CHO48; 50% 31 ± 4.0 PROT 8% of OF calories
FAT40;PROT12
5 days HF CHO32;
FAT60;PROT8
5 days Fructose
CHO50;FAT42;PROT8
5 days Sucrose
CHO50;FAT42;PROT8
5 days Glucose
CHO50;FAT42;PROT8
Minehira, K127 2003 3 days Cross-over 75% 5/4 26 ± 1 65.4 ± 1.9 kg 1.5 kg DNL increased by CHO
4 days Isocaloric 22 ± 1 feeding at expense of
CHO50;FAT35;PROT15 glycogen; Adipose mRNA
4 days OF CHO71; for fat synthesis genes up
FAT20;PROT9
Murgatroyd, P128 1999 2 days Cross-over 35% 8/0 35.9 ± 6.5 69.2 ± 8.4 kg EI not regulated over 2 day
2 days 35% FAT No Ex BMI < 25 period of either inactivity
ad lib or HF diet.
2 days 35% FAT + Ex ad
lib
2 days 65% FAT No Ex
ad Lib
2 days 65% FAT + Ex ad
Lib
OF Free activity
Nijhuis, J129 2008 7 days Cross-over 53% 2/9 39.1 ± 89.0 ± 18.9 kg 0.8 kg Wt Stable 3.4 yr after
7 days Eu OF; 18.5 31.3 ± 4.9 bariatric surgery; VBG in
7 days OF 3 packets/day 9; Lap-Band in 2; Insulin
900 kcal sensitivcity by SSPG;
Parry, SA130 2017 7 days OF (CHO35;FAT45; 50% 5/4 23 ± 1 65.6 ± 2.1 kg 0.79 ± 0.14 Mixed meal; GLP-1, GIP*,
PROT20) 22.3 ± 0.6 kg FFA, Ins, acylated-ghrelin
Peterson, C131 2016 1 day 28 days at 4 Celsius 5 50% 9/0 23 ± 3 71.5 ± 6.5 kg Cold exposure and OF on
days/week for 4 weeks 23.0 ± 1.8 BAT; CIT & DIT different
1 day OF
Sagayama, H132 2014 3d Bsln Body Comp twice 1500 kcal/day 10/0 23.1 ± 1.6 63.6 ± 4.5 kg 0.7 ± 0.5 kg Body composition; All SS
3 days OF repeat Body 21.6 ± 1.3 returned Bsln Wt < 2
Comp weeks
15

(Continues)
 16

TABLE 3 (Continued)

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
Schmidt, SL133 2012 3 days Cross-over; 25% 8/15 ObPr 28.5 ± 2.3 23.8 ± 2.7 Obesity Prone and Obesity
CHO50;FAT30;PROT20; 16/16 28.1 ± 2.9 20.6 ± 2.2 Resistant people in
3 days Eucaloric ObRes Chamber on eucal and
3 days Hypercaloric hypercal diet PAMS
Activity
Schwarz, JM134 1995 5 days 5 days Eu then 67% 6/0 35 ± 3 74.5 ± 5.2 kg 2.3 ± 0.2 kg DNL; Hepatgaic glucose
sequentially 28% 24.2 ± 1.2 −2.8 ± 0.4 production; Ra & Rd; CHO
5 days OF-50% CHO 55% kg and FAT oxidation
5 days 50% decrease in 2.1 ± 0.4 kg
CHO
2 weeks ad lib hiatus
followed by
5 days 25% excess CHO
5 days 25% decrease in
CHO
5 days OF 50% FAT
Sobrecases, H135 2010 7 days 7 days Isocaloric (C55; 35% 30/0 23.9 ± 0.4 22.6 ± 0.2 0.3 ± 0.1 kg Intrahepatic lipid by MRS. Fr
F30;P15) 30% 12/0 & Fat increase IHCL but
7 days Fructose 3.5 g/kg 30% 10/0 only Fat in TG VLDL
FFM 8/0
4 days Hi Sat Fat
4 days Fruc + Hi Sat Fat
Sun, G136 2007 7 days Parallel Arm Lean <21% 70% 24/0 23.0 ± 0.5 70.6 ± 2.2 kg 2.0 ± 0.23 Visfatin; Retinol Bindng
FAT 14/0 21.8 ± 0.8 77.8 ± 1.1 kg kg Proten; Intgerleukin-6; Ins;
OW 21–25.9% FAT 23/0 22.7 ± 0.5 93.0 ± 3.3 kg 1.6 ± 0.27 DXA body comp
Ob ≥ 26% FAT kg
2.6 ± 0.24
kg
Thearle, MS158 2013 1 day Randomized cross-over 200% 15/0 38.6 ± 7.8 84.1 ± 8.8 kg Part of a large on-going
1 day OF CLP CHO75; 0/5 30.7 ± 26.6 ± 1.9 study. Intrinsic EE
FAT22;PROT3 10.3 82.9 ± 19.6 kg response to OF negatively
1 day OF CNP 31.8 ± 7.4 associated with adiposity
CHO75;FAT5;PROT20 – but only small fraction
1 day OF LPF of ingested EI
CHO51;FAT46;PROT3
1 day OF FNP
CHO20;FAT60;PROT20
1 day OF Balanced OF
CHO50;FAT30;PROT20
1 day Fasting
BRAY AND BOUCHARD
TABLE 3 (Continued)

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
Utiger, RD137 1982 7 days 2 days Usual Diet 2000 kcal/day 4/4 25–30 N/A +2.1 ± 0.2 T3 Increase; TSH and T4
7 days OF dextrose 3/2 of 8 kg not; Follow-up 1–10
BRAY AND BOUCHARD

supplement −0.7 ± 0.2 mtonths later. Rx T4 not

1–10 months later, same kg OF
diet, plus
Rx T3 but Not OF
Van Aggel-Leijssen, DP138 1999 1 day Randomized cross-over 1123 kcal 8/0 23.5 ± 7.0 21.4 ± 2.3 Hourly measurement of
1 day EB No Ex 1577 kcal leptin in day every 2 h at
1 day E OF + Extra Ex night. Exercise decreases
1 day 1123 kcal Ex neg peak & avrg leptin; Pos EB
EB increases 24 h leptin
1 day Ex + extra food
Walhin, JP139 2013 7 days 7 days OF reduced 4000 50% 14/0 25 ± 7 75.5 ± 8.9 kg 2.7 kg Vigorous Exercise offsets
steps/day 75 ± 3% 12/0 78.2 ± 11.0 kg 1.6 kg most effects of
7 days OF + Vig Ex 45 overfeeding
min 75% max
Weststrate, JA140 1990 4 days Cross-over with two Graded increase OF 5/5 23 ± 0.8 64.2 ± 3.6 kg 2 × 2 design with Groups A
8-day periods 15%, 30%, 45% and B with CHO OF with
D 1–4 Wt Main ± Ex and 60% and without exercise;
Day 5–8 OF ± Ex Hood; RMR; DIT
OF schedule: D5 + 15%
D6 + 30% D7 + 45%
D8 + 60%
Weyer, C141 2001 2 days Randomized cross-over 100% 14/0 30.4 ± 6.1 117.4 ± 7.4 kg Chamber; Are Pima different
Mixed 7/0 Causas 31.7 ± 7.6 38.3 ± 17.2 from Caucsians? SMR and
Diet CHO50;FAT30; 7/0 Pima 29.2 ± 4.3 110.4 ± 49.0 kg EE increased with OF and
PROT20 6/0 Lean 27.5 ± 6.2 34 ± 16.5 decreased with fasting
3 days Bsln mixed diet 8/0 Obese 32.7 ± 5.2 124.3 ± 52.9 kg
2 days Bsln diet 42.6 ± 18.0
2 days Fasting 71.3 ± 5.2
2 days OF by 100% 22.4 ± 2.9
152.0 ± 36.7 kg
50.2 ± 12.9
Wijers, SL142 2007 3.5 days EB thermoneutral temp 60% 13/0 22.8 ± 1.7 79.6 ± 4.0 kg TDEE and PA increased in
22 C 23.0 ± 0.9 cold. Much Interindividual
3.5 days OF at 22 C Variation; Respiration
EB at reduced temp of Chamber 2 times
16 C
Diet (CHO47;FAT38;
PROT15%)

(Continues)
17
 (Continued)
18

TABLE 3

Author Year Days OF Design and diet OF % or kcal # M/F Age Initial BMI Wt gain Methods, comments
143
Wulan, SN 2014 4 days Eu CHO55;FAT30; 50% 10/0 S 27 ± 2 68.9 ± 7.4 kg S Asians and Whites
PROT15 Asian 24 ± 2 23.3 ± 3.0 matched for % Body fat. S
Hypercaloric 10/0 White 88.3 ± 18.1 kg Asians more adverse
CHO25;FAT60;PROT15 27.0 ± 4.2 effects on lipids
Zed, C144 1986 6 days 6 days Bsln (2390 1028 kcal/day 0/8 Lean 25 ± 8 50.2 ± 5.7 kg 1.1 ± 0.7 kg Women lived in metabolic
kcal/day) 0/8 Obese 37 ± 8 116.5 ± 21.1 kg 1.0 ± 1.1 kg unit. Metabolic Chamber
6 days OF (Fat 4.3 MJ/d) on 8 occasions
10 days Low EI Diet
(1195 kcal/day)
6 days Refeeding (Fat 4.3
MJ/d)

Abbreviations: BAT, brown adipose tissue; Bsln, baseline; BMI, body mass index (kg/m2); Bx, biopsy; CCK, cholecystokinin; CHO, carbohydrate; CIT, hydroxicitrate; CLP, high CHO, low PROT OF; CNP, high
CHO, normal PROT; Con, control; DIT, dietary induced thermogenesis; DLW, doubly labelled water; DNL, de novo lipogenesis; DXA, dual X-ray analysis; EB, energy balance; EI, energy intake; Eu, eucaloric; Ex,
exercise; F, Female; Fam Hx, family History; FFA, free fatty acids; FFM, fat free mass; FGF-21, fibroblast grown factor-21; FI, food intake; FNP, high fat, normal PROT OF; Fruc, fructose; GLP-1, glucagon like
protein-1; GIP, gastric inhibitory peptide or glucose-dependent insulinotropic peptide; HC, high carbohydrate; HF, high fat; hGH, human growth hormone; HGO, hepatic glucose output; Hi GI, high glycaemic
index; HP, high protein; IHCL, intrahepatocellular lipid; IHTG, intrahepatic triglyceride; IRS-1, insulin receptor substrate; IV, intravenous; IVF, in vitro fertilization; kJ, kilojoules; LCTG, long chain triglyceride; LPF,
low PROT OF; M, Male; MCTG, medium chain triglyceride; MF, medium fat; MJ, megajoule; MRS, magnetic resonance spectroscopy; NE, norepinephrine; NG, nasogastric; ObPr, obesity prone; ObRs, obesity
resistant; OF, overfeeding; O-GTT, oral glucose tolerance test; OW, overweight; OXPHOS, oxidative phosphorylation in mitochondria; PA, physical activity; PPARGC1A, peroxisome proliferator-activated
receptor-γ co-activator 1A; PAMS, physical activity monitoring system; PBO, placebo; PROT, protein; PSMF, protein-sparing modified fast; PYY, Protein YY; Ra, measurement of glucose appearance rate; Rd,
measurement of glucose disposal rate; REE, resting energy expenditure; RMR, resting metabolic rate; rT3, reverse triiodothyronine; SS, subjects; SSPG, steady state plasma glucose; T3, triiodothyronine; T4, thy-
roxin; TDEE, total energy expenditure; TEF, thermic effect of food; TG, triglyceride; TSH, thyroid stimulating hormone; UF, underfeeding; Urine E, urinary epinephrine; VBG, vertical banded gastroplasty surgery;
VLDL, very low density lipoprotein; VO2, oxygen uptake; Wt, weight.
BRAY AND BOUCHARD
BRAY AND BOUCHARD
body weight and BMI were within the normal range for most
of the studies. Half of the studies were conducted
44,67,70,71,73,74,78–80,83,85,89–94,99,100,105,110,118,137
inpatients.
the studies in listed in Table 2, one examined overfeeding in adoles-
cents with and without obesity.70 Some examined dietary effects
including nibbling versus gorging71 and high-fat versus high-
31,67,79,85,113
carbohydrate diets, and candy versus peanuts.
studies included medications, one of which was propranolol100 and
the other growth hormone.73 One study examined overfeeding in a
patient with anorexia nervosa77 and another one in subjects with a
family history of type 2 diabetes.98
It is useful to remember here that overfeeding protocols lasting
from 8 days to about 1 month are likely to generate the most
ambiguous findings as their post-overfeeding measurements are
likely to be influenced by the excess calories consumed, the
nutrient composition of the caloric surplus and the body weight
and adiposity changes that have occurred over the overfeeding
period.
The 46 studies in Table 3 lasted from 1 to 7 days. Twenty-one
of these studies were from 1 to 3 days in length and 19 of them
included women. Participants with overweight or obesity were rep-
resented in fifteen studies.103,108,114,115,125,126,141,144 One study
examined Pima Indian children and their parents.114 Measurement
of food intake at the end of overfeeding103,117,120 and neural-
imaging of the brain in response to food and nonfood cues109
were included in some studies. Individuals who were categorized
as being “at-risk” of developing obesity were contrasted to those
who were resistant to obesity109,155; individuals with normal or
low birth weight were compared156; individuals of South Asian
extraction were compared with Europeans143; individuals conceived
by in vitro fertilization were compared with those conceived nor-
mally106; and individuals who did or did not have human immuno-
deficiency virus (HIV) were compared.121 Contrasts between levels
of carbohydrate, fat and protein with or without exercise were
102,103,111,124,128,138,140,157–159
examined in several studies,
effect of added fructose was also examined.112,123,126
Short-term overfeeding studies over a few days are of
particular interest for the investigation of the effects of excess
calories or excessive consumption of specific nutrients on metabo-
lism because body weight and adiposity changes are generally
minimal.
5 | C H A N G E S I N B O D Y WE I G H T A N D B O D Y
C O M P O S I T I O N WI TH OV E R F E E D I N G
Two studies have focused on the maximal amount of body weight
that could be gained when subjects ate as much as possible for an
extended time period.59,63 In the Vermont Experimental Overfeed-
ing Study conducted on state prisoners by Ethan Sims and collabo-
rators, subjects ate as much as they could in order to gain at least
25% of their initial body weight over a period of about 200 days.
Three of their nine subjects could not achieve this goal even
19
though they increased their body weight by 18% to 21%. The
as mean weight gain of the group reached about 15 kg (extrapolated
Among from graph) or 25%.63 In Cameroon, nine young men participated
in the 2-month long traditional fattening session (the Guru
Walla).59 It was estimated that these young men ate on average an
excess of 228 000 kcal over 2 months, or about 3800 excess calo-
72
Two ries per day. The mean weight gain reached 17 kg with a range
from 12 to 23 kg.
In contrast, most overfeeding experiments with human subjects
have relied on a fixed number of excess calories or a caloric sur-
plus defined as a percentage of the habitual energy intake or of
the energy intake associated with weight maintenance or a target
weight gain as percentage of baseline weight. From the 13 long-
term studies comprising 19 different experimental groups from
which the number of calories can be calculated, the gain in body
weight, fat mass and fat free mass can be plotted as a function of
the overfed calories. The characteristics of these studies lasting
9 to 100 days are summarized in Table 4.
5.1 | Pattern of weight gain with overfeeding
Statistically significant body weight increases are generally not seen
until after a week of chronic overfeeding with a daily caloric excess of
about 1000 kcal.83,118,123,144 The weight gain in response to over-
feeding ranging from 17 000 to 84 000 kcal is displayed in Figure 2.
The weight gain ranged from 1.4 to 8.1 kg with heterogeneity in
weight change at any given level of overfed calories. The relationship
between the amount of weight gain and the number of overfed calo-
ries appears to be linear. There was no substantial difference in
weight gain between the six inpatient studies versus those performed
on free-living subjects. Two of the overfed groups received a low-
protein diet, and their weight gain is less than the other groups for the
same number of overfed calories (Figure 2). The r2 between the num-
and the ber of overfed calories and the weight gain reaches 0.82 (Panel A).
Assuming a linear relationship, the confidence intervals of the regres-
sion line are relatively small, about 0.9 kg for a given level of over-
feeding. If one excludes the two low-protein diet groups (Panel B), the
correlation between the total number of overfed calories and the
measured weight gain in the remaining 17 groups is r2 = 0.87, with a
standard error of estimate of 0.8 kg. This is a good fit with the
predicted weight gain if one considers all potential sources of energy
losses.
5.2 | Human variability in body weight gain with
overfeeding
The issue of the time course of the weight gain across the duration the
overfeeding was addressed using data on pairs of identical twins over-
fed for a period of 100 days (total excess energy intake reached
84 000 kcal).161 Body weight was measured daily and showed that
during the first few weeks of overfeeding, most of the energy surplus
20 BRAY AND BOUCHARD

TABLE 4 Overfeeding studies and experimental groups with complete data on excess calories consumed and body weight and composition
changes

Study OF kcal (total) Weight gain, kg FM gain, kg FFM gain, kg BE gain, kcal BE % of OF
1 Bouchard C, 1990 84 000 8.10 5.40 2.70 52 974 63%
2 Bray GA, 2012 50 729 3.16 3.66 −0.70 33 324 66%
3 Bray GA, 2012 50 729 6.05 3.45 2.87 35 012 69%
4 Bray GA, 2012 50 729 6.51 3.44 3.18 35 236 69%
5 Cornford AS, 2012 24 500 2.40 1.60 0.70 15 594 64%
6 Diaz EO, 1992 62 202 7.60 4.60 3.00 45 840 74%
7 Horton TJ, 1995 18 606 2.47 1.09 1.38 11 545 62%
8 Horton TJ, 1995 18 606 2.31 1.21 1.10 12 375 67%
9 Horton TJ, 1995 23 366 3.47 2.06 1.41 20 596 88%
10 Horton TJ, 1995 23 366 2.98 1.90 1.08 18 772 80%
11 Johannsen DL, 2014 66 156 7.60 4.20 3.40 42 528 64%
12 Joosen AMCP, 2005 18 732 1.45 1.05 0.40 13 893 74%
13 Levine JA, 1999 56 000 4.70 2.30 2.40 23 838 43%
14 Norgan NG, 1980 65 560 6.03 3.70 2.33 36 787 56%
15 Poehlman ET, 1986 22 000 2.24 1.17 0.99 21 842 99%
16 Ravussin E, 1985 17 226 3.21 1.80 1.41 18 178 106%
17 Samocha-Bonnet D, 2010 28 000 2.20 1.48 0.50 14 274 51%
18 Samocha-Bonnet D, 2010 28 000 3.40 1.96 1.00 19 248 54%
19 Webb P, 1983 30 000 2.39 1.67 0.72 16 265 54%
Abbreviations: OF, overfeeding; FM, fat mass; FFM, fat-free mass; BE, body energy.

was recovered as body weight and body energy gain, whereas these
gains represented only about 60% of the overfed calories towards the
end of the 100-day experimental treatment. Hence, an exponential
increase in body weight was observed with a half-time of 86 days,
which suggests that weight gain was slowing down towards the end of
the overfeeding period, possibly as a result of the extra energy needed
for resting energy expenditure and to move the heavier body.161
There is considerable human variability in responsiveness to
chronic overfeeding. To quantify inter-individual differences in weight
gain, we relied on studies in which a fixed number of overfeeding calo-
ries or a percentage of daily energy intake were prescribed. We focused
on studies with a total caloric overload of at least 50 000 kcal and with
individual data on weight gain. Five studies with seven experimental
groups met this criterion (Table 5). The studies lasted from 42 to
100 days and the caloric overload ranged from 53 000 to 84 000 kcal.
The main weight gain across the seven experimental groups was
between 3.2 to 8.1 kg, with the lowest weight gain registered in a low-
33
protein diet group. There were considerable inter-individual differ-
ences in weight gain in each of the seven groups. The difference
between the lowest and highest gainers in each experimental group
ranged from 1.8-fold to 5.1-fold, with a mean of about threefold across
all groups. There were no apparent differences between the inpatient
overfeeding studies33,44 and the studies performed on free-living sub-
47,56,58
jects. However, there is a strong trend in the Bray et al study for
a reduced heterogeneity in response of weight gain in the presence of
an increasing protein content in the caloric overload.33
What are the reasons for this heterogeneity in response to weight
gain under standardized overfeeding conditions? One must begin by
questioning the precision of the baseline estimates of the energy cost
of weight maintenance. This is clearly a challenging problem as under-
estimating or overestimating the true energy cost of weight mainte-
nance prior to overfeeding would cause a lower or higher caloric
overload than predicted during the overfeeding period. In the study
by Bray et al,162 energy intake estimated from weight balance was
2263 ± 250 kcal/day in the low-protein diet group,
2478 ± 512 kcal/day in the normal-protein diet group and
2487 ± 545 kcal/day in the high-protein diet group. The
corresponding values from doubly labelled water were
2233 ± 360 kcal/day in the low protein group, 2176 ± 528 kcal/day in
the normal protein group and 2090 ± 453 kcal/day in the high protein
group. Although the values were close in those assigned to the low-
protein diet, the weight balance method overestimated energy needs
by 300 to 400 kcal/day.162
Could this problem account for the variability in weight gain
observed in well-controlled overfeeding experiments? Probably not
entirely based on the following observation. In the long-term over-
feeding protocol (100 days) conducted with 24 young men (12 pairs
of identical twins) living as inpatients for the duration of the study,44
the correlation between the number of calories associated with
weight stability at baseline with the weight gain in response to over-
feeding was low and not statistically significant (r = 0.26). If the energy
cost of weight maintenance had been substantially underestimated or
BRAY AND BOUCHARD 21

F I G U R E 2 (A) Plot of weight gain in

relation to the number of overfed calories
in 19 experimental groups retrieved from
13 studies.31,33,44,47,52,56,58,66,73,81,95,98,160
Groups on a low-protein diet and groups
derived from inpatient studies are
contrasted to others. (B) Plot of weight
gain in relation to weight gain in the
remaining 17 experimental groups once
the low-protein overfeeding diet studies
have been removed. Both regressions,
p < 0.0001. The study numbers in both
panels are defined in Table 4

TABLE 5 Illustration of individual differences in response to experimental overfeeding

Overfeeding duration/excess Mean weight gain Difference between highest and lowest weight
Study N calories (kg) gain (fold)
Bray et al., JAMA 2012*
Group 1—5% Protein 8 56 days/ 3.2 4.4
Group 2—15% Protein 9 53 000 kcal 6.1 2.6
Group 3—25% Protein 8 6.5 1.8
Levine et al., Science, 1999 16 56 days/56 000 kcal 4.7 5.1
Diaz et al., Am J Clin Nutr, 9 42 days/62 000 kcal 7.6 2.1
1992
Bouchard et al., NEJM, 1990 24 100 days/84 000 kcal 8.1 3.0
Norgan, Durnin, Am J Clin 6 42 days/65 560 kcal 6.0 2.3
Nutr, 1980
22
overestimated, one would have found a strong correlation with the
amount of weight gained.
Another potential factor contributing to the variability in response
to standardized overfeeding is in the fidgeting and low EE physical
activities taking place constantly but that cannot be fully controlled
even under the most rigorous experimental conditions. It has been
proposed that this component plays an important role in variation in
weight gain.56 Individual biological characteristics may also contribute
to the variability of weight gain even under fully standardized and
controlled conditions. This hypothesis is supported by a detailed anal-
ysis of the biological traits that were correlated with weight gain in
the study of 24 young men (12 pairs of identical twins) overfed by
84 000 kcal over 100 days. The findings are depicted in Figure 3.34
The strongest and most consistent correlates of gains of body weight,
adiposity and energy between the six highest and six lowest gainers
were baseline low fat-free mass, low skeletal muscle oxidative poten-
tial, low cardiorespiratory fitness, low androgenicity profile from
plasma androgen levels and high plasma leptin levels. Other less pow-
erful predictors included large mean abdominal fat cell size, low post-
prandial EE, high plasma oestrogen levels, poor response of thyroid-
stimulating hormone (TSH) to a thyrotropin-releasing hormone (TRH)
challenge and low plasma cortisol and dehydroepiandrosterone
(DHEA). All of these observations need to be replicated in larger well-
controlled studies, but they nonetheless support the hypothesis that
there is a biology underlying some of the human variability in
response to chronic overfeeding.
5.3 | Fat mass and lean mass changes with
overfeeding
With overfeeding, most of the weight gain is in adipose tissue relative
to the gain in lean mass. The mean weight gain across the 19 experi-
mental groups (see Table 4) reached 4.12 kg whereas fat mass
increased 2.51 kg, which represents 61% of the increase in body mass.
Based on these 19 overfeeding groups, the gain in fat mass across an
OF range from about 20 000 to 84 000 kcal appears to be linear, with
BRAY AND BOUCHARD
one notable outlier (Study 13) (Figure 4, Panel A). Assuming a linear
relation, there is a R2 of 0.89, with a standard error of estimate of
0.46 kg of fat mass. Such a strong relationship is remarkable consider-
ing the unavoidable level of noise in aggregated data, particularly aris-
ing from the fact that body composition was measured by several
different methods and that control over energy intake and activity
level of the subjects was heterogeneous. Excluding the two studies
that fed low-protein diets did not materially influence this relationship
with these 17 study groups having R2 of 0.88 and standard error of
the estimate of 0.47 kg.
The gain in lean tissues is generally less than the increase
observed for fat mass, representing about 30% to 40% of the weight
gain depending on the overfeeding protocol. However, there are over-
feeding studies with higher lean tissue contribution to the weight
gain. Figure 4 (Panel B) depicts the gain in fat-free mass in response
to overfeeding in 19 groups. The increase in fat-free mass is generally
poorly predicted from the magnitude of the caloric overload (R2 of
0.43; SEE = 0.87 kg, with a mean gain of 1.57 kg). Interestingly, the
two low-protein overfeeding groups did not experience substantive
changes in fat-free mass. Removing these two groups from the analy-
sis resulted in a better association between the gain in fat-free mass
and the number of overfed calories (R2 = 0.69; standard error of esti-
mate = 0.56 kg), but the correlation remained lower than the gain in
fat mass. The potential contributions of age, gender and ethnicity to
this variability in body composition changes with sustained overfeed-
ing has not been investigated.
5.4 | Body energy gain with overfeeding
The gains in body energy expressed in kcal (Panel A) and as a percent-
age of the number of overfed calories (Panel B) are depicted in
Figure 5. The changes in body energy were calculated from the gains
in fat mass, using a caloric equivalence of 9300 kcal per kg, and the
gains in fat-free mass based on a caloric value of 1020 kcal per
kg. Data available do not allow for separate estimates of the caloric
content of the gains in fat-free mass versus muscle mass. A close fit
F I G U R E 3 Baseline predictors of the
response to long-term overfeeding. The
paths to high gains are identified from the
correlation studies, as well as the
comparisons of the six highest and six
lowest gainers in response to overfeeding.
The strongest and most consistent
baseline predictors are in the upper part
of the figure, indicated by thick red
arrows, whereas the weaker and less
consistent predictors are grouped in the
lower part of the figure, characterized by
narrow lighter arrows. Reproduced from
Bouchard et al34
BRAY AND BOUCHARD 23

F I G U R E 4 (A) Plot of fat mass gain in

relation to the number of overfed calories in
19 experimental groups retrieved from
13 studies.31,33,44,47,52,58,66,73,81,95,98,163
Groups on a low-protein diet and groups
derived from inpatient studies are contrasted
to others (p < 0.001). (B) Plot of fat-free
mass gain in relation to the number of
overfed calories based on the same studies
as in Panel (A) (p < 0.002). The study
numbers in both panels are defined in
Table 4

was observed between the number of excess calories and the increase
2
in body energy content with R of 0.88, standard error of esti-
mate = 4550 kcal (Panel A). Only one study appears to deviate sub-
stantially from the regression line (Study 13). No differences were
found between the inpatient studies versus the outpatient groups or
between the low-protein diets and the others.
The picture is more complex when one plots the % of the overfed
calories recovered as body energy gain as a function of the number of
overfed calories (Panel B). Here, the relationship is negative but reaches
a low R2 of 0.14 with a standard error of estimate of 15.3%. The associ-
ation with the body energy gain as a % of the overfed calories tends to
be particularly weak when the total caloric overload was in the range
from about 20 000 to 30 000 kcal. Two studies recovered all the excess
caloric intake as increases in body energy, suggesting a perfect effi-
ciency with zero change in any of the components of energy expendi-
ture (Studies 15 and 16). The relationship becomes better at higher
calorie intake but again with the exception of Study 13. The two low-
protein groups fall squarely on the linear regression line.
The graphs for the gains in fat mass, fat-free mass, body energy in
kcal and body energy as a % of the caloric overload based on
17 groups instead of 19 (after removing the two low-protein experi-
mental groups) are available in the Supplementary Material
(Figures S1 to S4).
In summary, on a group basis, there is an excellent fit between
the number of calories overfed and the measured weight gain. The
gains in fat mass and in total body energy content are closely related
to the caloric overload but the increase in fat-free mass is only mod-
erately associated with the number of excess calories. However,
there is on average a threefold differences in weight gain between
the lowest and highest gainers in any given experimental overfeed-
ing study, with a reduced heterogeneity when the protein content
of the overfed diet is increased. The biology underlying human
24
variability in response to experimental overfeeding remains poorly
understood.
6 | METABOLIC RATES AND ENERGY
E X P E N D I T U RE
One of the consequences of chronic overfeeding in humans is that
changes occur in the profile of metabolic fuel oxidized, in metabolic
rates and in whole body total daily energy expenditure which are
reviewed below.
6.1 | Resting metabolic rates
Resting metabolic rate or resting EE is commonly measured using
indirect calorimetry either with the ventilated hood technique or in
a room calorimeter. Resting EE can be measured under variable
BRAY AND BOUCHARD
F I G U R E 5 (A) Plot of body energy gain
in relation to the number of overfed calories
in 19 experimental groups retrieved from
13 studies.31,33,44,47,52,58,66,73,81,95,98,163
Groups on a low-protein diet and groups
derived from inpatient studies are contrasted
to others (p < 0.0001). (B) Plot of estimated
body energy gain as a percentage of overfed
calories based on the same studies as in
Panel (A) (p < 0.111). The study numbers in
both panels are defined in Table 4
resting conditions and this has generated distinctions among
sleeping or basal metabolic rate and resting metabolic rate, the lat-
ter being typically measured during a period of rest in a supine
position when a steady state of quietness has been reached. The
metabolic rate captured under adequate resting conditions reflects
the bioenergetic needs of the organism such as cardiac functions,
breathing, muscle tone, body temperature regulation and tissue
repair. The energy expended at rest represents the largest compo-
nent of total daily EE in people who are sedentary or only rec-
reationally active.
With overfeeding protocols lasting less than 3 days with the
excess caloric overload of the order of a few hundred calories,
there is no consistent indication that excess calories cause an
increase in resting metabolic rate. Some studies lasting 3 and 4 days
have observed significant increases in resting metabolic rate
expressed in absolute units of kcal/min or kcal/day,158 but others
have not.111,140 Increases in resting metabolic rate are not consis-
tently observed when the duration of overfeeding is 7 days and
BRAY AND BOUCHARD 25

more with an overfeeding stimulus that is in excess of 50% of
baseline energy requirements or about 1000 kcal
68,78,81–83,94,95,100,118,164
day. The changes or lack of change in rest-
ing metabolic rate in response to experimental overfeeding were
independent of the carbohydrate or fat content of the overfeeding
protocol.85,154 However, the response of resting metabolic rate
may vary with protein intake, because the study where protein
intake of the caloric overload varied from 5% to 15% and 25%
showed a significant increase in resting metabolic rate the next
day only in the normal and high protein diets.162 Notably, when
exercise was added to a 10-day overfeeding protocol, resting met-
abolic rate increased significantly.78 Interestingly, 3 days of over-
feeding by 50% over weight maintenance increased resting EE per
kg of fat free mass in patients with HIV lipodystrophy but not in
121
HIV-infected or in healthy controls.
The research findings are more consistent with exposure to pro-
longed periods of overfeeding lasting from 1 month to 100 days,
where increases in resting metabolic rate were consistently
observed33,43,45,47,55,58,59,165 with changes of 5% to 12%. In contrast,
low-protein diets did not increase resting metabolic rate compared
with 160 and 227 kcal/day increased with normal- and high-protein
overfed diet.33 This is illustrated in Table 6. Similar increases in RMR
have been observed in individuals with normal weight or obesity and
who were overfed until they gained 10% of their baseline body
weight.55 However, when the same subjects were assessed in a room
calorimeter, a nonsignificant increase of only 97 kcal/day was
observed in subjects who gained 20% of their initial body weight.
Resting metabolic rate increased similarly in adolescents with normal
weight and obesity70 whereas adults with normal weight register
larger increases compared with adults with obesity in response to
82
chronic overfeeding in some studies.
In contrast to data expressing resting metabolic rate in kcal, stud-
ies lasting from 1 week to 100 days have not found an increase in
resting EE when the changes are reported on a per kg body weight or
45,56,58,60,148,164,165
kg fat-free mass basis which suggests that the gain
in resting metabolic rate is essentially associated with the
per overfeeding-induced gain in body mass.
The increase in resting metabolic rate with chronic overfeeding is
correlated with the gain in body weight in protocols lasting about
3 weeks, with coefficients of about 0.4 to 0.6.74,165 The increase in EE
occurs in adipose tissue, muscle and residual nonmuscle, nonfat tis-
sues and is related to protein intake.162
One study has suggested that the calculated metabolic efficiency
of weight gain in response to overfeeding was not related to the
observed changes in resting metabolic rate.81 However, increases in
resting metabolic rate as a result of chronic exposure to overfeeding
vary among individuals by as much as twofold to threefold,68,94 con-
sistent with the heterogeneity of the changes observed in
weight gain.
6.2 | Thermic effect of food
The thermic effect of food or diet-induced thermogenesis is typically
measured by indirect calorimetry for 4 h and more following the con-
sumption of a standardized meal under resting conditions in the
supine or semi-reclined position. Thermic effect of food is influenced
by the nutrient composition of the food consumed but is variable
even when testing conditions are highly standardized. In a short-term
study, carbohydrate overfeeding for 4 days was shown to increase
thermic effect of food by as much as 39% in 10 young, normal weight
148
subjects.140 In contrast, no effect of overfeeding over 3 days was
observed on thermic effect of food in patients with HIV-lipo-
dystrophy, or those with HIV but no lipodystrophy as well as healthy
controls.121 Notably, a glycogen-depleting exercise bout the day prior
to the measurement of thermic effect of food following 4 days of car-
bohydrate overfeeding caused a 23% increase in the thermogenic
response.140
A number of overfeeding protocols lasting 7 to 14 days failed to
find significant changes in the thermic effect of food.70,78,110 One

TABLE 6 Changes in the components of energy balance following 8 weeks of overfeeding in relation to the protein content of the diet

Protein diet group, mean (95% CI)

Low: 5% Normal 15% High 25% p value*

Age, year 22.9 22.9 26.8 0.08
N 8 9 8
Weight, kg 3.16 (1.88 to 4.44) 6.05 (4.84 to 7.26) 6.51 (5.23 to 7.79) 0.002
Energy expenditure, kcal/day
Total 42 (−177 to 262) 522 (316 to 729) 454 (233 to 672) 0.007
Resting −20.6 (−82 to 41) 160 (102 to 218) 227 (165 to 288) <0.001
Physical activity level† 0.021 (−0.13 to 0.18) 0.16 (0.019 to 0.31) 0.079 (−0.078 to 0.23) 0.38
Nonresting energy expenditure, kcal/day 59 (−160 to 277) 310 (104 to 516) 181 (−37 to 399) 0.24
Excess energy intake, kcal/day 924 (SD = 50) 977 (50) 904 (53) 0.13
33
Note. Adapted from Bray et al.
*
Calculated using analysis of variance.
†
Calculated as TEE divided by resting energy expenditure.
26
study did not find any interaction between the thermic effect of food
and exercise in response to 7 days of overfeeding by 50%.110 In
another study, thermic effect of food increased slightly with 9 days of
overfeeding, but it remained at about 10% of the energy intake when
measurements were extended to 16 h in a room calorimeter.95 Over-
feeding with medium chain triglycerides for 1 week significantly
increased the thermic effect of food by 12% whereas a similar over-
feeding protocol with long chain triglycerides increased it by a nonsig-
nificant 7%.118 In 12 young men subjected to 22 days of overfeeding
by 1000 kcal/day, thermic effect of food increased by about 6%, with
94
a range from 4% to 21%.
In studies lasting from 1 month to 100 days, the changes in ther-
mic effect of food were typically small, but they often reached statisti-
cal significance. When 24 young, lean men were exposed to a caloric
surplus of 84 000 kcal over 100 days of overfeeding, thermic effect of
food was unchanged, amounting to 65 kcal over 4 h before overfeed-
ing and 68 kcal in the post-overfed state.165 Leibel and co-workers
observed a slight increase in the thermic response to a liquid meal
with a caloric content specified as 60% of resting EE measured the
55
same morning. The thermic effect of food reached 3% of the calo-
ries ingested at the initial weight in 13 subjects without obesity but
5% after they had gained 10% of their body weight, whereas the
changes in the thermic effect of food went from 2% to 4% in 11 sub-
jects without obesity, differences that were reported to be significant.
Under similar experimental conditions in 23 patients (14 males), the
increase in thermic effect of food following a 10% gain of body weight
148
reached 36 kcal per day. In 16 normal weight subjects overfed by
1000 kcal/day for 8 weeks, the thermic response to a meal challenge
of 200 kcal remained the same at 11 kcal before and 12 kcal post-
overfeeding.56 However, in nine men overfed by an average of
228 000 kcal over 65 days (3508 kcal/day) as part of the Gulu Walla
ritual, the thermic response to a meal from 60 to 100 min after the
meal increased from 1.5 kcal to 1.9 kcal/min with the massive over-
feeding, a mean increase of 26% with a range from 7% to 48%.59
Unfortunately, the test meal could not be fully standardized in the lat-
ter study; thus, some of these large increases could be explained by
the variable caloric content of the test meal. The thermic effect of
breakfast was increased by exercise but not by the size of the break-
46
fast in one overfeeding experiment. In another study, weight gain
did not increase thermic effect of food even in the presence of pro-
longed hypercaloric diets with variable protein content,33 but thermic
effect of food was increased in response to a meal test matching the
166
low-, normal- or high-protein content of the overfeeding protocols.
Globally, the data suggest that if there is an increase in thermic
effect of food with overfeeding, although it is rather small and gener-
ally not statistically significant. It is often not clear whether the slight
increase in thermic effect of food is mainly the result of the slight gain
in resting metabolic rate. Thus, the change in the thermic effect of
food often became nonsignificant when resting EE over the same
period was taken into account. Notably, the protein content of the
test meal influences the magnitude of thermic effect of food with
low-protein content associated with a lower thermic effect of food
response.
BRAY AND BOUCHARD
6.3 | Exercise energy expenditure
An important question is whether exercise influences the response to
experimental overfeeding. Two questions were examined: (1) does
overfeeding increase sedentary time and reduce EE associated with
fidgeting and spontaneous physical activity, and (2) is there a change
in energy cost at a given external workload with overfeeding.
A few reports have generated data bearing on the first ques-
tion. In one study, sedentary time estimated with accelerometers
during 3 days of overfeeding was positively associated with weight
gain (r = 0.49) adjusted for age, sex and initial weight.117 In
another report, 8 days of overfeeding increased nonresting EE by
316 kcal per day.83 Levine et al56 studied 16 normal weight sub-
jects overfed by 1000 kcal per day for 8 weeks and observed that
nonresting EE (called NEAT by the authors) increased by a nonsig-
nificant 328 kcal/day (SD = 256). This nonresting EE included
changes associated with fidgeting, posture and activities of daily
life and was significantly associated with resistance to gaining body
fat (r = 0.77; p < 0.001).56 This observation is interesting especially
because the changes in resting metabolic rate and thermic effect
of food were not associated with the gains in adiposity. However,
an important limitation of the study is that subjects were enrolled
as outpatients throughout the experimental overfeeding protocol.56
In contrast, in an inpatient setting, when three groups of subjects
were overfed for 56 days with diets that varied in protein content,
the change in nonresting EE was nonsignificant and was identical
among the three groups, as well as being more modest than the
changed reported by Levine and colleagues.33
The study of overfeeding with different levels of dietary protein33
also showed effects of overfeeding on physical activity measured by
accelerometry.167 Change in activity, using the vector magnitude from
the accelerometer which is independent of weight, and activity energy
expenditure were positively correlated with weight gain, but these
changes in activity were not affected by the level of protein in the
diet. Thus, overfeeding seems to produce an increase in physical activ-
ity and in energy expended in physical activity after adjusting for
changes in body composition, suggesting that increased activity in
response to weight gain might be one mechanism to support adaptive
thermogenesis.167
Several studies are available to address the second question of
whether there is a change in energy cost at a given external work-
load with overfeeding. One study reported that 15 days of over-
feeding by 1500 kcal/day increased the absolute energy cost of
cycling, walking and climbing stairs by 19% to 29% as assessed by
the measurement of oxygen uptake during exercise.68 In contrast,
18 days of overfeeding by 1000 kcal per day did not increase the
energy cost of exercise in subjects who were lean or obese,82 nor
did overfeeding with weight gain affect the energetic efficiency of
muscle measured on a cycle ergometer at four incremental
workloads.168
In longer overfeeding protocols, the energy cost of various
postures and activities increased although there were occasional
negative results. For instance, in nine men overfed by 50% for
BRAY AND BOUCHARD
42 days, the energy cost of stepping and cycling did not change.47
When six underweight males were overfed by 720 kcal/day for
8 weeks, there was an increase in the energy cost of sitting and
standing but no change in the cost of walking, shoveling or push-
ing an empty or a loaded wheel barrow, all expressed as
kcal/min.60 In contrast, Norgan and Durnin58 found that the energy
cost in kcal/min of lying down increased by 12%, sitting by 11%,
walking at 4 km/h by 9% and at 5.6 km/h by 12%. However, none
of these changes were significant when they were expressed in
kcal/min per kg body weight.58 Similarly, in 24 young men overfed
by 84 000 kcal over 100 days, the energy cost of walking at three
different speeds increased by 7.1%, 10.6% and 11.3% when data
were expressed in kcal/min but was unchanged when reported per
kg of body weight.165 In the massive overfeeding ritual of the
Guru Walla, the excess consumption of 228 000 kcal over 65 days
resulted in an increase of estimated activity metabolic rate from
59
4.7 to 5.6 kcal/min.
In conclusion, whether exercise EE is influenced by chronic
overfeeding seems to be dependent on two main factors: whether
subjects were enrolled in the overfeeding protocol as inpatients or
outpatients and how the EE data are reported. Non-resting EE
seems to be generally higher in protocols in which the subjects
were enrolled as free living outpatients. Overfeeding-induced
changes in nonresting EE or in the energy cost of specific postures
or activities tend to be significant when expressed in absolute unit
(e.g., kcal/min) but not on a per kg weight of kg fat-free mass
basis reflecting that the change in energy costs results mainly from
the gain in body mass.
6.4 | Total daily energy expenditure
Total daily EE during overfeeding has been evaluated in free-living
participants and in others confined to a metabolic ward using
either 24-h calorimetric measurements or the doubly labelled water
method.
Short-term overfeeding lasting 1 to 4 days marginally increases
total daily EE.111,114,141,142,144,155,158 It has been shown that carbohy-
drate overfeeding increases total daily EE more than overfeeding with
111
dietary fat, but the latter increases EE in individuals who were
obese or of normal weight.144 Of interest is the observation that low-
protein overfeeding increases total daily EE less than a high-protein
33,158,159
caloric overload. One study has reported that total daily EE
was increased to about the same extent by overfeeding or mild cold
exposure for 3.5 days.142
In studies lasting 1 to 3 weeks, total daily EE did not increase in
78,81,96 70,79,83,95
some studies but it did in others. In two overfeeding
reports with 8- and 9-day protocols, 25% of the excess caloric intake
was dissipated as increased total daily EE.83,95 Similar increases in
total daily EE have been reported in obese and nonobese adolescents
70
exposed to 2 weeks of carbohydrate overfeeding. Two weeks of
carbohydrate overfeeding increased total daily EE in subjects who
were lean or obese whereas fat overfeeding did not.31
27
In longer overfeeding protocols extending from 1 month to
100 days, most studies report an increase in total daily EE in kcal/day,
but the data are less consistent when expressed per unit of body
weight. Three studies did not observe significant increases in total
daily EE.58–60 In contrast, Leibel and collaborators found an increase
of total daily EE in 24 subjects who were weight stable after gaining
10% of their initial body weight.55 Figure S5 depicts the changes in
total daily EE in experimental subjects who were confined to a meta-
bolic ward for long periods of time. The change in EE from baseline
was computed from a model in which predicted EE based on the
regression of fat-free mass and fat mass was subtracted from the
observed EE. Subjects were measured at their initial weight, after
gaining 10% of their weight, again after a 10% weight loss and, for
some subjects, after 20% weight loss. The overfeeding-induced 10%
gain of body weight was accompanied by a robust increase in total
daily EE of about 500 kcal/day.55 Notably, reductions of body weight
by approximately 10% or 20% decreased total daily EE by only about
300 kcal/day.
In the same experimental framework, and based largely on the
same subjects, changes in total daily EE were detected when the
assessment was based either on caloric titration (Diet total daily
EE) or on doubly labelled water assessment of TDEE (Isotope total
daily EE) but not when the measurement was derived using room
calorimeters.148 After a gain of body weight of 10%, total daily EE
increased by 593 kcal/day with the Diet total daily EE method,
328 kcal/day by the Isotope total daily EE method but only by
43 kcal in the confines of the room calorimeter. The latter has
been attributed to the lower spontaneous physical activity level in
the calorimeter.
However, another overfeeding protocol lasting 42 days reported
a mean calorimeter-assessed increase in total daily EE of about
430 kcal/day but no change was observed when it was evaluated by
the doubly labelled water method.47 In 16 volunteers who gained
4.7 kg of body weight over a period of 8 weeks, total daily EE
increased from 2807 to 3361 kcal/day, a mean increase of
554 kcal/day.56 In one overfeeding experiment lasting 100 days con-
ducted with 12 pairs of identical twins, the estimated total energy
cost of weight maintenance increased from 2756 to 3001 kcal/day,
an increase of 5.5% in the presence of a weight gain of 8.1 kg.44,165
Interestingly, when volunteers were overfed in an inpatient setting for
56 days, those on a low-protein diet (5% of calories) did not experi-
ence an increase in total daily EE from doubly labelled water.33 In con-
trast, total daily EE increased by 522 kcal/day in the normal-protein
diet (15% of calories) group and by 454 kcal/day in the high-protein
diet (25% of calories) subjects as shown in Table 6. Interestingly, the
dose response of total daily EE in relation to the protein content of
the overfeeding protocol extends to the observed changes in resting
EE, with a range from no change for the low-protein diet to an
increase of 227 kcal/day for the high-protein group, but not to non-
resting EE or physical activity level for which changes were not
significant.
Although the topic has not been addressed systematically in the
studies reviewed herein, the ratio of total daily EE to basal metabolic
28
rate reached 1.8 and, in one report, was found to be identical before
and after an overfeeding protocol.47
6.5 | Respiratory exchange ratio
The respiratory exchange ratio or respiratory quotient (RQ) is used to
assess whole body oxidation rates of carbohydrate and lipid sub-
strates under resting, postprandial or active but standardized condi-
tions. Short-duration overfeeding protocols have generated mixed
findings. For instance, 3 days of overfeeding resulted in an increase in
carbohydrate oxidation over 24 h in a metabolic chamber in both obe-
sity prone and obesity resistant subjects.155 In the same study, fat oxi-
dation decreased and protein oxidation (estimated from urinary urea
nitrogen excretion) increased in both groups. Carbohydrate overfeed-
ing increased carbohydrate oxidation during resting conditions com-
pared with baseline measurement (RQ = 0.83 to 0.91) as it did in
response to a meal challenge as measured by postprandial substrate
140
oxidation rates. In the latter case, RQ increased from 0.87 to 0.94
when the thermic effect of food reached 12% of the caloric content
of the ingested calories.
When the resting RQ is assessed in overfeeding protocols of lon-
ger durations of up to 3 weeks, an increase in RQ is typically
observed, reflecting a growing reliance on carbohydrate oxida-
tion.31,79,85,96,154,169 The increase in carbohydrate oxidation is
observed during resting or sleeping conditions in a calorimeter and is
most prominent following exposure to carbohydrate overfeeding. In
contrast, chronic overfeeding with dietary fat has negligible effects on
RQ.31,85 The rate of protein oxidation declines or is unchanged with
dietary fat or carbohydrate overfeeding.31,79 In a one-week study,
medium or long chain triglyceride overfeeding resulted in a lower RQ
and thus an increase in lipid oxidation.118 In contrast, carbohydrate
overfeeding for 17 days did not enhance carbohydrate oxidation in
response to a meal challenge.169 In fact, carbohydrate overfeeding
was accompanied by a nonprotein RQ greater than 1.0 suggesting
lipogenesis from glucose in the fasted state. The response to a meal
test did not stimulate net lipogenesis from glucose following the over-
feeding protocol.169
In longer overfeeding protocols, lasting 1 month and more,
targeting a 5% increase in body weight, did not change RQ under rest-
ing conditions45 but a 10% gain was associated with a decrease in RQ
from 0.92 to 0.86 (see Leibel et al55). In a study in which nine men
were overfed by 50% of their baseline energy requirements for 42
days, the RQ derived from room calorimeter measurement reflected
an increase in carbohydrate oxidation from 316 to 491 g/day whereas
fat oxidation decreased from 126 to 71 g/day in spite of an increase
47
in dietary fat intake from 131 to 204 g/day. Similarly, in eight men
who were exposed to a daily caloric surplus of 550 kcal/day for
28 days, the RQ associated with the resting metabolic rate measure-
ment increased from 0.84 to 0.88 (see Meugnier et al88).
Long-term exposure to overfeeding with a balanced macronutri-
ent composition (15% protein, 35% lipid, 50% carbohydrate) seems to
attenuate and even prevent the major fluctuations in the respiratory
BRAY AND BOUCHARD
exchange ratio observed in short term studies. This was evidenced in
the 24 young men who were overfed by 84 000 kcal over a 100-day
period (C. Bouchard and A. Tremblay, unpublished data). Thus, no sig-
nificant changes in RQ were observed when assessed during resting
metabolic rate, thermic effect of food in sitting or standing positions
or when walking on the treadmill under steady state conditions at 4.5,
5.5 or 6.5 km/h. Thus, it is critical to distinguish between early and
long-term adaptation to overfeeding and the macronutrient composi-
tion of the diet both of which can make a substantial difference
on RQ.
In summary, changes in resting metabolic rate have been consis-
tently observed with long-term overfeeding, particularly when the
metabolic rate measurements were performed a day or two following
the end of the overfeeding treatment. However, when resting EE data
are adjusted for body weight or fat-free mass, the overfeeding-
induced changes are generally nonsignificant. There is a slight trend
for an increase in thermic effect of food with overfeeding, but the net
effect is small and often not statistically significant. The protein con-
tent of the test meal influences the magnitude of thermic effect of
food. The energy cost of physical activity and nonresting EE may
increase with experimental overfeeding but only in proportion to the
weight gained. In long-term overfeeding studies, there is strong evi-
dence that total daily EE is increased although it may not always be
detected in the confine of a room calorimeter. In overfeeding studies
lasting from a few days to about 1 month, the changes in RQ suggest
a growing reliance on carbohydrate oxidation. This trend is attenuated
with longer periods of overfeeding.
7 | E F F E C T S OF D I E T A N D D I E T A R Y
C O M P O S I T I O N ON TH E R E S P O N S E T O
O V E RF E E D I N G
Experimental overfeeding can be done using excess energy from bal-
anced diets, as well as changing the amount of fat, carbohydrate or
protein independently. In addition, the type of fat or carbohydrate
may also make a difference in the response.
7.1 | Overfeeding with high-carbohydrate or high-
fat diets
High-fat and high-carbohydrate diets have been compared in a
number of studies31,66,67,69,71,85,102,111,113,120,124,158,159,170–173 with
several being conducted with participants living in a research
facility,31,159,174,175 but in most studies, participants lived in outpa-
tient settings where meals were provided or where individuals were
asked to follow recommended dietary plans. The study by Adochio
et al102 illustrates the effect of short-term overfeeding and included
11 men and 10 women who ate both high-carbohydrate and high-fat
diets for 5 days each. No sex differences in response were identified.
The high-carbohydrate diet increased plasma triglycerides and insulin
but lowered plasma free fatty acids (FFA). In contrast, the high-fat diet
lowered triglycerides but had no effect on either plasma FFA or
BRAY AND BOUCHARD
plasma insulin. Neither the high-carbohydrate nor high-fat diet chan-
ged plasma glucose, glycerol, adiponectin, leptin, tumour necrosis
102
factor-alpha (TNFA) or interleukin-6 (IL6). After 5 days of overfeed-
ing, whole body insulin sensitivity estimated from the M-Value (mg/kg
fat-free mass/min), the glucose disposal rate, the metabolic clearance
rate or the resting hepatic glucose output did not change with either
diet. In contrast to the lack of whole body changes in insulin
sensitivity, there were changes in insulin response at the cellular level.
Skeletal muscle insulin signalling through tyrosine phosphorylation of
insulin receptor substrate-1 (IRS1) was increased as was the
IRS1-associated phosphatidylinositol 3-kinase (PI3K) activity following
the high-carbohydrate diet. In contrast, overfeeding with the high-fat
diet increased skeletal muscle serine phosphorylation of IRS1.
Lower plasma FFAs after overfeeding a high-carbohydrate diet or
a balanced diet were observed in many studies of
overfeeding.43,63,73,82,88,95,104,107,111,112,130,139,142,154,176–188 How-
ever, not all studies reported a reduction of FFA after
overfeeding.49,52,76,77,87,97,111,121,143,189–197 These differences in
response may reflect, in part, differences in the duration of overfeed-
ing because Samocha-Bonet et al98,198,199 reported that FFA were
suppressed at 3 days but not after 28 days of overfeeding with a 46%
fat diet. An increase in plasma FFA, triglycerides and hepatic fat was
found after 3 days of overfeeding with a high-fat/high-energy diet in
15 healthy men.200 This temporal difference is consistent with the
suppression of nocturnal FFA during the first day of overfeeding.125
7.2 | Comparison of candy and nuts as an alternative
way to overfeed sugar and fat
A second approach to testing the effect of high-carbohydrate and
high-fat diets used snacks with candy for the high-carbohydrate diet
and nuts for the high-fat diet to overfeed 11 men and 14 women by
20 kcal/kg body weight. Body weight increased significantly by 0.8 kg
in the group eating candy, but the 0.3 kg increase in the group eating
the peanuts was not significant.72 The main finding in this study was
that overeating sugar in the candy diet produced a more unfavourable
change in lipids which were independent of changes in weight.
Palmitic acid, oleic acid and linoleic acid all increased significantly in
the peanut diet, with no change in the candy diet. Apolipoprotein B
rose in the high-carbohydrate group eating candy but fell slightly in
the peanut group producing a significant difference in response
between the groups.
7.3 | Effect of types of dietary fat on the response
to overfeeding
7.3.1 | Polyunsaturated fatty acids compared with
saturated fatty acids
The quality as well as the quantity of dietary fat may influence the
response to overfeeding. To test diet quality, Rosqvist et al62 overfed
29
39 healthy men and women for 7 weeks using diets in which muffins
were enriched with either sunflower oil as a rich source of linoleic
acid, a polyunsaturated fatty acid or palm oil, which is rich in palmitic
acid, a saturated fatty acid. The intake of muffins, which were 51%
energy as fat, was adjusted to assure a 3% weight gain over 7 weeks.62
Both groups had a weight gain of 2.2%. Despite comparable weight
gains, consumption of the polyunsaturated fatty acid supplemented
diet prevented the deposition of liver fat and visceral adipose tissue
compared with the saturated fatty acid supplemented diet. Con-
versely, polyunsaturated fatty acids were associated with a nearly
threefold greater increase in lean tissue than the group overeating sat-
urated fatty acids from palm oil. The increase in liver fat was directly
correlated to the changes in plasma saturated fatty acids and inversely
with polyunsaturated fatty acids. A number of genes involved in regu-
lating energy dissipation, insulin resistance, body composition and fat-
cell differentiation in subcutaneous adipose tissue were differentially
regulated between diets and associated with increased polyunsatu-
rated fatty acids in subcutaneous fat.62 Total cholesterol, high-density
lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) choles-
terol and apolipoprotein B:apolipoprotein A1 ratios decreased signifi-
cantly in the individuals eating muffins enriched with polyunsaturated
fats compared with the palm oil enriched muffins, but other car-
diometabolic risk markers were unchanged. Independent of fat intake,
there were increases in plasma proinsulin (+21%), insulin (+17%), pro-
protein convertase subtilisin/kexin type 9 (PCSK9) (+9%), fibroblast
growth factor 21 (FGF21) (+31%), endothelial markers vascular cell
adhesion molecule 1 (VCAM1), intercellular adhesion molecule
1 (ICAM1) and E-selectin (+9%, +5% and +10%, respectively).201
7.3.2 | Effects of overfeeding with medium chain
triglycerides compared with long chain triglycerides
The fatty acids released from medium chain fatty acids are
absorbed as such and transported directly to the liver. In contrast,
long chain fatty acids are converted to triglycerides in the gastroin-
testinal tract and packaged into chylomicrons before entering the
lymphatic system and then the general circulation. To test whether
this difference in routes of absorption would affect thermogenesis,
Hill and colleagues studied 10 healthy young men ages 22–44 who
were overfed liquid formula diets containing 40% fat as either
medium-chain or long-chain triglycerides.118 Each man was studied
for 1 week on each diet using a double-blind, crossover design.
Although resting metabolic rate did not change during either week
of 50% overfeeding, the thermic effect of food was greater on
Day 1 and was 12% greater following a 1000 kcal meal containing
medium chain fatty acids than following an isocaloric meal con-
taining long chain fatty acids. Energy expenditure during a 20-h
continuous enteral infusion was significantly higher with the
medium chain fatty acid than long chain fatty acid diet on Day
7 (15.7 ± 1.7% vs. 7.3 ± 0.9% of ingested energy). The hypothesis
that the increased EE was the result of increased lipogenesis in
the liver during the medium chain fatty acid diet was supported by
30
a reduction in fasting serum total cholesterol concentrations with
long chain fatty acids but not with medium chain fatty acids and a
threefold increase in fasting serum triglycerides concentrations with
medium chain fatty acid but not long chain fatty acid diet.202
7.4 | Effects of different types of sugars on the
response to overfeeding
7.4.1 | Effect of glycaemic index
The glycaemic index is a measure of the rise in blood glucose fol-
lowing a fixed dose of food containing readily digestible carbohy-
drate. This is determined by the rate of cleavage of the
carbohydrate into simple molecules like glucose and fructose in the
gastrointestinal tract and can be modified by the presence of other
foods. Overfeeding might modify the response to diets with differ-
ent glycaemic loads based on adaptive change in gastrointestinal
tract with overfeeding.
Significant changes in glucose metabolism are evident within
7 days of controlled overfeeding or underfeeding.182 During refeeding
after an initial period of overfeeding followed by underfeeding, partici-
pants assigned to a 65% carbohydrate diet during the second over-
feeding period gained more body weight compared with a 50%
carbohydrate diet particularly when high glycaemic index foods were
incorporated into the meals.182 After refeeding, serum triglycerides
and liver fat were elevated above baseline only with the 65% carbohy-
203
drate diet. Refeeding a high-glycaemic index diet led to impaired
basal fat oxidation when compared with the low-glycaemic index
diet.204 During the second overfeeding period, insulin sensitivity
decreased, an effect that was associated with increases in adiponectin,
free triiodothyronine (T3), thyrotropin stimulating hormone (TSH) and
glycaemic load—all of which influenced the response to the diet. In
contrast, ghrelin levels decreased.205 Endotoxin levels significantly
increased by 30.8% during the 7-day overfeeding period and again by
24.7% during the subsequent overfeeding after caloric restriction, but
this was not influenced by the glycaemic content of the diet.206
7.4.2 | Effect of overfeeding dietary fructose
Fructose is a 6-carbon hexose which is found in many fruits and some
vegetables and comprises 50% of the sucrose (sugar) molecule. The
major source of fructose in most diets is from sucrose which is a dimer
of fructose and glucose that is widely used in food preparation and in
sugar-sweetened beverages, energy drinks and fruit drinks. The sweet
taste of these products is primarily due to fructose. Fructose alone is
poorly absorbed from the gastrointestinal tract, but in the presence of
glucose absorption, absorption increases sharply. The fructose that is
absorbed into the portal blood is almost completely cleared in the first
pass through the liver. Thus, fructose has a “low glycaemic index” (see
above) because it does not appear in the circulation as glucose and
thus does not stimulate insulin.
BRAY AND BOUCHARD
Within the liver, fructose is converted to fructose-1-phosphate
which is readily cleaved to make glycerol-3-posphate, the backbone
of triglycerides. It is for this reason that fructose can stimulate triglyc-
eride formation.207 Overfeeding of fructose has been examined in
several studies112,123,126,135,154,183,194,208–211 and compared with glu-
cose212 or fish oil.208 McDevitt et al126 recruited eight lean and five
obese women to spend 96 h living continuously in a whole-body calo-
rimeter to assess energy dissipation as well as glycogen and fat stor-
age. Five diets were provided in random order: a control diet at
energy balance levels (control) and diets with predominantly fat, glu-
cose, fructose or sucrose which were overfed by 50% of energy
requirements. Increased energy expenditure after overfeeding dissi-
pated 7.9% of the energy excess, but there were no significant varia-
tions related to the diets fed. Energy expenditure during overfeeding
with fat significantly exceeded that during the control treatment in
the lean but not in the obese women. There were no differences in fat
balance during controlled overfeeding with either fat, fructose, glu-
cose or sucrose.126 There were also no differences between lean and
obese women in macronutrient oxidation or energy balances for indi-
vidual carbohydrates.
The effects of overfeeding fructose on hepatic lipid and plasma
triglycerides have been reported in several studies.123,135,154,210,213
Fructose at doses above 1.5 g/kg/day had significant effects on
hepatic lipids and serum triglycerides, but glucose at comparable
doses also produced effects on hepatic lipids indicating that both can
be detrimental at higher doses.
Adding essential amino acids to the diet194 or coffee214 both
reduced the detrimental effect of fructose on lipid metabolism during
overfeeding. In contrast, supplementation with leucine, isoleucine,
valine, lysine and threonine which blunted the effects of fructose on
intrahepatocellular lipid levels did not affect the induction of hyper-
triglyceridemia.194 Coffee, which contains chlorogenic acids, also sig-
nificantly decreased hepatic glucose production, but it did not
significantly blunt the rise in intrahepatocellular lipids.214
Fructose and starch as whole wheat have been compared in
17 individuals who received four treatments each lasting 24 h. One
was an energy balanced diet; the second was a 24-h fast; the third
and fourth were overfeeding at 200% with 5% fat diets that contained
75% of carbohydrate either as high fructose corn syrup (HFCS) or
whole wheat.209 Both types of carbohydrate—high fructose corn
syrup and whole wheat—increased 24-h EE by 12.8% ± 6.9%. Inter-
estingly, sleeping metabolic rate was significantly higher after the high
fructose corn syrup diet (+ 35 ± 48 kcal), but not the whole wheat
diet. Overfeeding high fructose corn syrup also increased urinary cor-
tisol concentrations more than overfeeding whole wheat
(107.6 ± 46.9 nmol/24 h vs. 82.8 ± 35.9 nmol/24 h).209
7.5 | Comparison of dietary protein on the response
to overfeeding
Interest in dietary protein and the response to overfeeding was kin-
dled by Miller and Mumford (1967)57 who reported less weight gain
BRAY AND BOUCHARD
when overeating a low-protein diet.57 Subject DM gained 1.3 kg as
opposed to the expected 6.2 kg when overeating 37 300 kcal and
subject PM only gained 1.2 kg when overeating 20 050 kcal as
opposed to an anticipated 3.3 kg based on the energy values of the
foods they were overeating. Stock19 found a “V”-shaped relationship
between protein intake and feed efficiency of weight gain when plot-
ting the energy cost of weight gain (in kcal per kg) against percent pro-
tein in the diet. The nadir in a group of 12 studies reviewed by
Stock19 was at 12% with a higher energy cost of weight gain when
eating either lower or higher protein diets. This review prompted the
Protein Overfeeding Study (PROOF) in which 25 healthy men and
women living in an in-patient facility were overfed with diets con-
taining either 5%, 15% or 25% protein for 56 days.33 Figure 6 shows
that less weight was gained when eating the low protein diet, but that
this was a result of less increase in fat free mass, but not in fat stor-
age. The gain in total body fat was directly related to the increase in
energy intake; however, individuals eating the low-protein diet stored
a higher percentage of the extra energy in fat and showed an increase
162,177
in percent fat compared with the higher-protein group. Fat-free
mass and intrahepatic lipid increased more with the high-protein diet,
whereas percentage body fat and fasting FFAs increased more on the
low-protein diet. The thermic effect of food was related to acute
intake of dietary protein but was not altered by prolonged intake of
high-energy diets with either high or low protein content.166
F I G U R E 6 Effect of 8 weeks of overfeeding three levels of dietary protein on changes in body weight, body fat and lean body mass.
Reproduced from Bray et al33
31
Metabolomic analysis of blood samples from this 8-week protein
overfeeding study (PROOF) identified three patterns of change in
plasma amino acids.215 One showed increasing concentration of five
essential and three nonessential amino acids with increasing protein
intake, as one might expect from the increased protein level. The sec-
ond pattern had higher levels of six nonessential amino acids with the
low-protein diet and finally a third pattern where the relationship was
flat or “V”-shaped. Dietary fat and protein were both correlated with
changes in valine, phenylalanine, leucine/isoleucine/norleucine and
tyrosine, but energy intake was not. The changes in fat mass and
weight were also related to the change in several amino acids. Base-
line fat cell size and the interaction between glucose disposal and fat
cell size were associated with changes in several amino acids during
overfeeding.216
Metabolomic analysis of fatty acyl carnitines found that higher
protein intake was associated with significantly lower 8-week levels
of medium chain fatty acids and C2, C4-OH and C6:1 but higher
values of C3 and C5:1 acyl-carnitines derived from essential amino
acid.216 In contrast, energy and fat intake were only weakly related to
changes in fatty acyl-carnitines. A decease or smaller rise in 8-week
medium chain acyl-carnitines was associated with an increase in
sleeping EE and fat-free mass and a decrease in FFA concentrations.
In contrast, changes in short-chain fatty acyl-carnitines were related
to changes in resting EE and fat-free mass, and C4-OH was positively
32
related to FFA. The low-protein diet was associated with higher
values of most carnitine acyl-fatty acids. The association of dietary
protein and fat intake may explain the changes in EE and metabolic
variables resulting in the observed patterns of fatty acyl carnitines.216
Two studies have examined the effect of a low-protein diet on
change in EE in the first 24 h of overfeeding.173 Schlogl et al173 over-
fed individuals by 200% for 1 day and reported that the low-protein
diet increased EE by 2.7% in contrast to a 10.9% rise when protein
content was 20%. The effects reported by Bray, Redman et al177 who
overfed by 40% differed somewhat from Schlogl et al,173 in that nei-
ther total daily EE nor sleeping EE increased in the metabolic chamber
in the first 24 h when overeating the 5% protein diet. However, both
24-h EE and sleeping EE increased in those eating the normal or
higher protein diet. Over the 8 weeks of the study, the 24-h EE in the
metabolic chamber was correlated with protein intake (r = 0.60) but
not energy intake in all three groups.162
7.6 | Effect of frequency of food intake
The effect of frequency and timing of meals on the response to over-
feeding has been examined in two studies.71,190 In the first study, six
patients with obesity were fed 5000-calorie diets for 4 weeks.71 Dur-
ing one period of 2 weeks, the calories were consumed over 4 h (gorg-
ing) and during the other 2 weeks, the dietary intake was spread over
20 h (nibbling). Each of these periods followed a low caloric intake
which lasted at least 10 days. Three male patients (group I) were stud-
ied at or near their maximal weight and three females (group II) after a
weight loss of 50–70 kg. The patients in group II gained more weight
than those in group I. Lipogenesis from pyruvate was greater in group
II than in group I. Gorging was accompanied by a significant increase
in lipogenesis in adipocytes. These data suggest that the frequency of
food intake may be inversely related to obesity.
To test further eating frequent meals versus large meals Koopman
190
et al randomized 36 lean, healthy men who overate by 40% for
6 weeks using a 2 × 2 randomization with an additional control group
that did not overeat. Regular meals in two groups were supplemented
with either high-fat/high-sugar foods or high-sugar foods only (large
meals). The other two groups ate the same meals, but their supplements
were given between meals (frequent meals). At the end of 6 weeks,
weight had increased by about 3% in all men. Increasing meal frequency
during overfeeding increased liver fat by 45% in the high-fat/high-sugar
food group and 110% in the high sugar group whereas increasing meal
size did not. Abdominal fat increased in the high-fat/high-sugar group
eating frequent meals and tended to increase in the high sugar frequent
meal group as well. Hepatic insulin sensitivity tended to decrease in the
high-fat/high-sugar frequent meal group, whereas peripheral insulin
sensitivity was not affected. The authors conclude that overeating with
more frequent meals increases intrahepatic triglycerides and abdominal
fat independent of caloric content and gain in body weight, whereas
increasing meal size does not. This study suggests that snacking, a com-
mon feature in the Western diet, independently contributes to hepatic
steatosis and obesity.190 In the same study, the binding of serotonin to
BRAY AND BOUCHARD
receptors in the central nervous system was examined by single-photon
emission computerized tomography. Although there were no differ-
ences in total calories consumed between the four diet groups, the
group overfed the high-fat/high-sugar snacking diet significantly
decreased serotonin transporter (SERT)-binding by 30%. More work is
needed to understand this difference.54
We conclude that although diet does not play a role in the
amount of fat stored—only excess energy does that—diet does play a
role in the distribution of body fat. Effects of the duration of overfeed-
ing were evident in several studies suggesting that the mechanisms
called into play initially may be replaced by other response mecha-
nisms with continued overfeeding. Low levels of dietary protein
reduced weight gain, but not fat storage. Specific nutrients can also be
singled out including the quantity of unsaturated fatty acids compared
with polyunsaturated fatty acids which is an area that merits further
investigation. Finally, the pattern of eating whether frequently or
infrequently produced differences that warrant additional study.
8 | M E D I A T O R S OF TH E R E S P O N S E T O
O V E RF E E D I N G
By mediators, we refer to baseline factors such as age, sex, ethnicity,
birth weight, family history of diabetes and other traits that may influ-
ence the response to overfeeding. Genetic factors are addressed in a
separate section.
8.1 | Response to overfeeding in older and younger
individuals
Energy requirements decline with age; thus, age may modify energy reg-
ulation in response to overfeeding. Only one study has addressed this
question directly and then only in men. Roberts et al217 compared the
effects of overfeeding on changes in EE, substrate oxidation and energy
deposition between younger men, aged 23.7 (standard error of 1.1 years),
and older men, aged 70.0 (± 7.0), who were of normal body weight. They
were overfed by 1000 kcal/day for 21 days. There were no differences
in weight gain or in measures of EE when comparing the young and old.
There was a tendency towards a smaller increase in resting energy
expenditure (REE) in the older men compared with the young men
(p = 0.07) which was accounted for by their lower fat-free mass. There
was also a significantly smaller increase in resting energy expenditure
averaged over fasting and fed states in the older men. Thus, older men
appear to have a reduction in the ability to increase EE and an alteration
in the pattern of substrate utilization in response to overfeeding.150
8.2 | Differences in the response to overfeeding
between men and women
Men and women have been included in many
studies.33,65,69,103,159,162,172,177,189,215,216,218,219 In one study, where
BRAY AND BOUCHARD
men and women with similar initial BMI and age were overfed a bal-
anced diet for 3 days, men and women gained similar amounts of
body weight and increased body fat, visceral and subcutaneous, by
similar amounts.220 There were no gender differences in response to
overfeeding in fasting glucose or insulin. The similarity in the response
of men and women to other variables was indicated by the lack of
gender by diet interactions. Acute overfeeding did not alter blood
pressure, resting metabolic rate, peripheral insulin sensitivity or meta-
bolic flexibility as measured by the change in RQ in response to the
hyperinsulinaemic-euglycaemic clamp in either men or women. There
were also no significant gender differences in changes in plasma levels
of insulin-like growth factor-1 (IGF1), angiopoietin like 6 (ANGPTL6),
selenoprotein P (SEPP1), c-reactive protein (CRP), monocyte chemo-
attractant protein-1 (MCP1) or high molecular weight adiponectin fol-
lowing overfeeding in men or women. Interestingly, weight gain in
response to overfeeding, was positively and significantly associated
with changes in HOMA-IR and MCP1 (r = 0.39, p = 0.03).220
In contrast to the effects with a balanced diet, sex may play a role
in the response to fructose during overfeeding where fasting triglycer-
ide concentrations increased significantly more in men (71%) than
women (16%).208 Endogenous glucose production was increased by
12%, alanine aminotransferase concentration by 38% and fasting insu-
lin concentrations by 14% after fructose overfeeding in men but not
in women. In men, fasting plasma FFA and lipid oxidation were
inhibited by fructose, but this did not occur in women. Thus, short-
term overfeeding with fructose produces hypertriglyceridemia and
hepatic insulin resistance in men to a greater degree than in
women.208
8.3 | Response to overfeeding in people who are
obese or lean
A number of studies have compared the response to overfeeding in
people with obesity versus people who are
lean.47,55,70,136,141,144,221–225 Several different diets were used to
increase energy intake, including a balanced
diet,47,55,136,141,221–223,226,227 a high-fat diet,144 or a high-
70,126,212
carbohydrate diet. Some studies were 7 days or
longer55,70,136,221–224,227 and others shorter in length.126,141,225,227
In a study of obese and lean adolescents, Bandini et al70 found
that increases in body fat were greater in the group with obesity
(1.5 ± 0.3 kg) than the nonobese group (0.8 ± 0.5 kg) but that the
changes in body composition with overfeeding between the two
groups were not different. There were no differences between adoles-
cents with or without obesity in response of plasma glucose and insu-
lin, resting metabolic rate, thermic effect of food or total daily
EE. Triiodothyronine increased with overfeeding in both groups
whereas thyroxine decreased with overfeeding—a finding which dif-
fers from that in adults (see Endocrine Section). Urinary catecholamine
excretion responded similarly to overfeeding in both groups of adoles-
cents. In conclusion, obesity had only minor effects on the response
to overfeeding in adolescents with different degrees of obesity.70
33
In the study of adults with and without obesity who were overfed
until they gained 10% of their baseline body weight, there were no
differences in fecal energy losses and the thermic effect of food
between the two groups of subjects.55 The baseline profile of total
daily EE, resting and nonresting energy expenditure in individuals who
were obese and those who were not was largely maintained with the
overfeeding-induced weight gain.
8.4 | Effects of overfeeding in individuals with
reduced obesity or who are prone to develop obesity
as compared with individuals who are resistant to
obesity
It is obvious that many people are susceptible to gaining excessive
amounts of weight. Although the underlying mechanisms are not fully
understood, it is clear that more energy is consumed over time, usu-
ally many years, than is required for daily energy needs. Overfeeding
such individuals or individuals who have been reduced from their
obese state provides one way of testing whether there are differences
compared with individuals who are resistant to obesity. That is, is
there a “thrifty” phenotype that predisposes to conservation of energy
or alternatively to wasting of energy during times of excessive energy
intake?
In one study, individuals with “reduced-obesity” had greater
body weight and BMI as well as percent body fat and fat-free
mass than individuals who were naturally lean. They also had
higher levels of fasting glucose, insulin and leptin.228 Three days of
overfeeding did not increase body weight in either group and did
not change the basal rate of glucose appearance (Ra), but it
reduced insulin suppression of glucose appearance (Ra) from 80%
to 60% in lean women whereas it remained unchanged in the lean
men and in the groups with reduced obesity. Individuals who were
lean had greater rates of insulin-stimulated glucose uptake
(Rd) than people with “reduced-obesity.” These findings demon-
strate that insulin action is reduced in lean women who are resis-
tant to obesity after short-term overfeeding primarily because of
an inhibition of insulin-mediated suppression of endogenous glu-
cose production. In contrast, short-term overfeeding does not
appear to affect the action of insulin in lean men or in individuals
who have reduced from their obese state. This finding may explain
the ability of lean women to maintain weight in the face of an
obesogenic environment.228
Individuals who are lean or have a reduced obese status
showed differences in the amount of energy expended by various
body compartments throughout a 24-h period. Using a respiratory
chamber, Schmidt et al155 found that 24-h EE was not different
between those who were and were not prone to obesity when
eating a eucaloric diet (2367 ± 80 vs. 2285 ± 98 kcals) and that it
increased significantly in both groups after 3 days of overfeeding.
In contrast, nighttime RQ during overfeeding increased by +6.7% in
those prone to obesity but did not change in those who were
resistant to obesity. This suggests that fat oxidation during the
34
night was down-regulated to a greater extent in those who were
prone to obesity following overfeeding.155
The concept of a thrifty phenotype was explored further by
overfeeding 200% excess energy and contrasting it with 1 day of
fasting.229 A smaller reduction in 24-h EE during fasting and a
larger response to overfeeding predicted more weight loss over
6 weeks when eating a 50% calorie restricted diet, even after
accounting for age, sex, ethnicity and baseline weight. A greater
percentage decrease in EE with fasting correlated with a smaller
percentage increase in EE with overfeeding (r = 0.27). In men, a
greater percentage decrease in EE in response to fasting was asso-
ciated with a lower 24-h core body temperature.230 The thrifty
phenotype defined by a larger reduction in EE with fasting was
also more likely to have greater overall body fat and abdominal
adiposity as well as lower core body temperature, all consistent
with a more efficient metabolism.229 These metabolic changes were
related to diet composition. The percentage decrease in 24-h EE
with fasting correlated with the percent energy expenditure during
overfeeding with a low-protein diet (r = 0.46). Also with a high
carbohydrate diet, the percentage increase in 24-h energy expendi-
ture correlated with the percentage decrease in 24-h energy
expenditure during a 24-h fast (r = 0.40). The mean percent energy
expenditure in response to overfeeding across four different over-
feeding diets was inversely related to the percentage of body fat
(r = 0.43).173
8.5 | Overfeeding in individuals who are
“constitutionally” thin or have anorexia nervosa
2
Individuals who maintain a BMI less than 20 kg/m may be
labelled as “constitutionally” thin. Whether this thinness reflects a
thrifty phenotype can be assessed by overfeeding such individ-
uals.151 Germain and colleagues76 carried out such a study of eight
constitutionally thin women and eight women of normal weight
who were overfed for 4 weeks with a high fat diet. The control
group gained 0.6 kg, compared with only 0.2 kg in the group
labelled as constitutionally thin, levels of weight gain that are
below the regression line and the lower confidence interval as
derived from 19 studies depicted in Figure 2, Panel A. It is difficult
to interpret these findings, because food intake was estimated by
dietary recall.
This problem of weight gain in “thin” people is further illustrated
by a patient with anorexia nervosa.77 The investigators reported that
increasing energy intake from 1615 kcal/day to 3200 kcal/day for
50 days increased body weight by 10 kg, from 31 to 41 kg. This period
76
is twice as long as that used by Germain et al, but the weight gain at
the half way point was still 5 kg. With the higher level of energy con-
sumed, we would have expected a weight gain in the study by Ger-
main et al to be about 2.5 kg if their participants had eaten all the
76
food provided.
BRAY AND BOUCHARD
8.6 | Overfeeding individuals with different levels of
insulin sensitivity and in those classified as having
metabolically abnormal obesity
8.6.1 | Insulin resistance
Insulin resistance is a mediator that may influence the response to
overfeeding. To test this hypothesis McLaughlin et al87 measured
subcutaneous fat cell size, insulin suppression of lipolysis and
responses of regional adipose tissue depots before and after 4 weeks
of overfeeding. Individuals were classified as insulin sensitive or insu-
lin resistant based on their response to the Steady State Plasma Glu-
cose (SSPG) test. Insulin sensitive subjects were those with glucose
<120 mg/dL and insulin resistant subjects were those with a glucose
>150 mg/dL. These two groups were matched for BMI. At baseline,
insulin resistant subjects had more visceral adipose tissue, higher
levels of intrahepatic triglycerides, higher plasma FFA levels, larger
diameter of fat cells and a higher percentage of small adipose cells.
After a weight gain of 3.1 ± 1.4 kg (3.0% in the insulin resistant
group and 3.9% in the insulin sensitive group), the individuals with
insulin resistance demonstrated no change in adipose cell size, vis-
ceral adipose tissue or insulin suppression of lipolysis, but only an 8%
worsening of insulin-mediated glucose uptake. In contrast, the indi-
viduals who were insulin sensitive showed an enlargement of their
fat cells, a decrease in the percentage of small adipose cells, an
increase in visceral fat, intrahepatic triglyceride and lipolysis, a 45%
worsening of insulin mediated glucose uptake and a decrease in
expression of lipid metabolizing genes. Smaller baseline fat cell size
and greater enlargement of fat cells with weight gain thus predicted
a decline in insulin mediated glucose uptake, as did an increase in
intrahepatic triglyceride and visceral adipose tissue and a decrease in
insulin suppression of lipolysis. Weight gain in the individuals who
were sensitive to insulin caused maladaptive changes in fat cells,
regional fat distribution and insulin resistance. The correlation
between development of insulin resistance and changes in fat cell
size, visceral fat, intrahepatic triglyceride and insulin suppression of
lipolysis highlight those factors as potential mediators of the
response to overfeeding.87
The large number of phenotypes examined in this comparison of
individuals matched for BMI but with different levels of insulin sensi-
tivity offered an opportunity to explore the potential contributions of
various omics technologies.87,231 Weight gain was associated with
strong inflammatory and hypertrophic signatures for cardiomyopathy.
Weight loss that followed the overfeeding reversed some, but not all,
of these changes, such that a number of signatures persisted indica-
tive of long-term physiological changes. There were “omics” signa-
tures for insulin resistance that may serve as the basis for novel
diagnostic techniques. Specific molecules were highly individualized
and stable in response to the perturbations of overfeeding, and these
might well serve as targets for predictive approaches about risk for
obesity.231
BRAY AND BOUCHARD
8.6.2 | Metabolically abnormal obesity
Obesity is commonly associated with insulin resistance and increased
hepatic triglyceride content which are key risk factors for diabetes and
cardiovascular disease. However, a subset of people with obesity do not
manifest these metabolic derangements, and these individuals might
respond differently to overfeeding than those with risk factors for car-
diovascular disease. To test this idea, 12 people with “metabolically nor-
mal obesity,” “defined by having intrahepatic triglyceride levels <5.6%”
were matched for BMI with eight people who had metabolically abnor-
49
mal obesity defined as an intrahepatic triglyceride level > 10%). After a
weight gain of 5–7% induced by overfeeding, the increase in body fat
mass was the same in both groups, but many metabolic responses were
different. Those with metabolically abnormal obesity had a deterioration
of hepatic, skeletal muscle and adipocyte insulin sensitivity, as well as an
increase in VLDL apolipoprotein B100 concentration and secretion rates
whereas people with metabolically normal obesity did not show any of
these changes. Gene expression in adipose tissue from these two groups
revealed a number of striking differences. Genes involved in fatty acid
biosynthesis, lipid biosynthesis and lipid metabolism were markedly
increased by weight gain in the group with metabolically normal obesity
but not in the group with metabolically abnormal obesity. These data
indicate that biological pathways and genes associated with lipogenesis
in adipose tissue are increased in people with metabolically normal obe-
sity but not in those who were metabolically normal.
8.6.3 | Family history of diabetes or obesity
Diabetes results from a combination of insulin resistance reflecting
low response of tissues to circulating insulin and a reduced rate of
insulin secretion by the pancreas in the face of elevated levels of
serum glucose. A family history of diabetes might thus be expected
to alter the response to overfeeding even before diabetes mani-
98
fests itself. To test this hypothesis, Samocha-Bonet et al overfed
41 sedentary individuals with and without a family history of type
2 diabetes for 28 days. Liver fat, subcutaneous fat and visceral adi-
pose tissue increased similarly in both groups. In contrast, body
weight, fasting insulin and C-peptide increased significantly more at
3 and 28 days in those with a positive family history of diabetes
compared with those who did not have this history. Peripheral
insulin sensitivity measured by HOMA-IR also decreased to a
greater extent in those with a family history of T2D than in those
without it. Fasting glucose increased at both time points but there
98
was no effect of family history.
In an analysis of the relationship between family history of diabe-
tes and the response to overfeeding, two orthogonal factors were
identified which were interpreted as frame size and adiposity.
Between them they accounted for 80% of the variance in the relation-
ship of overfeeding to family history of diabetes. The family history
positive group was associated with significantly higher adiposity
scores without affecting frame size scores. Adiposity scores in family
history positive individuals conformed to a bimodal normal
35
distribution, consistent with dominant expression of major susceptibil-
ity genes.232
Low circulating levels of testosterone may be another potential
modulator for the metabolic differences in those with a family history
of diabetes and their response to overfeeding. Low testosterone has
been associated with development of type 2 diabetes in obese men
and in men with a family history of diabetes, overfeeding increased
body weight, fat mass and fasting insulin levels more.233 Men with a
family history of diabetes were more susceptible to deleterious out-
comes of overfeeding with greater reductions in serum testosterone
and dihydrotestosterone and greater increases in markers of insulin
resistance, which may contribute to increased risk of developing type
2 diabetes.233
A family history of diabetes may also differentially affect the
response to overfeeding with glucose, fructose and sucrose. Offspring of
positive family history had significantly higher intrahepatic triglyceride
and total triglycerides, but lower whole-body insulin sensitivity than did
a control group. The effects of fructose on VLDL-triacylglycerols were
greater in those who were the offspring of patients with type 2 diabe-
tes.183 A study by Seyssel et al211 of high fructose feeding in the off-
spring of patients with type 2 diabetes showed that a number of genes
were altered transcriptionally including peroxisome-proliferator activated
receptor-α (PPARA) and nuclear receptor subfamily 4 group A member
2 (NR4A2), but there was no nondiabetic control group to help separate
the effects of family history from the effects of fructose alone.211
8.6.4 | Parental obesity
Parental obesity is a strong marker for weight gain in offspring and
might influence their response to overfeeding.114 A lower resting EE
has previously been reported to predict the risk of obesity in some
studies234–236 suggesting that peri-pubertal children born to obese
parents might have a lower 24-h resting EE during “weight mainte-
nance” and/or in response to overfeeding when compared with chil-
dren born to normal-weight parents. A group of eight offspring of
parents with obesity were heavier than the eight offspring of thin par-
ents.114 Twenty-four-hour EE, adjusted for differences in body com-
position, was similar in both groups at baseline and rose similarly
during overfeeding. However, offspring of normal weight parents
were less receptive to overfeeding and returned more food than their
obese counterparts. The fact that children born to parents with obe-
sity were already overweight might have obscured an initial impair-
ment in energy expenditure.114
8.7 | Response to overfeeding of individuals who
were small for gestational age (low birth weight) and
those who were appropriate weight for gestational
age (normal birth weight)
The hypothesis that low birth weight may be a risk factor for develop-
ing cardiometabolic diseases was formulated by Barker et al237 and
36
later expanded by Wadhwa et al238 and is generally referred to as the
“Development Origin of Health and Disease (DoHAD)” hypothesis.
Overfeeding provides a tool to explore the developmental abnormalities
of individuals with low birth weight. When adults were challenged with
5 days of high-fat overfeeding with 50% extra energy, subjects with low
birth weight developed peripheral insulin resistance and reduced gene
expression of peroxisome proliferator-activated receptor gamma
coactivator 1-alpha (PPARGC1A) was significantly higher in subjects with
low birth weight during the control diet. After overfeeding, methylation
of PPARGC1A increased in a reversible manner only in the men with nor-
180
mal birth weight. There was lower DNA methylation plasticity in skel-
etal muscle from men with low birth weight compared with men with
normal birth weight. This difference might contribute to the link between
low birth weight and the increased risk of type 2 diabetes.237 Resting
muscle phosphocreatine and total ATP increased significantly after over-
feeding only in the subjects with low birth weight. Other measurements
of mitochondrial function were unaffected. Men with low birth weight
had increased fat oxidation during insulin infusion compared with men
with normal birth weight, despite similar levels of FFA. Plasma leptin
levels were higher in individuals with low birth weight and did not
increase with overfeeding, in contrast to the increase observed in those
with normal birth weight.156 There were smaller increases in plasma
glucose-insulin polypeptide (also called gastric inhibitory polypeptide)
(GIP) and pancreatic polypeptide (PP) levels in men with low birth weight
when overfed a high-fat diet. Thus, overfeeding a high-fat diet to men
with differing birth weights unmasked a number of metabolic derange-
ments. During overfeeding a high-fat diet, men with low birth weight sig-
nificantly increased nighttime EE in a metabolic chamber compared with
normal birth weight men. At the same time, nighttime glucose oxidation
rate was decreased, while both adjusted 24-h and nighttime fat oxidation
rate was elevated in low birth weight subjects. Although plasma retinol
binding protein-4 (RBP4) is associated with components of the metabolic
syndrome, it is not determined by birth weight and is not involved in
short-term high-fat diet-induced resistance to insulin.239
In a metabolomic analysis of fatty acyl-carnitines, men with low birth
weight had higher C2 and C4-OH levels after the control diet than men
with normal birth weight, indicating increased beta-oxidation of fatty
acids relative to their flux through the tricarboxylic acid cycle.239 Men
with low birth weight also had higher levels of C6-DC, C10-OH/C8-DC
and total hydroxyl-/dicarboxyl-acylcarnitine levels suggesting increased
omega-oxidation of fatty acids in the liver. Metabolomics of amino acids
revealed that men with low birth weight had higher levels of alanine, pro-
line, methionine, citrulline and total amino acid levels after overfeeding
than men with normal birth weight.188 Because the alanine and total
amino acid levels tended to be negatively associated with insulin-
stimulated glucose uptake after overfeeding, the higher amino acid levels
in men with low birth weight could be a consequence of reduced skeletal
muscle insulin sensitivity after overfeeding which could contribute to
overall impaired insulin sensitivity. In addition, alanine was negatively
associated with the plasma acetylcarnitine levels and positively associ-
ated with hepatic glucose production after overfeeding, suggesting that
the higher alanine level in men with low birth weight could be accompa-
nied by an increased anaplerotic formation of oxaloacetate and thereby
BRAY AND BOUCHARD
an enhanced tricarboxylic acid cycle activity as well an increased
gluconeogenesis.
8.8 | Effects of exercise on the response to
overfeeding
Mann et al86 published one of the early experiments on the interac-
tion between overeating and exercise (see Table 2). This experiment
was performed with four medical students and consisted of three
periods: an initial baseline period of 1 week at energy balance with
caloric intakes that ranged from 2788 to 3093 kcal/day and fat intake
from 155 to 175 g/day or 50% to 51% of the daily calorie intake. The
overfeeding period of 7 weeks consisted of two parts. During the first
3 weeks of overeating, the men ate a high-calorie diet averaging
5462–6213 kcal/day but exercised so as to keep their weight con-
stant. Fat intake remained the same as at baseline, but with the higher
energy intake, fat now represented only 24.7 to 25.2% of energy
intake. During this period, plasma lipoproteins and cholesterol
remained unchanged. However, when exercise ended but the high
caloric intake maintained during the last 4 weeks, body weight and
body fat rose and so did cholesterol and lipoproteins. These experi-
ments clearly show that EE of exercise modifies the lipoprotein
response to overfeeding.
Two studies examined altering energy balance by exercise and
overfeeding (Table S2).240,241 The first study tested the interaction of
overfeeding and physical activity as modulators of total energy expen-
diture, resting metabolic rate, the thermic effect of food and energy
expended in physical activity, whereas the second study evaluated
the effect of these variables on fat and carbohydrate oxidation on
19 men living in a metabolic unit. The study included four groups of
men all of whom spent 10 days while sedentary and in energy bal-
ance. For the next 10 days they were divided into four groups: two
groups continued the baseline food intake with one half (Group A)
remaining sedentary and thus in energy balance and the other half
(Group B) increasing energy expenditure by 50% thus inducing nega-
tive energy balance; the other two groups were overfed by 50% for
10 days with half of the men remaining sedentary (Group C) which
produced a positive energy balance, and the other half increasing
physical activity by 50% (Group D) to maintain energy balance. Rest-
ing metabolic rate increased significantly during overfeeding whether
in positive energy balance (Group C) and energy balance at high
energy flux (Group D) relative to the change in resting metabolic rate
when energy balance was maintained at low energy flux (Groups A
and B). A significant increase in resting metabolic rate was also noted
during negative energy balance after adjustment for changes in fat-
free mass. There was no effect on the thermic effect of a meal across
the treatment groups. These studies show that simultaneous increases
in energy intake and physical activity in Group D did not act synergis-
tically to raise resting metabolic rate.240,241
In another report comparing short-term positive energy balance
produced by overfeeding or reduced physical activity resulted in
impaired metabolic outcomes and altered the expression of several
BRAY AND BOUCHARD
key genes within adipose tissue that relate to energy balance, metabo-
lism and insulin action.139 These detrimental effects were largely
prevented by the addition of vigorous daily exercise even in the face
of energy surplus. Gene expression of sterol regulatory element-
binding protein 1c (SREBP1C), fatty acid synthase (FAS) and glucose
transporter type-4 (GLUT4) in adipose tissue was significantly up-
regulated and pyruvate dehydrogenase kinase 4 isozyme (PDK4), insu-
lin receptor substrate-2 (IRS2), hormone sensitive lipase (HSL) and
visfatin were significantly down-regulated by the positive energy bal-
ance produced by lower levels of exercise. Thus, a higher level of vig-
orous physical activity, at the same level of positive energy balance,
counteracted most of the effects of short-term overfeeding at the
139
whole-body level and in adipose tissue.
8.9 | Response to overfeeding in different ethnic
groups
Ethnicity is another potential contributor to differences in the meta-
bolic response to overfeeding. In a group of South Asian men and
White (non-Hispanic European) men with similar percent body fat, the
South Asian men had lower body weight and less fat free body mass,
smaller waist circumference and a smaller hip circumference, but simi-
lar waist/hip ratio.156 Short-term overfeeding with a high-fat diet had
more adverse effects on the LDL-cholesterol in the South Asian men
than the white men.143 However, despite a relatively higher percent-
age of visceral fat area, liver fat increased similarly in South Asian and
242
Caucasian men in response to overfeeding with a high fat diet.
8.10 | Response to overfeeding in individuals whose
birth followed normal conception and those whose
birth followed in vitro fertilization
106
Chen et al examined glucose metabolism in young adults conceived
by in vitro fertilization (IVF, n = 14) versus natural conception (n = 20)
under conditions of energy-balance and after overfeeding 1250 kcal/day
with 45% fat for 3 days. There was a greater increase in systolic blood
pressure and a significantly lower insulin sensitivity in patients conceived
by in vitro fertilization. Thus, in vitro fertilization, including the exoge-
nous hormonal preparation of the mother for implantation, may increase
the risk of the development of metabolic diseases later in life.
8.11 | Response to overfeeding in individuals with
lipodystrophy
8.11.1 | Lipodystrophy appearing in individuals
with HIV
The lipodystrophy associated with HIV and other syndromes of lipo-
dystrophy are characterized by extensive loss of subcutaneous adi-
pose tissue and are also associated with increased resting EE. This
37
hypermetabolism may be an adaptive response to an inability to store
triacylglycerol fuel in a normal manner. After 3 days of eating a
eucaloric diet, resting EE was significantly higher in patients with HIV
lipodystrophy (n = 9) than in either HIV-infected individuals without
lipodystrophy (n = 10) or healthy controls (n = 9) (see Kosmiski et al121)
and increased significantly after 3 days of overfeeding in patients with
HIV lipodystrophy but not in the other groups. The thermic effect of
food did not differ among the groups after a “normal” test meal but
tended to be higher in patients with HIV lipodystrophy than in healthy
controls after a large test meal. Augmented thermogenesis may be a
feature of the HIV lipodystrophy syndrome and may be due to an
inability to store lipid fuel in a normal manner.121
8.11.2 | Congenital lipodystrophy
Humans respond to an acute excess of ingested energy by storing
the surplus energy as triglycerides mainly in white adipose tissue.
In one study, total fat mass in patients with lipodystrophy was
approximately 70% lower than in control subjects (6.1 kg
vs. 22.0 kg). Energy expenditure and macronutrient oxidation rates
were assessed in a metabolic chamber on two separate occasions
lasting 40 h each, during which time subjects consumed either an
energy-balanced diet or a diet incorporating 30% excess energy as
fat. On the energy-balanced diet, both resting and total daily EE
were linearly associated with fat-free mass in both groups
(r2 = 0.83) and were not significantly different between groups
when corrected for lean mass. In response to the fat challenge,
total daily EE did not increase significantly in healthy controls and
substrate oxidation showed that the excess fat was predominantly
stored. In contrast, the lipodystrophic subjects significantly
increased total daily EE which was largely attributable to a 29%
increase in fat oxidation. Thus, subjects with congenital lipo-
dystrophy respond to overfeeding with a higher energy expendi-
ture as a result of increasing fat oxidation.191
We conclude that many variables affect the metabolic and endo-
crine responses to overfeeding. High on this list are the effects of base-
line insulin resistance and area that deserves continuing exploration. Age
is a second factor which has not been adequately explored. Exercise can
counteract some of the effects of overfeeding and deserves additional
study. Finally, the effects of age need further study. Gestational age (low
birth weight) may have significant effects on the response to overfeeding
through effects on genetic-metabolic adaptations. Further studies are
also required to confirm the finding of decreased response of resting
energy expenditure (REE) with overfeeding in older men and to deter-
mine the underlying cause of this change.
9 | H O R M O N E S A N D H O R M O N A L SY S T E M S
IN RESPONSE TO OVERFEEDING
One of the reasons that Ethan A.H. Sims and colleagues initiated
the overfeeding Vermont Studies in the 1960s was to establish
38
which of the many endocrine alterations in patients with obesity
were caused by obesity per se, and which could be replicated sim-
ply by experimental overeating. The men in his first two groups (I-
II) ate 2000 kcal/day extra of a mixed diet over 200 days and
gained 24.0 ± 4.4 % (range 21–31%) above their baseline
weight.243 In contrast to people with “spontaneous obesity,” the
obesity induced experimentally by overfeeding had no increase in
the number of fat cells244 and lower levels of FFA than people
with spontaneous obesity. Other variables, including serum choles-
terol, fasting insulin, insulin/glucose ratio, intravenous glucose toler-
ance, growth hormone levels and cortisol levels were similar
between individuals who were obese or of normal weight in the
Vermont overfeeding study.63 Sims and colleagues thus concluded
that overfeeding was a valuable model to assess metabolic changes
seen in obesity because most of the endocrine changes associated
with spontaneous obesity were reproduced with experimental
63,245
overfeeding.
9.1 | Effect of overfeeding on growth hormone
Plasma human growth hormone (hGH) concentrations are low in
people with obesity. Sims et al found that after overfeeding with a
mixed diet, the hGH peak following a glucose load fell from
30 mμg/mL before overfeeding to 16 mμg/mL after weight gain.
One participant in the Vermont Studies had no effect on the rise
in hGH after a 2% weight gain but a drop from 23.6 mμg/mL peak
at baseline to 6.3 mμg/mL peak after a 15% weight gain. At his
peak weight, which was 21% above baseline, plasma GH only rose
to 2.3 mμg/mL.11,63,243,246,247 When overfeeding was accomplished
with a high fat diet, hGH was not suppressed, suggesting the high
carbohydrate diet was responsible for the hGH suppression. These
findings in men are consistent with those of Votruba and
Jensen248 who also reported a suppression of GH in men but,
interestingly, not in women.
The most detailed studies of overfeeding and change in GH were
performed by Cornford et al.73,178,179 To assess the temporal changes in
hGH, they sampled blood every 20 min for hGH and every 2 h for insulin
and cortisol. There were six peaks of hGH rising in magnitude during the
day and falling off after midnight. During overfeeding from Day 3 through
Day 13, the hGH peaks were attenuated by over 80%. Cortisol was
unchanged, but integrated insulin rose twofold by Day 3. Plasma FFA
were suppressed by overfeeding, triglycerides were increased, and glu-
cose was unchanged. The suppression of hGH was the result of a change
in pulse amplitude of hGH secretion with no change in hGH pulse fre-
quency or interpulse level.178 Insulin like growth factor binding protein
1 (IGFBP1) was suppressed, but insulin like growth factor binding protein
3 (IGFBP3) was increased with a net effect of an increase in free insulin
like growth factor 1 (IGF1) after 2 weeks.178 Neither hGH nor IGF1 were
significantly affected by heavy resistance training while overeating a sup-
64
plemental formula for 8 weeks.
To assess whether the fall in hGH with overfeeding influenced
73
other variables, Cornford et al administered hGH or a placebo
BRAY AND BOUCHARD
four times a day during 2 weeks of overfeeding to restore circulat-
ing hGH to the preoverfeeding levels. Preventing the fall in plasma
hGH during overfeeding increased insulin resistance. It also
increased fasting plasma triglyceride concentrations and whole
body proteolytic rate after an overnight fast. Thus, the suppression
in hGH secretion during the early stages of overeating may attenu-
ate the insulin resistance and hyperlipidaemia that typically accom-
pany overeating.
9.2 | Effect of overfeeding on thyroid hormones
Thyroid hormones have a strong effect on EE. Overfeeding
increases EE providing a rationale for examining the interaction
with thyroid hormones during overfeeding and this relationship has
been evaluated in several studies6,249–251 and the main findings are
summarized in Table S3. Triiodothyronine (T3) was increased in all
but a few overfeeding studies.95,108,252,253 In the Vermont study,6
men ate both a low carbohydrate (400 kcal/m2/d) and a high car-
bohydrate (1200 kcal/m2/d) diet with similar levels of protein both
before and again after gaining weight. T3 was significantly higher
when the men were eating the high carbohydrate diet
(152 ± 6 ng/dL) than when eating the low carbohydrate diet
(136 ± 10 ng/dL). In a 3-month study from the Vermont group,
subjects were overfed 895 kcal/day as extra fat.6 T3 did not
increase after overeating with this high fat diet, suggesting that
there may be differences in thyroid response as a function of the
diet used and duration of overfeeding.
The effect of duration of overfeeding was clearly shown by
Oppert et al250 who found that T3 was significantly increased after
25 days of overfeeding but not after 50 or 100 days. In this study,
plasma thyroid hormones and thyrotropin (TSH) were investigated
during the course of overfeeding in 24 (12 pairs of identical twins)
young, normal weight men for 100 days.250 Plasma thyroxine (T4) and
free T4 were unchanged with overfeeding. In contrast, reverse-
triiodothyronine (rT3) levels were persistently decreased whereas the
TSH response to stimulation with thyrotropin-releasing hormone
(TRH) was enhanced throughout the overfeeding protocol. Within-
twin pair resemblance was observed for the changes in T4 and free T4
levels. Baseline T3 was positively associated with changes in body
weight. Moreover, the TSH response to TRH at baseline was positively
correlated with the changes in resting metabolic rate during
overfeeding.
The interaction of diet and overfeeding is clearly shown in the
study of Davidson and Chopra,249 who used isocaloric diets with
20%, 40% and 80% carbohydrate during 5 days of overfeeding. T3
was increased progressively with each carbohydrate level in the
isocaloric diets. Overfeeding also increased T3. In multiple regres-
sion analysis, the highest correlation was with calories rather than
carbohydrate, although both carbohydrate and calories increased
T3 levels. T3 is also increased with overfeeding in adolescents with
obesity70 and in one study in adults with obesity,251 but not
another.82 It was also increased during 3 weeks of overfeeding
BRAY AND BOUCHARD
with a high protein diet,6 but only with the low protein diet in the
56-day overfeeding of Bray et al.162 Danforth et al6 showed that
after 3 weeks of overfeeding that both T3 and its metabolic clear-
ance were increased, resulting in a marked increase in the produc-
tion rate of T3 irrespective of the composition of the diet that
was overfed. The higher concentrations of T3 were usually associ-
ated with lower levels of rT3, the alternative metabolic pathway
for deiodination of thyroxine. In contrast to the changes in T3, the
concentration of circulating thyroxine and TSH were largely unaf-
fected by overfeeding or changes in dietary carbohydrate. Thus,
overfeeding seems to have two effects on the hypothalamic-pitui-
tary-thyroid axis. First it increases conversion of thyroxine to T3
and with a reduction in formation of rT3 through the alternative
pathway. Raising circulating T3 by exogenous treatment reduced
TSH and thyroxine, something that does not happen with over-
feeding.137 This implies that overfeeding also reduces the feed-
back inhibition of circulating T3 on the hypothalamic-pituitary-
thyroid axis.
9.3 | Effect of overfeeding on catecholamines and
the sympathetic nervous system
Activity of the sympathetic nervous system was assessed during
overfeeding using several strategies, including measurement of uri-
nary catecholamine excretion, response to the infusion of catechol-
amines, the synthesis and degradation of catecholamines, and
electrical activity of sympathetic nerves. In a study with adoles-
cents70 and in studies of adults70,113,162,254,255 urinary excretion of
catecholamines was unaffected by overfeeding. Urinary norepi-
nephrine excretion was also unaffected by 24 h of overfeeding in
men and women when overfed with a balanced diet, a high-carbo-
hydrate, a high-fat or a low-protein diet.159 A higher urinary epi-
nephrine concentration was correlated with smaller reductions in
EE. Three days of overfeeding with either a high-carbohydrate or a
high-fat diet did not change plasma catecholamines.111 Similarly,
Welle and Campbell254 reported that plasma norepinephrine in the
recumbent position was not increased after overfeeding, but that
the peak after standing was increased by 9%. Urinary norepineph-
rine over 24 h increased in response to a 10% weight gain in
adults who were never obese as well as in subjects with obesity
(Table 7). The urinary epinephrine changes did not reach the
threshold of significance.
TABLE 7 Effect on 24-h urinary catecholamine excretion of a 10% weight gain produced by overfeeding
Never obese
Baseline
Norepinephrine, nmol/day 180 ± 20
Epinephrine, nmol/day 26 ± 7
Note. Adapted from Rosenbaum et al.223
*
Significantly different from baseline at p < 0.005.
39
The metabolic clearance of norepinephrine was similar between
subjects with obesity and lean controls at baseline and was
unaltered by overfeeding as was the metabolic responses to nor-
epinephrine.82 Propranolol, a drug that blocks beta-receptors of the
sympathetic nervous system, did not modify the stimulation of
resting metabolic rate during overfeeding with a high-carbohydrate
diet.100 Similarly, propranolol did not prevent the rise of triiodothy-
ronine during overfeeding with carbohydrates. These data do not
support an important role for the sympathetic nervous system in
mediating the thermogenic response to carbohydrates during over-
feeding, but the data below provide a different perspective.
Vinales et al159 examined catecholamine metabolism in two
groups that they labelled as “thrifty” and “spendthrift” phenotypes.
The individual with a “thrifty” phenotype has a greater reduction in
24-h energy expenditure during fasting and smaller increases with
overfeeding than those with the “spendthrift” phenotype.229 The
thrifty phenotype lost less weight in a carefully monitored inpatient
weight loss programme compared with individuals who were
defined as having the spendthrift phenotype. The spendthrift group
also had a greater increase in EE during overfeeding with a high-
fat diet but not during overfeeding with other diets. Sleeping EE
did not differ between thrifty and spendthrift phenotypes. Urinary
catecholamines during overfeeding also did not correlate with the
percent changes in EE. However, urinary epinephrine did correlate
with the percent change in fasting EE (r = 0.36), such that the
higher the urinary epinephrine concentration, the smaller the
reduction in EE during fasting. When comparing the thrifty and
spendthrift phenotypes, urinary epinephrine and dopamine were
significantly lower in the thrifty phenotype using the average of all
dietary interventions. The average urinary epinephrine across all
individual measurements correlated with percent body fat
(r = 0.35), but the other catecholamines and urinary free cortisol
did not.159
Catecholamine turnover and electrical activity from the auto-
nomic nervous system have also been used to assess the response to
overfeeding. The electrical activity of muscle sympathetic nerves
expressed either as burst frequency or burst incidence were corre-
lated with the percent increase in body weight, body fat and percent
body fat.147 The metabolic clearance rate and the appearance rate of
norepinephrine were also positively related to energy intake.89 These
studies on metabolic clearance of norepinephrine and the neural
activity of sympathetic nerves indicate that overfeeding has a modest
stimulatory effect on activity on the sympathetic nervous system,
Obese
10% weight gain Baseline 10% weight Gain
250 ± 20* 260 ± 30 330 ± 50*
47 ± 10 34 ± 7 40 ± 7
40
which contrasts from the conclusion reach from catecholamine
excretion.
In individuals who are overweight, resting postganglionic sympa-
thetic nerve discharge rate using intraneural microelectrodes on skele-
tal muscle blood vessels was directly correlated with BMI and percent
body fat.256 In addition, muscle sympathetic nerve activity was corre-
lated with age, and plasma insulin and lactate concentrations.
Together, these covariates accounted for 58% of the variance of mus-
cle sympathetic nerve activity (p < 0.0001). The rate of sympathetic
nerve discharge to calf blood vessels was also directly correlated with
calf vascular resistance (r = 0.40) but did not predict EE. Thus, body
fat is a major determinant of the resting rate of muscle sympathetic
nerve discharge. Overweight-associated sympathetic activation could
represent one potential mechanism contributing to the increased inci-
256
dence of cardiovascular complications in overweight subjects.
Muller et al252 also noted that heart rate variability, an index of
increased sympathetic and parasympathetic activity, was increased
following 7 days of overfeeding.
Blockade of the sympathetic nervous system with esmolol and
the parasympathetic nervous system with atropine provide another
way to assess the balance of the autonomic nervous system.257
The differences between people with obesity and people who
were never obese are similar to the effects produced by overfeed-
ing258 and suggest that the activity of both components of the
autonomic nervous system of individuals with obesity are chroni-
cally altered in a way that would tend to oppose excessive adipos-
ity. However, these autonomic changes are more likely to respond
to forces that induce obesity, rather than being primary agents that
produce obesity.257
Mental stress can modulate activity of the sympathetic nervous
system. Mental stress increased plasma norepinephrine and epineph-
rine to the same extent whether the healthy men were eating an iso-
caloric diet or were overeating, regardless of whether the diet was
high in carbohydrate or high in fat. Stress also increased heart rate,
blood pressure and oxygen consumption independent of type of diet
or level of calorie intake.185
9.4 | Effect of overfeeding on adrenal
glucocorticoids
Steroids from the adrenal glands are metabolized and excreted in
the urine, and individuals with obesity excrete larger quantities of
many of these metabolites, although plasma cortisol levels are usu-
ally not elevated.259 Plasma cortisol was significantly lower at
7 AM after experimental weight gain but not at other times of the
day. Cortisol production rate was significantly increased by 29%
after overfeeding, but this increase was eliminated after correction
for the change in body weight. Increased urinary excretion of
17-hydroxycorticoids (17-OHCS), a metabolite of cortisol, was also
a function of increased body weight. Normal values of plasma cor-
tisol have been noted in other studies where both high-fat or high-
carbohydrate diets were fed111 and when a balanced diet was
BRAY AND BOUCHARD
overfed.73 Urinary free cortisol did not differ by diet between indi-
viduals with the thrifty or the spendthrift phenotype.159 Thus, the
effects of overfeeding on cortisol metabolism appear to be a func-
tion of the increase in body weight. However, overfeeding a high
fructose corn syrup diet significantly increased urinary free cortisol
compared with the effect of an energy-matched overfeeding a diet
composed of whole wheat.209
Because hormones differ by gender, overfeeding might have
different effects on male versus female hormones. Testosterone in
overfed women was significantly lower, but it was unchanged
when men were overfed.252,260 In the twin overfeeding study,
baseline levels of sex hormone-binding globulin (SHBG), testoster-
one, dihydrotestosterone, androsterone glucuronide, androstene-3
α,17β-diol glucuronide and dehydroepiandrosterone-sulfate (DHEAS)
were normal. SHBG levels fell and 3α-DIOL-glucuronide rose with
weight gain. In contrast, Muller et al252 did not find any changes in
SHBG with overfeeding. The change in testosterone had a signifi-
cant inverse correlation with the change in upper body fat.260 It
was also noted that testosterone was strongly and inversely related
to visceral adipose tissue, whereas DHEAS was strongly and
inversely related to abdominal subcutaneous fat. Using a bimodal
split of weight gain, the group gaining the least fat mass had signif-
icantly greater changes in testosterone and androsterone
glucuronide.
9.5 | Effect of overfeeding on glucose and insulin
Insulin resistance during overfeeding has been measured by
changes in the ratio of glucose to insulin, by the homeostatic
assessment of insulin resistance index (HOMA-IR), by glucose dis-
posal during an euglycaemic-hyperinsulinaemic clamp, by suppres-
sion of FFA and by the change in hepatic glucose metabolism.
Several components of the overfeeding paradigm might influence
the response of glucose and insulin: (1) the quantity of energy that
is overfed; (2) the duration of overfeeding; (3) the macronutrient
composition of the food consumed; and (4) the change in body
composition.
Insulin is consistently increased by overfeeding, but glucose is
often unaffected. This is a reflection of regulatory responses that
are designed to maintain glucose within a narrow range. Insulin
and glucose can change within the first day of overfeeding, partic-
ularly at night125 and the responses are influenced by the diet
which is overfed. The metabolic status of the individual before
overfeeding also affects the response. This is particularly true if
the individual is insulin resistant, has centrally located obesity, low
birth weight or lipodystrophy. Figure 7 provides a visual schematic
summary of the effects of overfeeding on glucose and insulin as
well as a number of other systems as summarized from multiple
studies (see Table S4).
An increase in glucose and insulin can be detected as early as
14–24 h after the onset of overfeeding.125 Overfeeding with a
mixed diet increased baseline insulin as well as hepatic glucose
BRAY AND BOUCHARD
appearance and hepatic insulin resistance within the first 24 h in
men who were obese or overweight.225 Hypercaloric, compared
with isocaloric, feeding in men who were obese or overweight
increased the rate of glucose appearance in plasma, the hepatic
insulin resistance index, and the concentration and secretion rate
VLDL–apoB-100. In contrast, the clearance rate of plasma VLDL–
apoB-100, the secretion and plasma clearance rates of VLDL-triglyc-
eride, and the appearance rate of free fatty acids in plasma were
not affected by overfeeding.225
After 1 day of overfeeding a high-fat diet, insulin sensitivity, mea-
261
sured by the Matsuda index, and FFA were reduced. However,
baseline glucose and insulin were not different in this study nor in the
study by Ravussin et al,95 but the glucose response to a 2-h glucose
tolerance test was significantly increased.95 In the first 3 days of over-
feeding a high-fat diet, intramuscular lipid levels increased and insulin
sensitivity was demonstrated with an euglycaemic hyperinsulinaemic
clamp. The increase in C-peptide and insulin observed by after 4 days
of overfeeding by Clore et al107 is consistent with an increase in insu-
lin secretion. After 5 days of overfeeding, hepatic glucose production
and fasting glucose levels were significantly increased. However,
peripheral action of insulin, muscle mitochondrial function and gene
expression for general and specific enzymes of oxidative phosphoryla-
tion were unaffected by high-fat feeding. Insulin secretion increased
appropriately to compensate for hepatic, and not for peripheral, insu-
lin resistance, and it is thus clear that increased insulin secretion pre-
cedes the development of peripheral insulin resistance, as well as
mitochondrial dysfunction in response to overfeeding, suggesting a
role for insulin per se in this process.157 In another paper, increasing
dietary carbohydrate intake increased the appearance rate of glucose
and oxidation of glucose in a dose-dependent manner. These effects
of carbohydrates on whole-body fuel selection occur both by
F I G U R E 7 Schematic overview of the effects of overfeeding on glucose and insulin and on major organs participating in their metabolism
such as skeletal muscle, adipose tissue, liver and other organs. See text for additional details and Table S4
41
providing substrate and by altering serum insulin concentrations. In
contrast to carbohydrate, surplus dietary fat had little effect on inter-
mediary metabolism or whole-body fuel selection.134
Perfusion of the forearm with a solution containing insulin pro-
vides a direct way to assess the action of insulin primarily on muscle.
Horton et al11 used intra-arterial infusions of insulin to measure
changes in arteriovenous differences in glucose, amino acids and
other relevant metabolites before and after overfeeding in the Ver-
mont study. They found diminished glucose uptake in response to
insulin after overfeeding as compared with before overfeeding. In this
study of forearm metabolism volunteers were overfed a mixed diet.
Basal glucose uptake by muscle was unchanged but the response to
the intra-arterial infusion of insulin was markedly blunted after weight
gain in four of the five overfed individuals who were studied.11 In
these same subjects, six of 16 individual amino acids (leucine, isoleu-
cine, tyrosine, phenylalanine, glycine and threonine) were increased
after weight gain.262 This was associated with increased release of
those amino acids from muscle in the basal state and diminished inhi-
bition of their release by insulin. Thus, the development of insulin
resistance in muscle, as well as in adipose tissue,244 occurred follow-
ing experimental weight gain in normal subjects and is similar to that
seen in spontaneous obesity.11,263 To evaluate whether these effects
of insulin sensitivity were affected by the amount of carbohydrate in
the diet, a high-fat diet was used to induce weight gain. Before weight
gain, basal glucose uptake and the response to intra-arterial insulin
infusion were the same as in normal controls. After weight gain, the
basal glucose uptake was slightly but not significantly higher than that
observed at initial weight, but the response to insulin infusion was
markedly blunted.11
The reduction in plasma growth hormone (hGH) with overfeeding
may play a role in the changes in insulin concentration and insulin
42
resistance, because replacement of hGH reduced the usual
decrease in FFA that occurs with overfeeding and enhanced the
rise in fasting and post-prandial insulin resistance along with
blunting of lipolysis.178 The fall in hGH may help attenuate the
insulin resistance and hyperlipidaemia that typically accompany
overeating. In muscle biopsies, phosphorylation of insulin receptor
substrate-1 (IRS1-Ser312), protein kinase B (also known as Akt-Ser
473) and c-Jun N-terminal kinase were not altered, leading
Cornford et al73 to conclude that muscle was probably not a major
site for the insulin resistance after 2 weeks of overfeeding. Cornier
264
et al concurred and reported that tyrosine and serine phosphor-
ylation of IRS1 and serine and threonine phosphorylation of ribo-
somal protein S6 kinase beta-1 (p70S6 kinase) were unaffected by
3 days of overfeeding. However, they went further and found that
the reduction in insulin sensitivity was associated with increased
expression of skeletal muscle p85α, an alteration in the ratio of
p85α to p110 and a decrease in the amount of IRS1-associated
p110 along with a decrease in phosphoinositide 3-kinase (PI3K)
activity. Increases in expression of p85α and in the p85α:p110 ratio
were inversely correlated with insulin sensitivity, and changes in
PI3K activity correlated with changes in insulin sensitivity.264
FGF21 provides another potential mechanism for insulin resis-
198
tance with overfeeding. Heilbronn et al found that insulin sensitiv-
ity after an euglycaemic hyperinsulinaemic clamp was decreased and
metabolically protective cytokines, including FGF21 and adiponectin,
were increased after 3 days of overfeeding. The effects of overfeed-
ing on FGF21, however, differed between diets. With a high-fat diet,
there was no change in FGF21, but a significant eightfold increase
with a high-carbohydrate diet.124 Interestingly, the change in FGF21
correlated with leg glucose uptake and with increased hepatic glucose
production. These authors also noted that the phosphorylation of
Ser660 in hormone sensitive lipase and the level of perilipin-1 in adi-
pose tissue were increased suggesting that adipose tissue is a site for
changes in insulin responsiveness with overfeeding.124 Dietary pro-
tein may also affect FGF21 levels. Concentrations of FGF21 did not
change significantly between baseline and Day 28 of overfeeding in
subjects fed a normal 15% protein diet, but they rose significantly
over the same time interval in individuals eating a 5% low protein diet.
Expressed as percentage change from baseline, FGF21 concentrations
increased by 171% in subjects overfed the low protein diet.265
Karschin et al205 also argue that changes in adiponectin, leptin and
their ratio might partly explain the change in insulin sensitivity. In con-
trast to the changes described above, several other cytokines, includ-
ing plasminogen activator inhibitor 1 (PAI1), lipocalin-2 (LCN2) and
fatty acid binding protein-4 (FABP4) were not changed by
overfeeding.198
Although 5 days of overfeeding a high-fat diet increased hepatic
glucose production and fasting insulin, there were no changes in the
expression of genes related to oxidative phosphorylation in muscle.157
Changes in glucose concentration could be attributed to increased
glucose production, because there was no change in peripheral insulin
resistance using an euglycaemic hyperinsulinaemic clamp. Thus, 5 days
of overfeeding with a high-fat diet unmasked a dissociation between
BRAY AND BOUCHARD
insulin resistance and mitochondrial dysfunction but only in individ-
uals with a low birth weight.156
The interaction between dietary glucose and energy intake during
overfeeding was teased apart further by Davidson and Forsythe.266
They studied isocaloric diets containing 20%, 40% or 80% carbohy-
drate and diets containing 20% and 40% carbohydrate that were
overfed by 2000 kcal/day for 5 days. Insulin responses were posi-
tively related to both the caloric intake and carbohydrate content of
the diets, both of which affected insulin secretion, but these effects
were independent of each other. Glucose disappearance rates were
highly correlated with acute insulin response to glucose. Raising either
carbohydrate content isocalorically or increasing calories in diets with
normal carbohydrate content increased fasting plasma glucose. How-
ever, if either calories or carbohydrate were already high, increasing
the other one had no effect on fasting plasma glucose. Their study
also showed that glucose disappearance rates were tonically modu-
lated by cholinergic receptors, because the anticholinergic drug, pro-
pantheline bromide, increased glucose disappearance.266
To study the effects of overfeeding on changes in insulin,
Olefsky90 et al fed eight (6M/2F) normal weight individuals an isocalo-
ric liquid formula diet (35 Cal/kg) which consisted of 43% carbohy-
drate, 15% protein and 42% fat for 7 days. This was followed by
3 weeks of overfeeding an additional 2000 cal/day which was pro-
vided with an average composition of 50% carbohydrate, 15% protein
and 35% fat. After 1 week of increased caloric intake, they found a
significant 22% increase in fasting plasma insulin, a 5% increase in glu-
cose and a 30% increase in triglycerides. Most of these changes ret-
urned towards baseline values during the succeeding 2 weeks
although they remained significantly elevated even at the end of
3 weeks. However, insulin resistance, as estimated by indirect mea-
surement of insulin responsiveness, did not change as a result of
2 weeks of increased caloric intake, suggesting that some of the
abnormalities of carbohydrate and lipid metabolism may be attributed
to an increased caloric intake, because these changes occurred before
significant weight gain. Alligier et al43 reported an increase in insulin
after 14 days of overfeeding that was no longer significantly elevated
at 56 days. In contrast, other studies of 56 days of overfeeding have
consistently shown increased levels of insulin.48,61,177,195,196,248,267
Others have also failed to find changes in overall glucose disposal rate
in healthy men and women fed high-carbohydrate or high-fat
diets.102,264
Overfeeding of men and women for 3 days did not alter blood
pressure, resting metabolic rate, peripheral insulin sensitivity or
metabolic flexibility as measured by the changes in RQ in response to
an euglycaemic hyperinsulinaemic clamp. There were also no
detectable changes in plasma levels of insulin-like growth factor-1
(IGF1), angiopoietin like 6 (ANGPTL6), selenoprotein (SEPP1),
c-reactive protein (CRP), monocyte chemoattractant protein-1
(MCP1) or adiponectin as a result of acute overfeeding or in the
expression of inflammatory genes in subcutaneous adipose tissue.220
Interestingly, weight gain in response to overfeeding was positively
related to changes in HOMA-IR (r = 0.4) and MCP1 (r = 0.39) (see
Chen et al220).
BRAY AND BOUCHARD
The first-phase insulin response to an intravenous glucose bolus
is a sensitive measure of β-cell function.268 Using this technique,
157
Brons et al found that first-phase insulin response to an intrave-
nous glucose tolerance test was significantly elevated after 5 days of
overeating a high-fat diet. These authors also found that the disposi-
tion index was unaltered by feeding a high-fat diet, indicating appro-
priate compensation of insulin secretion from the β-cell to changes in
insulin resistance. When they included peripheral insulin action in
their calculations, there was a disproportionately elevated insulin
secretion relative to peripheral insulin action. Thus, the hyper-
insulinaemia of overfeeding may be causally related to the develop-
ment of peripheral insulin resistance after short periods (5 days) of
eating a high-fat diet.157 Despite the observed hepatic insulin resis-
tance with overfeeding, whole-body insulin-stimulated glucose dis-
posal was unaffected by 5 days of fat overfeeding or with fat and
fructose overfeeding112 or carbohydrate overfeeding,228 and over-
feeding also did not affect whole-body insulin sensitivity in lean,
healthy subjects.157
223
A study by Rosenbaum et al compared a 10% weight gain
above baseline weight in both individuals who are obese and those
who are nonobese and found no significant change of either glucose
or insulin concentrations in the individuals who were never obese.
However, there was an increase in both glucose and insulin during a
3-h glucose tolerance test in individuals who were previously obese.
Following 8 weeks of overfeeding, insulin sensitivity was
decreased 18% following an euglycaemic hyperinsulinaemic clamp
52
along with a reduction in insulin suppression of fatty acids. In the
overfeeding study with 5%, 15% or 25% protein diets, whole-body
glucose disposal was not altered,162 but insulin suppression of FFA,
another measure of insulin resistance, was significantly reduced
after overfeeding.177 When healthy men gained an average of 5% in
body weight, insulin, C-peptide and glucose were all elevated
resulting primarily from reduced insulin clearance which contributed
more to the baseline plasma hyperinsulinaemia after weight gain
48
than did insulin secretion. The associated basal and stimulated
hyperinsulinaemia resulted from differential changes of insulin secre-
tion and clearance, respectively,48 but the insulin resistance that
developed during weight gain in these men remained within the
normal range.
The magnitude of insulin resistance with overfeeding appears to
87
depend in part on the initial degree of insulin resistance. After a
comparable weight gain of 3 kg, insulin resistant individuals showed
no insulin suppression of lipolysis and only an 8% worsening of insulin
resistance. In contrast, the insulin sensitive subjects had a significant
45% worsening of insulin resistance as measured by insulin-mediated
glucose uptake.
Glucagon produced in the alpha-cells of the pancreas was mea-
sured in short-term and longer-term studies with divergent results. In
one short-term study comparing high-fat and high-carbohydrate diets,
glucagon was unchanged,111 but when fructose was overfed glucagon
increased, and the rise was not prevented when essential amino acids
194
were added to the diet. Glucagon levels were increased in one
43
long-term study45 and one intermediate length one153 but not in
another.190
In the 100-day overfeeding study of 12 pairs of identical male
twins, the 84 000 kcal overload resulted in a mean weight gain of
about 8 kg.269 Even though fasting plasma glucose increased only
slightly, more substantial increases were observed for fasting plasma
insulin and glucagon. In a 3-h glucose tolerance test (75 g glucose),
the glucose and glucagon areas under the curve were unaltered by
the long-term overfeeding but the insulin area increased by 30%. In
contrast, during a meal test of mixed composition lasting 4 h, the
plasma insulin and glucagon areas were both increased by about 30%
with no change in the glucose area. Thus, exposure to a glucose load
and a balanced meal test uncovered a shift in the response of gluca-
gon following exposure to chronic overfeeding. Low gainers for vis-
ceral adipose tissue in response to overfeeding were characterized by
lower fasting plasma glucagon levels with no difference in fasting glu-
cose or insulin levels.
Changes in level of “inflammation” might be involved in the
changes in insulin sensitivity. In one study, there was a decrease in
insulin sensitivity, an increase in circulating C-peptide, and a rise in
MCP1 with overfeeding, but no changes in other inflammatory
markers, including IL6, intercellular adhesion molecule-1, the number
of macrophages in adipose tissue, the number of T cells, inflammatory
gene expression or changes in circulating immune cell numbers of
surface activation in the study by Tam et al,218 or the one by Chen
et al.220
9.6 | Effect of overfeeding on gastrointestinal
peptides
Because the food associated with overfeeding enters the gastrointes-
tinal tract as its first stop on its metabolic journey, changes in hor-
mones secreted by the gastrointestinal tract might be expected to
change with overfeeding or influence the response to overfeeding.
Most reports on gastrointestinal hormones showed no response to
overfeeding (Table S13). Ghrelin, a peptide produced in the epsilon
(P/D1) cells, was the peptide most often measured. In short-term
studies, ghrelin was increased in subjects overfed either a whole
wheat source of carbohydrate or high fructose corn syrup,209 but
there were no changes in glucagon like peptide-1, pancreatic polypep-
tide and polypeptide YY.209 Ghrelin, but not pancreatic polypeptide,
was increased in individuals who were lean, overweight or obese in
the study by Wadden et al.227 In most short-
117,130,142,157,205,252,270
term and intermediate-term studies80,88,97
ghrelin was unchanged, but in long-term overfeeding studies, ghrelin
was increased in one study76 but decreased in two others.48,80 The
basis for these discrepancies is unclear.
Glucose-insulin dependent peptide (GIP), produced in the L cells,
was increased in the two short-term studies in which it was mea-
sured.130,157 There were no measurements of this peptide in longer-
term studies.
44
PYY3–36, produced in the L cells, had a variable response to
overfeeding. In one short-term study, 24-h integrated values for
PYY were significantly increased in response to 3 days of overfeed-
ing in individuals who are resistant to obesity but not in the individ-
uals who are prone to obesity. This study also showed that a
greater response of PYY following overfeeding was associated with
lower ad libitum energy intake, which is consistent with the idea
that individuals prone to obesity may be more susceptible to
reduced satiety and greater energy intake during the evening.271
PYY increased in two other short-term studies,157,222 but in one
117 97
short-term and one long-term study there was no change. In
contrast, a decrease in PYY3–36 was reported in one short-term
study158 and one intermediate study.76 Again, the reasons for these
differences are not apparent.
Glucagon-like peptide-1 (GLP-1), which is produced in “K” cells,
was reported to be increased in four short-term studies130,157,209,272
117,130
but unchanged in two other short-term studies. One long-term
study reported no change.45 It was reported to be decreased in one
76
arm of another intermediate-term study, but not the other arm.
Pancreatic polypeptide produced in the F cells was increased in
76,157
two short-term studies, but unchanged in one short-term
study.130 This peptide was not reported in any of the long-term stud-
76
ies but was increased in one intermediate-length study and
unchanged in another.97
Cholecystokin (CCK), produced in entero-endocrine cells, was
one of the earliest peptides shown to reduce food intake when admin-
istered to animals and human beings. Plasma CCK after overfeeding
was only measured in one study75 where its rise after a standard
breakfast was greater at 90 and 120 min after overfeeding than
before. These elevated CCK levels might suggest a corresponding
down-regulation in CCK receptors responsible for feedback inhibition
75
of CCK release.
In summary, the initial exploration of overfeeding as a model to
separate causes from effects on the endocrine system has been
greatly extended since the work of Sims and colleagues. The increase
in insulin is a consistent change occurring early and related to initial
state of insulin resistance. The rise of T3 with overfeeding is
influenced more by carbohydrate in the diet than by excess energy.
The suppression of growth hormone occurs rapidly, and influences
insulin sensitivity, and the cellular mechanisms responsible for the
rapid suppression of growth hormone still need to be clarified. The
gastrointestinal hormones have been explored, but there is consider-
able inconsistency in the responses leaving an area for further
understanding.
1 0 | O V E RF E E D I N G A N D B R A I N I M A G I N G
Food choices and food intake are largely controlled by the brain.
Great strides have been made in understanding the biology and psy-
273
chology of food intake regulation and its relation to obesity, which
has been facilitated by the development of techniques to image the
brain. Overfeeding leads to a number of alterations which involve the
BRAY AND BOUCHARD
brain thus providing a tool to examine the response of this system
(Table S6).
To examine neural signalling, Cornier and colleagues have used
three strategies: the first was to compare 2 days of overfeeding with
an eucaloric period in thin individuals; the second was to compare
responses to overfeeding in individuals who are thin and those who
had been reduced from their obese state; the third was to compare
response to overfeeding and underfeeding in individuals who are
prone to obesity and those who are resistant to obesity. Before over-
feeding in short term studies with thin individuals, pictures of foods of
high hedonic value elicited greater activation of the inferior temporal
visual cortex, posterior parietal cortex, premotor cortex, hippocampus
and hypothalamus than did pictures of neutrally rated foods,274,275
which is consistent with visual processing and attention. Two days of
overfeeding led to significant attenuation of these responses and also
reduced hunger ratings and increased satiety ratings.275 Two days of
overfeeding also significantly attenuated the response in the insula
and visual cortex as well as hypothalamic responses in thin as com-
pared with individuals with reduced obesity.276 Using the second
strategy, Cornier et al276 compared 22 individuals who were thin (BMI
19–23 kg/m2) with no family history of obesity and a stable weight
for 10 years to 19 individuals who were reduced from their obese
state before and after overfeeding by 30% for 2 days. Functional MRI
was performed while subjects using images of foods with either high
hedonic value or neutral nonfood objects and elicited significantly
greater activation of the insula and inferior visual cortex in the thin
individuals as compared with individuals with obesity who had been
reduced in weight. The 2 days of overfeeding led to significant attenu-
ation not only of the insula and visual cortex but also of the hypotha-
lamic response in the thin as compared with individuals reduced from
their obesity. Using the second strategy, Cornier and colleagues277
found that underfeeding produced greater differential effects than
overfeeding. Underfeeding of individuals resistant to obesity signifi-
cantly increased activation of the insula, the somatosensory cortex,
the inferior and the medial prefrontal cortex, parahippocampus,
precuneus, cingulate, as well as the visual cortex. In contrast, individ-
uals who were prone to obesity had minimal responses to underfeed-
ing. In contrast, overfeeding was associated with a reduction in
activation of the inferior visual cortex, but there was no difference
between individuals who were thin and those prone to obesity. This is
clearly an area in need of further research.
In summary, the study of brain imaging in response to overfeeding
is a promising area for further research, particularly comparing individ-
uals before and after weight gain to individuals with obesity.
11 | E F F E C T S OF OV E R FE E D I N G O N T H E
C A R D I O V A S C U L A R SY S T E M
Cardiovascular disease is one of the undesired consequences of cor-
pulence.278 Excess energy intake, diet composition and degree of obe-
sity, among others, can each play a role in the risk of heart disease.
Changes in heart rate, myocardial function, blood pressure,
BRAY AND BOUCHARD
endothelial function and inflammatory markers are all responses that
might be affected by either excess energy intake or dietary composi-
tion during overfeeding, and these are reviewed below.
11.1 | The heart
Using the parenteral route for overfeeding, Casper et al105 found that
when the nutrient intake of eight healthy men was doubled from mainte-
nance levels, myocardial oxygen consumption and pulse rate increased
105
over the first 4 days of overfeeding and then plateaued. An increase
in extracellular sodium balance was a major cause of weight gain over
the first 3 days. Variation in blood pressure was small and no major
changes were observed throughout the study.105 In another 3-day study
of overfeeding achieved by supplementing the regular diet with
800 mL/day of cream, intrahepatic triglyceride, as assessed by magnetic
resonance spectroscopy, was significantly increased in parallel with the
rise in plasma FFA and triglycerides. In spite of these systemic changes in
triglycerides, myocardial triglycerides and myocardial function remained
unchanged in the 15 healthy men in this study.200 Thus, there appears to
be differential, tissue-specific partitioning of excess fat intake between
cardiovascular tissues and other ectopic tissues such as liver during over-
200
feeding of a high-fat diet.
Cardiorespiratory fitness is an important predictor of future
cardiovascular problems.279 To assess the effects of fitness on the
response to overfeeding, Gentile et al51 measured maximal oxygen
uptake in a group of 13 men and divided into the upper and lower
half by fitness. Those with higher fitness had less total body fat
(13.0 ± 1.7 vs. 16.9 ± 1.3 kg) as well as less visceral fat (49 ± 6
vs. 80 ± 14 cm2). Resting blood pressure and 24-h ambulatory
blood pressure were similar in the two groups at baseline, but after
gaining ≈5 kg by overfeeding there was an increase in resting sys-
tolic blood pressure (1 ± 2 mm Hg vs. 7 ± 2 mm Hg) and diastolic
blood pressure (−1 ± 4 mm Hg vs. 5 ± 1 mm Hg) which was
smaller in those with the higher cardiorespiratory fitness, even
though weight gains were similar between groups. Cardiorespira-
tory fitness was inversely correlated with the increase in resting
systolic blood pressure (r = −0.64) and diastolic blood pressure
(r = −0.80). These relationships between cardiorespiratory fitness
and blood pressure remained significant after adjusting for changes
in the proportion of total abdominal fat gained as visceral adipose
tissue during this 8-week overfeeding study.147
11.2 | Heart rate and blood pressure
200,280
Heart rate was increased in some studies of overfeeding, but
not in others.51,113 Similarly, blood pressure was unchanged in many
overfeeding studies,87,106,113,157,200,201,281 but in others either dia-
stolic, or more usually systolic blood pressure increased in other
studies.89,147
Even though standard blood pressure measurements may not
change, mean arterial pressure may still be significantly
45
increased.282 The response of blood pressure, like the response of
insulin and glucose, may also be affected by the initial metabolic
status of those being overfed. Individuals with metabolically normal
obesity showed only a small change in systolic blood pressure
(before 123 ± 12 mm Hg; after 118 ± 13 mm Hg) with overfeed-
ing. In contrast, individuals with metabolically abnormal obesity had
a significant increase in systolic blood pressure from 128 ± 13 mm Hg
to 139 ± 6 mm Hg as well as an increase in diastolic blood pres-
sure from 71 ± 8 mm Hg to 81 ± 6 mm Hg.49 A similar change
was seen between individuals with insulin sensitivity who signifi-
cantly increased their diastolic, but not systolic, BP during over-
feeding for 4 weeks, whereas those with insulin resistance had no
changes in either diastolic or systolic BP.87
Two studies of ambulatory blood pressure during overfeeding
provide data on both systolic and diastolic blood pressure throughout
the 24 hours.51,280 In the data reported by Gupta et al280 partici-
pants increased their body weight by 7.4 kg, body fat by 2.0% and
visceral adipose tissue by 0.2 L. Intrahepatic triglycerides also
increased as did subcutaneous adipose cell size along with signifi-
cant changes in plasma variables such as hs-CRP which increased
as did leptin, and fasting plasma glucose, fasting insulin and
HOMA-IR.283 Circadian systolic blood pressure variability increased
significantly as did pulse pressure, but circadian diastolic blood
pressure was only marginally increased. In another study of ambu-
latory blood pressure, Gentile et al51 compared those with high
and low cardio-respiratory fitness. Daytime ambulatory systolic
blood pressure increased in both groups, but ambulatory diastolic
blood pressure was significantly lower in the group with high car-
diovascular fitness. Most interesting, nighttime ambulatory systolic
and diastolic blood pressure showed a time by group interaction
with both systolic and diastolic blood pressure increasing in the
low fitness group, but decreasing in the group with high cardiore-
spiratory fitness.51
11.3 | Renal function
Overfeeding also affects the cardio-renal system. In moderately obese
men, 7 days of overfeeding reduced sodium content of red blood cells
from 9.7 mmol/L to 8.4 mmol/L, and the sodium efflux rate constant
which increased from 0.45 to 0.54 h−1 (see Fagerberg et al113). Body
water also increased from 44.3 to 45.9 kg during 8 weeks of over-
feeding and as expected from the increased volume load, serum albu-
min declined from 4.4 to 4.3 g/dL.282 Glomerular filtration rate and
creatinine clearance increased282 as did plasma renin which rose from
1.2 to 2.0 mg/mL/h.147
11.4 | Endothelial function
Endothelial function assessed by flow-mediated dilatation of the
brachial artery also responds to overfeeding. In healthy normal-
weight volunteers who gained approximately 4 kg (n = 35), flow-
46
mediated dilatation decreased significantly from 9.1 ± 3% to
7.8 ± 3.2%. With return to baseline weight, endothelial function
returned to baseline levels. Interestingly, there was a significant
correlation between the decrease in flow-mediated dilatation and
the increase in visceral fat (r = −0.42), but no such relationship
was seen with subcutaneous fat.61 Blood pressure and overnight
polysomnography to measure sleep quality and quantity did not
change after gain of body fat. The resting reactive hyperemia
index, as a measure of endothelial function, declined significantly in
one overfeeding study by Gupta et al.283
In contrast to many other systems covered in this review, changes
in the cardiovascular system with experimental overfeeding are rela-
tively unimpressive.
1 2 | E F F E C T S OF OV E R F E E DI NG ON
L I P O P R O T E I N S A N D LI V E R L I P I D
High-fat and high-carbohydrate diets are known to influence circulat-
ing levels of lipids and lipoproteins.284,285 Plasma TG concentrations
are particularly sensitive to the amounts and types of dietary carbohy-
drate and fat as well as the caloric intake. These effects are exerted
through changes in triglyceride formation, triglyceride removal or
both, which in turn may be influenced by changes in FFA turnover,
insulin secretion or body weight. Overfeeding with diets differing in
fat and carbohydrate will reflect the dietary composition but can also
provide additional insight into how excess energy of each macronutri-
ent can influence lipids and lipoproteins above and beyond the effect
of the macronutrients themselves.
12.1 | Effects of overfeeding on lipids
The Vermont study by Sims et al63 documented a significant increase
in serum cholesterol and triglycerides and conversely a reduction in
plasma FFA. Sims and colleagues emphasized, however, that these
variations in blood lipids remained within the normal limits. Plasma
lipoprotein electrophoretic patterns did not deviate from normal, and
the pre-beta lipoproteins did not exceed 20% of the total. Similarly,
the ratio of the beta to the alpha lipoprotein fractions was not signifi-
cantly affected.63
Many subsequent studies of overfeeding have reported on plasma
or serum levels of triglycerides, total cholesterol, LDL-cholesterol,
HDL-cholesterol, FFA, glycerol and beta-
hydroxybutyrate32,43,48,49,52,61,63,67,69,72,73,76,77,82,87,88,90,95,97,98,101,
102,106–108,111,112,118,119,121,124,125,127,130,131,134–136,139,143,154,156,170,1-
76,178–180,182–186,189–203,208–210,212,218,220–222,224–228,243,261,264,267,270-
,272,278,281–283,286–308
(Table S7).
The effects of diet composition are shown clearly in several stud-
ies, two of which are described in more detail here. The effects of diet
on lipids were examined by using diets high in sucrose, glucose, fruc-
tose or starch as well as the response to saturated and polyunsatu-
rated fatty acids.184 They also examined the effect of overfeeding on
BRAY AND BOUCHARD
the appearance and removal of fatty acids in triglycerides by using an
infusion of radio-labelled palmitate and concluded that inadequate
removal of lipoproteins was a major cause of hypertriglyceridemia that
occurred during overeating. The rise in triglyceride concentration in
response to insulin resulted from a modest increase in the rate of
appearance of FFA in triglycerides whereas the fractional rate of
equilibration of FFA with triglyceride fatty acids fell strikingly.184
The effects of dietary fat and carbohydrate on the changes in
plasma lipids were explored further in five male volunteers living in a
metabolic unit during six stays of 5 days each when they ate, on dif-
ferent occasions, a eucaloric diet, this same diet reduced in carbohy-
drate by 25% or 50% on separate occasions or supplemented by 25%
or 50% carbohydrate or 50% fat on three separate occasions.134
Fasting hepatic glucose production varied by more than 40% between
the diets low in carbohydrate and those with the highest carbohy-
drate. Increased hepatic glucose production continued on the higher
carbohydrate diets despite significantly higher levels of serum insulin.
Lipolysis correlated inversely with carbohydrate intake, that is, the
higher glucose/insulin suppressed FFA release from adipocytes. Frac-
tional de novo lipogenesis increased more than tenfold on the higher
carbohydrate diets and was unmeasurable when eating the low carbo-
hydrate diets. De novo synthesis of lipid in the liver amounted to less
than 5 g/day even on the highest carbohydrate diet. High-
carbohydrate diets increased carbohydrate oxidation sixfold and
decreased fat oxidation. Carbohydrate oxidation was highly correlated
with hepatic glucose output which could account for 85% of fasting
carbohydrate oxidation on the +25% carbohydrate and 67% on +50%
carbohydrate diets. The +50% fat diet did not affect de novo lipogen-
esis, hepatic glucose production or metabolic fuel selection. Schwarz
et al134 concluded that carbohydrate intake alters hepatic glucose pro-
duction in a dose-dependent manner, and surplus carbohydrate is not
converted to fat in the liver.
The two overfeeding studies of monozygotic twins have dealt
with the response of plasma lipids and lipoproteins. Following a
22-day overfeeding protocol with six pairs of MZ twins, there were
significant increases in serum cholesterol and LDL-cholesterol, with
large inter-individual variations in the response.291 In the 100-day
overfeeding study with 12 pairs of male monozygotic twins, plasma
triglyceride levels increased significantly, but plasma cholesterol did
not change. Plasma VLDL cholesterol, VLDL-apo B, LDL cholesterol
and LDL-apo B rose, whereas HDL cholesterol levels fell raising the
CHOL/HDL-C ratio by 20%.300
Exercise modifies the changes in lipids produced by overfeed-
ing.86 When four healthy medical students doubled their caloric
intake, sufficient exercise to prevent weight gain prevented a rise in
serum lipoproteins or cholesterol. When the exercise terminated,
these men gained weight and their serum lipids increased.86
Two studies applied metabolomic techniques to the study of
changes in lipids. In one study of 21 men and 20 women who were
nondiabetic and who overate 1250 kcal/day for 28 days, c-reactive
protein, high-density lipoprotein-cholesterol (HDL-C) total cholesterol
and urinary F2-isoprostanes increased significantly with overfeeding,
but circulating concentrations of FFA, triglycerides and low density
BRAY AND BOUCHARD
lipoprotein cholesterol (LDL-C) were unchanged.189 Of the 333 serum
lipids measured in this study, 13% increased and 20% decreased with
overfeeding. The most notable increases were observed in the HDL-
associated phosphatidylethanolamine-based plasmalogens and their
precursor alkyl-phosphatidylethanolamine (18 ± 5% and 38 ± 8%,
respectively). Total diacylglycerol and lysoalkyl-phosphatidylcholine
concentrations decreased, whereas total ceramide, Cer22:0 and
Cer24:0, increased. These increases in ceramides, if left unchecked,
may promote systemic IR and increase cardiovascular risk. Uniform
increases in plasmalogens and their precursors were also observed.
Because plasmalogens are powerful antioxidants, they may represent
an appropriate response to the increased oxidative stress generated
by overfeeding.189
In a second study using metabolomic/lipidomic techniques, Morio
196
et al studied 19 lean and 19 overweight male volunteers who over-
ate a lipid-enriched diet of 785 kcal/day for 56 days. After overfeed-
ing, urinary metabolomic profiles were characterized by an increase in
short- and medium-chain acylcarnitines, as well as bile acids in the
overweight subjects. The time-course for changes in metabolomic
parameters were similar in both normal weight and overweight indi-
viduals. Plasma unsaturated lysophosphosphatidyl choline (22:6)
decreased more over time in the subjects with normal weight whereas
most of the saturated species increased in both groups. There were
subtle changes in plasma and urine metabolites, mostly related to dif-
ferences in the adaptation of β-oxidation and inflammation indicating
a lower metabolic flexibility in the subjects who were overweight dur-
196
ing overfeeding. This increase in lipid storage was associated with
increased expression of stearoyl coenzyme A desaturase 1 (SCD1);
diacylglycerol O-acyltransferase 2 (DGAT2), sterol regulatory element
binding transcription factor 1c (SREBP1c) as well as lipid droplet regu-
lation (perilipin 1) cell death inducing DNA fragmentation factor like
effector A (CIDEA).176
12.2 | Effects of overfeeding on lipogenesis
Lipogenesis, the formation of fat, can be assessed in two ways. The
first is by measuring the respiratory exchange ratio (RQ) and the
second is by using isotopes to trace the formation of fat. When
the RQ exceeds one, carbohydrate is being converted to
fat.67,170,309 In one study, 16 healthy men were overfed for 3 days
with one of three diets: a high-fat diet, a mixed diet or a high-
carbohydrate diet.170 At the end of 3 days, they entered a meta-
bolic chamber for 24 h after ingesting 500 g of glucose. The stimu-
lation of oxygen consumption with a high-carbohydrate meal was
higher than when eating the high-fat diet or the mixed diet, and net
lipogenesis, that is, RQ >1.0, occurred sooner and lasted longer
when they ate the high-carbohydrate diet. In another study these
authors compared six lean and six individuals with obesity over a
14 h time period following a challenge with a 500 g drink of malt-
170
ose dextrin. The increase in the RQ was similar in both lean indi-
viduals and those with obesity, and both groups were synthesizing
fat. In the final study, they showed that a large quantity of fat was
47
stored during overfeeding with carbohydrate after glycogen stores
had been filled.67 At the peak, these individuals were storing over
200 g of fat per day almost all of which was synthesized in periph-
eral tissues.
Arsland et al101 explored de novo lipogenesis by measuring both
RQ and the disposition of isotopic tracers. They depleted their sub-
jects of carbohydrate for 3 days with a low-carbohydrate diet before
they entered the metabolic chamber for 10 days. From Days 4–10,
they were progressively overfed with a high-carbohydrate diet to
reach 5000 kcal/day. Respiratory exchange ratio or RQ rose to
1.15 ± 0.022 on Day 4 of overfeeding, indicating net fat synthesis.
Although there was net fat oxidation in the basal state
(955 ± 139 mg/kg/min), net fat synthesis increased from Day
1 (481 ± 205 mg/kg/min) until Day 4 (2243 ± 253 mg/kg/min). The
contribution to lipogenesis by nonhepatic tissues was many times
greater than that of the liver, even though hepatic lipid synthesis
increased by 35-fold. Hepatic secretion of fat synthesized de novo in
the basal state was 1.0 ± 0.3 mg/kg/min, rising to 13.8 ± 6.8 mg/kg/min
on Day 1 and 43.3 ± 16.3 mg/kg/min on Day 4, but this only
accounted for a small portion of total fat synthesis, most of which
occurred in adipose tissue.101
Hepatic de novo lipogenesis versus lipid balance was also
explored by McDevitt et al212 in eight lean and five women with
obesity. De novo lipogenesis increased after overfeeding of either
glucose or sucrose to the same extent in the lean individuals and
the women with obesity but did not contribute greatly to total fat
balance. Estimated amounts of absolute VLDL production ranged
from a minimum of 2 g/day (control) to a maximum of 10 g/day
after overfeeding. This compares with a mean fat balance of
approximately 275 g or 3.6% after 96 h of overfeeding.126 Thus,
the largest amount of fat was made in tissues other than the liver,
almost certainly carried in fat cells.310
Lipogenesis has also been measured by the incorporation of
carbon-13 from acetate into lipid in 10 pairs of men with one member
of each pair being overfed a high-carbohydrate diet and the other
member a high-fat diet for 21 days at 2460 cal/day above their base-
line energy requirements.85 Both body weight and fat gain were simi-
lar between the groups, but, as expected, fractional de novo
lipogenesis was higher in the men eating the high-carbohydrate diet
because they had to synthesize more fat.
Fructose also affects de novo lipogenesis. In a cross-over study,
seven men received a high-fructose diet for 6 days with 25% extra
energy, or fish oil (7.2 g/d) for 28 days, or the combination of fish oil
and high-fructose for 6 days, or a control diet for 6 days. The high-
fructose diet significantly increased fasting glucose, triglycerides and
fractional de novo lipogenesis, as well as endogenous glucose produc-
tion.112 Fructose also impaired insulin-induced suppression of lipolysis
in adipose tissue and endogenous glucose production but did not
change whole-body glucose disposal.112 Fish oil significantly
decreased triglycerides after high-fructose diet compared with high-
fructose diet without fish oil and tended to reduce de novo lipogene-
sis but had no other significant effects. It is thus clear that overfeeding
a high-fructose diet can induce dyslipidemia, de novo lipogenesis and
48
insulin resistance in the liver and adipose tissue. Fish oil reversed the
dyslipidemia but not the insulin resistance.112
12.3 | Effects of overfeeding on liver fat
Increased liver fat is often found in patients who are obese or have dia-
311
betes. The role of dietary energy and specific macronutrients can be
untangled using imaging techniques of liver fat in individuals who have
been overfed. In response to an overfeeding protocol lasting 3 or more
days, intrahepatic triglyceride content increased101,123 but most of the
101,126
fat gain occurred in adipose tissue, with an estimated 12% of the
overfed calories stored as glycogen and 88% as fat.126
However, because accumulation of intrahepatic triglycerides
takes time, this has been explored almost entirely in studies with over-
feeding lasting more than 7 days. Despite the many metabolic differ-
ences between individuals who are insulin resistant versus those who
are insulin sensitive,87 these two groups accumulated similar amounts
of intrahepatic lipid during overfeeding.87
There were differences in the effect of overfeeding on liver fat in
individuals with metabolically abnormal obesity as compared with
those with metabolically normal obesity.49 Metabolically abnormal
subjects with obesity had liver fat levels >10%, compared to those
without metabolic abnormalities whose intrahepatic lipid was
<5.6%.49 Overfeeding produced a similar increase in body weight and
fat mass, but in those with metabolically abnormal obesity, hepatic fat
and fat in skeletal muscle increased, along with concentrations and
secretion rates for VLDL and apoB100, whereas insulin sensitivity in
adipose tissue deteriorated.
Three weeks of overfeeding healthy men and women by
1000 kcal/day, where 98% of the extra energy was from carbohydrate
produced a significant increase in liver volume and an increase in
intrahepatic lipid, and three liver enzymes (alanine aminotransferase
(ALT), aspartate aminotransferase (AST) and gamma-glutamyl transfer-
ase (GGT).287 Overfeeding with saturated fatty acids markedly
increased liver fat compared with overeating a diet rich in polyunsatu-
rated fatty acids. The saturated fatty acid diet also caused a twofold
larger increase in visceral adipose tissue than the polyunsaturated
fatty acid diet. The increase in liver fat was directly correlated with
changes in plasma saturated fatty acids and inversely with polyunsatu-
rated fatty acids.62
Overfeeding for 56 days increased intrahepatic lipid but not intra-
muscular lipid.52 This is consistent with the idea that increased liver
fat is more detrimental than other forms of ectopic fat.312 Another
56-day overfeeding study also showed an increased intrahepatic lipid
from 16 ± 2% to 18 ± 2%.43,186 Although individuals with a family his-
tory of diabetes mellitus gained more weight during overfeeding, they
increased liver fat to the same degree as those without a family his-
tory of diabetes.98 In one study, the change in liver fat was directly
related to the change in weight after 21 days of overeating a high-
carbohydrate diet.192
In brief, the effects of overfeeding lipoproteins and liver lipids
depended strongly on the composition of the diet, as well as the mag-
nitude of overfeeding.
BRAY AND BOUCHARD
13 | E F F E C T S OF OV E R FE E D I N G O N
A DI P OSE TI S SU E BI O LO GY
Human adipose tissue is profoundly impacted by chronic overfeed-
ing. Besides the expected increase in the adipose tissue mass read-
ily detected as augmented subcutaneous fat deposition, other
changes take place at the organ and cellular levels. Alterations in
adipose tissue induced by overfeeding are commonly associated
with insulin resistance, inflammation and perturbation in adipose
tissue metabolism with potential metabolic and health conse-
quences. Here, we review early and late changes in adipose tissue
and fat cell morphology and metabolism resulting from experimen-
tal overfeeding.
13.1 | Effects of overfeeding on fat cell size and
numbers
Obesity is most often characterized by larger subcutaneous adipo-
cytes (hypertrophic obesity) compared with normal weight individ-
uals. However, a substantial fraction of human obesity has only a
mild increase in fat cell size and a major augmentation in the num-
ber of fat cells, particularly in subcutaneous fat
depots (hypertrophic-hyperplasic obesity). Two critical issues need
to be taken into account in the interpretation of the studies
on the effects of chronic overfeeding on fat cell size and
number. First, a mean change in fat cell size is commonly observed
in response to overfeeding, but this does not reflect the
distribution of fat cell sizes which is highly heterogeneous. The
mean size does not inform us on whether the observed changes in
response to overfeeding occurred mainly in a specific size
category (e.g., small fat cell size) or was observed across all
sizes of cells. This may have implications for the interpretation of
the metabolic effects of the adipose tissue expansion. Second, con-
trary to what was assumed for decades, the number of fat cells is
not constant across the lifespan. Fat cells are turning over at
the rate of about 10% per year in human subcutaneous fat.313
These two critical characteristics are at play when we try to
understand the impact of chronic overfeeding on adipocyte
morphology.
Short-term overfeeding causing a small increase in body weight
of less than 3 kg is seldom associated with a measurable increase
in mean fat cell size218,314 although there may well be shifts in the
size distribution of fat cells. Changes in mean fat cell size begin to
be observed consistently when the weight gain is more than
3 kg.177,315–317 In one study in which 20 adults (five women) were
overfed for 8 weeks by about 40% of their baseline caloric intake,
mean abdominal fat cell size increased by more than 30%, but the
increase was not affected by the protein content of the diet.177 In
another 4-week overfeeding study which produced a 3 kg weight
gain, the peak abdominal fat cell size increased and the percentage
of small fat cells decreased only in the 16 insulin-sensitive individ-
uals, but not in the 15 insulin resistant ones.87
BRAY AND BOUCHARD
The ability to recruit or differentiate new fat cells to meet the
extra storage demands resulting from chronic exposure to over-
feeding is thought to be a critical step in successful adaption to
metabolic stress.318 In early studies, it was thought that the expan-
sion of adipose tissue in adults with overfeeding was dependent
only on increases in fat cell size.317 The topic was explored further
by Tchoukalova and colleagues in an 8-week overfeeding protocol
in which 28 healthy (15 men), normal weight adults gained on
average 4.6 kg of body weight.65 Fat cell hyperplasia was found in
lower body femoral fat depot but not in abdominal subcutaneous
fat, suggesting that abdominal fat responded by increasing fat cell
size whereas the changes in femoral fat were driven mainly by the
gain in fat cell numbers. No differences in preadipocyte
abundance, proliferation characteristics or in preadipocyte sensitiv-
ity to apoptosis could be found to explain the depot differences in
response to overfeeding. However, the expression levels of the
peroxisome proliferator activated receptor gamma (PPARG2) and
CCAAT enhancer binding protein alpha (CEBPA) genes were
higher in abdominal than femoral preadipocytes, which may in part
explain the observed gain in abdominal fat cell size. One report
has shown that adipocyte macrophage colony-stimulating factor
may contribute to the increase in fat cell hyperplasia with over-
feeding, but the effect is inhibited by tumour necrosis factor-α
(TNFA).319
Adipocyte size and number may have implications for glucose
and insulin metabolism as well. in vitro studies with adipocytes
316
from overfed individuals showed diminished response of glucose
metabolism to the addition of insulin in vitro. Both adipose cell
size and dietary composition influenced the in vitro metabolism of
glucose and the response to insulin by human adipose cells. When
eating weight-maintaining isocaloric diets of similar carbohydrate,
fat and protein composition, increasing adipose cell size is associ-
ated with unchanged rates of glucose oxidation and increased rates
of glucose carbon incorporation into glyceride-glycerol in the
absence of insulin. There is also decreased stimulation of glucose
oxidation by insulin. The relationships among adipose cell size, die-
tary composition, and the metabolism of adipose tissue are similar
in spontaneous and in experimental obesity.
13.2 | Effects of overfeeding on brown and beige
adipocytes
The role of brown adipocytes in human physiology and the regulation
of energy balance has been a topic of interest for decades.320 In
human beings, overfeeding at 200% of the energy requirements for
24 h did not activate brown adipose tissue as assessed by
175
fluorodeoxyglucose-positron emission tomography. Nor was
brown adipose tissue thermal activity, assessed by infrared imaging of
the supraclavicular fat depot, increased after 8 weeks of exposure to
298
overfeeding in 13 men. Even though some evidence has been
reported regarding a lower amount and/or lower activity of brown
adipose tissue in individuals with obesity compared with controls of
49
normal weight,321–323 the role of brown fat in experimental overfeed-
ing seems to be in doubt.
In 2012, the laboratory of Bruce Spiegelman reported on a
novel type of adipocytes, which they labelled “beige adipocytes,”
and its thermogenic properties.324 Although brown and beige adi-
pocytes share some features, they are from different cell
precursors. Brown adipocytes arise from mesodermal origin and
from cells expressing transcription factors such as En1, Myf5 and
Pax7 between Days 9 and 11 of the mouse embryonic life.325 In
contrast, beige adipocytes emerge within white adipose tissue
depots although beige adipocyte precursors appear to be
different from white adipose cell precursors.325 Brown and beige
adipocytes share the ability to induce uncoupling protein 1 (UCP1)
levels with cold exposure. However, UCP1-independent beige adi-
pose tissue thermogenic pathways have been identified, with cal-
cium cycling being the most prominent and important in
thermogenesis. Even though a role of beige adipose cells and of
UCP1 dependent and independent pathways in adaptation to over-
feeding can be reasonably hypothesized, it remains to be tested
experimentally.
13.3 | Effects of overfeeding on fat cell matrix
Adipocytes are surrounded by an extracellular matrix and this matrix
undergoes constant remodelling when the adipose tissue grows or
regresses in response to positive or negative energy balance.326 Sev-
eral collagen proteins are present in the extracellular matrix of adipo-
cytes. Type VI collagen expression seems to be specific to adipose
cells, and it has been shown to interact with other extracellular pro-
teins such as fibronectin, fibulin, lumican, heparin, hyaluronan, proteo-
glycans and other cell surface receptors.326 Collagens V and VI are
thought to be involved in the process of adipogenesis and their
expression levels is increased with obesity. Adipocytes expend energy
on the maintenance and remodelling of the extracellular matrix, pro-
cesses that are regulated by insulin.327 In obesity, the remodelling of
the extracellular matrix plays a role in modulating the growth in fat cell
size whereas diminished angiogenesis favours local hypoxic conditions
resulting in altered composition and reduced stability of the extracel-
lular matrix favouring an inflammatory response and insulin resis-
tance.327,328 It has been suggested that extracellular matrix alterations
observed in obesity lead to fewer adipose tissue capillaries.329 How-
ever, if a picture is emerging on the adipocyte extracellular matrix dis-
ruptions in obesity, only a little is known on the changes resulting
from experimental overfeeding.43,193,330
Proteomic studies have identified more than 70 proteins that are
part of the adipocyte extracellular matrix.327 Because collagen VI is
specific to adipose tissue, particularly visceral adipose tissue, it was
perceived as a candidate to monitor in response to chronic overfeed-
ing. In nine subjects who gained 7.7% (about 6 kg) of their body
weight on average after 4 weeks during which they consumed 144%
of their energy requirements,330 the abdominal adipose tissue expres-
sion of collagen type 6 alpha 3 chain (COL6A3) increased by about
50
2.5-fold. Even with a smaller weight gain (2.5 kg), the expression of
COL6A3 in abdominal fat cells increased after overfeeding a lipid-rich
diet as evidenced by transcript abundance in microarray studies and
in quantitative reverse transcription PCR (RT-qPCR) validation
assays.43 The remodelling of the adipose extracellular matrix appears
to be a late event in the course of adaptation based on data compar-
ing baseline, Day 14 and Day 56 of the overfeeding protocol.
13.4 | Effects of overfeeding on adipose tissue
lipolysis
Adipose tissue is not merely a depot for storage of excess energy; it is
a key tissue in the regulation of the flow of energy and metabolic sub-
strates, its main metabolic functions being regulated by hormones and
the sympathetic nervous system. It also serves as an organ producing
and secreting multiple adipokines.331 Adipose tissue lipolysis is char-
acterized by the hydrolysis of triglyceride molecules into three FFAs
under the concerted action of three lipases. The first enzyme adipose
triglyceride lipase converts triglycerides to diglycerides, which are
then converted to monoglycerides by hormone-sensitive lipase, and
finally monoglycerides are cleaved by monoglyceride lipase into glyc-
332
erol and FFA. Adipose tissue lipolysis is mainly activated by cate-
cholamines and natriuretic peptides, and suppressed by insulin. The
obese state is characterized by poorly regulated adipose tissue lipoly-
sis and by a diminished ability of insulin to suppress lipolysis when
required for metabolic fuel homeostasis. Few studies have investi-
gated the effects of chronic overfeeding on adipose tissue lipolysis
and there remain many unanswered questions.
One of the first papers dealing with the effects of experimental
overfeeding focused on five males who gained a mean of 11 kg of body
weight over a period of about 6 weeks.174 Lipolytic assays were per-
formed on fat cells obtained from abdominal subcutaneous fat. Three
novel observations were made. First, basal, unstimulated fat cell glycerol
release was strongly correlated with fat cell size when obese and normal
weight subjects were considered (r = 0.84, N = 24) but not when the
computation was performed only on normal weight subjects before the
exposure to overfeeding. Second, overfeeding did not change dibutyryl
cyclic AMP stimulated lipolytic rate from abdominal fat cells. Third, iso-
proterenol (a nonselective β adrenoreceptor agonist) stimulated abdom-
inal fat cells yielded higher rates of lipolysis after overfeeding,
particularly in subjects with larger fat cells.174 Kashiwagi and colleagues
investigated the effects of 14 days of overfeeding resulting in a weight
gain of 3 kg in seven moderately obese Native Americans.314 Basal and
insulin stimulated glucose transport rates by isolated abdominal adipo-
cytes increased by about 100% with overfeeding. The lipolytic rates of
isolated abdominal adipocytes in the absence and presence of two iso-
proterenol concentrations decreased by 25% to 75%, with no differ-
ence in the antilipolytic effects of insulin.
In a long-term overfeeding protocol conducted in 12 pairs of
identical twins, abdominal adipose tissue lipolytic data could be
obtained before and after a mean weight gain of 8.4 kg in 20 of these
315
young men. Using collagenase-isolated adipocytes, basal,
BRAY AND BOUCHARD
epinephrine and isoproterenol stimulated lipolysis increased several
fold, but there was no overfeeding-induced changes in lipolysis
expressed per fat cell or per adipocyte surface area under any of the
conditions examined. In a more recent study in which 41 adults were
overfed for 56 days resulting in mean weight gain of 2.5 kg, the
expression levels of the lipolytic-related genes hormone-sensitive
lipase (HSL), adipose triglyceride lipase (ATGL) and monoglyceride
lipase (MGL) (assayed in 20 subjects) were not affected by the over-
feeding protocol.176 Thus, it appears that overfeeding does not sub-
stantially alter basal and stimulated fat cell lipolysis.
13.5 | Effects of overfeeding on adipose tissue
lipoprotein lipase
Lipoprotein lipase (LPL) is a complex enzyme produced by tissues such
as adipose tissue, skeletal muscle, cardiac muscle and others.333 Once
LPL is synthesized in one of these tissues, it is secreted and anchored
to the endothelial vascular luminal surface. LPL can be released from
the endothelium by heparin. LPL driven hydrolysis of triglycerides car-
ried in chylomicrons and LDL particles is the main mechanism provid-
ing FFAs for uptake and storage in adipose tissue. In human obesity,
LPL activity in adipose tissue is increased when expressed on a per
cell basis while its responsiveness to insulin or caloric challenges is
diminished.333
A few studies have examined the effects of overfeeding on
adipose tissue LPL. Adipose tissue LPL obtained from suprailiac fat
biopsies was assayed in a study of six pairs of male identical twins
who were overfed by 22 000 kcal over 22 days152 and was
unchanged by overfeeding. In a second twin study, adipose tissue
biopsies were obtained from the suprailiac region and LPL activity
expressed in micromole of FFA released per hour per 106 cells
was unchanged with overfeeding.334 In another study, adipose tis-
sue LPL gene expression was unchanged after 56 days of overfeed-
ing in 20 subjects who gained 2.5 kg on average.176 This lack of
change in LPL gene expression in abdominal adipose tissue was
confirmed in insulin sensitive (N = 16) and insulin resistant
(N = 16) adults (50% females) who gained an average of 3 kg over
4 weeks of overfeeding.87
13.6 | Effects of overfeeding on markers of
inflammation
Inflammatory changes have been implicated in the risk for cardiovas-
cular disease and are increased in people with obesity.335 Overfeeding
is one strategy for separating factors related to energy and macronu-
trient intake and the increase in body fat (Table S8). A few inflamma-
tory markers are increased with overfeeding including macrophage
colony stimulating factor319 and MCP1193,220,336 and FABP4.198 In
contrast, interleukin-6 (IL6)193,196,197,224 was unaffected by overfeed-
ing as was interleukin-8 (IL8),193 interleukin-10 (IL10)193 and
TNFA.193
BRAY AND BOUCHARD
The level of circulating c-reactive protein (CRP) has been related
to changes in body weight. Both CRP and monocyte chemotractant
protein-1 (MCP1) increased significantly after a weight gain of 2.3 kg
and a significant 11% decrease in insulin sensitivity. However, there
were no changes in interleukin-6 and intercellular adhesion molecule-
1, fat cell size, the number of macrophages or T-cells in adipose tissue
218
or inflammatory gene expression. In contrast, CRP was unaffected
129 61
by overfeeding in one short-term and one long-term study. Over-
feeding increased microvascular density and connective tissue deposi-
tion in adipose tissue, but there was no increase in the number of
43
macrophages or inflammatory cells. As might be expected in a set-
ting where fat cells are enlarging there was an increase in mRNA for
330
collagen VI, which is involved in extracellular maintenance. Muscle
shared in this vascular remodelling193 with significantly higher levels
of mRNA for collagen type 1 (COL1) and collagen type 3 (COL3) as
well as matrix metalloproteinase 2 (MMP2), whereas a number of
other markers were unchanged (matrix metallopeptidase 9 (MMP9),
TIMP metallopeptidase inhibitor 1 (TIMP1), cluster of differentiation
337
68 (CD68) and integrin expression).
The quality of the diet contributes to the inflammatory
response.201 Despite comparable weight gain in groups overfed
with saturated or polyunsaturated fatty acids, there were differ-
ences in lipids and inflammatory responses. The ratios of total:
HDL cholesterol, LDL: HDL cholesterol and Apo B: A1 decreased
in the group overfed with the polyunsaturated fatty acid diet ver-
sus the saturated fatty acid diet, whereas no significant differences
were observed for other cardiometabolic risk markers independent
of the type of dietary fat that was overfed. In this same study,
plasma proinsulin increased by 21%, insulin by 17%, proprotein
convertase subtilisin/kexin type 9 (PCSK9) by 9%, FGF21 by 31%,
endothelial markers including vascular cell adhesion molecule–1,
intercellular adhesion molecule–1 and E-selectin by 9%, 5% and
10%, respectively, whereas nonesterified fatty acids decreased by
28%. In another study, Von Willebrand Factor and endostatin
showed no change with overfeeding.201 Although the secretion of
liver-derived cytokines, angiopoietin like-6 (ANGPTL6), IGF1,
selenoprotein-P (SEPP1), CRP, adiponectin and MCP1 are altered
in individuals with obesity, and they directly induce insulin resis-
tance in both cellular and animal models, a 3-day overfeeding
period with a 45% fat diet showed no detectable effect on their
220
plasma levels.
13.7 | Effects of overfeeding on adipokine levels
Adipose tissue is a metabolically active organ as illustrated by the
diversity of the proteins it produces and secretes. These proteins are
part of the large cytokine family and when originating in the adipose
tissue are referred to as adipokines. The first adipokine identified was
adipsin (human complement factor D) in 1985.338 Leptin is a major
339
adipokine that was uncovered 10 years later. Since then, more than
200 adipokines have been identified from a secretome exploration
performed on human adipocytes in culture.340 Adipokines influence a
51
whole array of physiological and metabolic processes including regula-
tion of energy balance, hormone sensitivity, inflammation and
immunity.341–343 Examples of adipokines contributing to energy
metabolism and hormone sensitivity include leptin, adiponectin,
resistin, adipsin, apelin, vifstatin, vaspin, omentin, lipocalin-2 (LCN2),
retinol binding protein-4 (RBP4), FGF21, chemerin, preadipocyte
factor-1 (PREF1), folloistatin-like-1 (FSTL1) and osteonectin.342,343 On
the other hand, adipose tissue-secreted pro-inflammatory cytokines
are numerous (interleukin 1 [IL1], IL6, IL8, MCP1 chemokine, TNFA,
CRP, serum amyloid, angiotensinogen and plasminogen activator
inhibitor 1 [PAI1]), whereas anti-inflammatory adipokines are fewer
(IL1 receptor A, IL10).342
Plasma levels of visfatin, IL6, retinol binding protein-4, MCP1 and
CRP were unchanged by a 7 days of overfeeding.136,220,224 Short-
term fat overfeeding increased PAI1 levels in normal birth weight and
low birth weight young men.156 In contrast, CRP, TNFA, IL6, IL8 and
IL10 were unchanged by 8 weeks of overfeeding at 40% above base-
line energy requirements resulting in an approximately 10% weight
gain.193 The expression levels in abdominal subcutaneous adipose
cells were also unchanged for adiponectin, IL6, chemokine ligand
2 (CCL2), CD68 and other adipokines.
Most adipokine studies in response to overfeeding have dealt
with leptin and adiponectin and they are summarized here.
13.7.1 | Effect of overfeeding on leptin
Leptin is produced almost exclusively in adipose tissue, and when
genetically deficient is associated with massive obesity.344 Leptin con-
centrations are higher in females than males, and the bulk of the evi-
dence indicates that the sex dimorphism is not entirely accounted for
by the sex difference in total adiposity.345–347
Following its isolation in 1994, the effects of overfeeding on
plasma leptin levels were evaluated in two protocols, one with acute
overfeeding for 12 h and a second protocol that produced a weight
gain of 10%.53 There was a strong linear relationship (r = 0.88)
between gain in weight and change in plasma leptin.53 In a 14-day
study of overfeeding, leptin increased from 5.9 ± 2.5 to
10.5 ± 5.3 ng/mL.348 Leptin levels have been increased in most short-
term overfeeding studies,111,138,142,156,157,183,185,190,191,270,293 inter-
mediate length studies,193,197,348 and long-term stud-
ies.48,49,52,53,61,147,163,196,248,345,349,350 The increase in leptin
concentration is found in subjects defined as metabolically normal or
metabolically abnormal obesity.267 Only four studies did not report an
increase in leptin with overfeeding.88,129,205,252 Leptin gene expres-
sion in adipocytes is up-regulated by overfeeding, and leptin concen-
trations are correlated with body fat both before and after
overfeeding.163 Leptin gene expression in adipocytes was increased
with 8 weeks of overfeeding in both the abdominal and femoral fat
depots.163
Exercise may blunt the rise in leptin during overfeeding. When
volunteers were overfed, leptin increased, but when exercise was
added and calories increased to maintain the positive energy intake,
52
leptin concentrations did not rise any further.270 The increase in leptin
produced by overfeeding results from a significantly higher amplitude
of the 24 h plasma leptin curve. These effects of overfeeding and
exercise are manifest within 24 h. The variations of average 24 h FFA
and average 24 h glucose concentrations are almost entirely explained
by the variation in average 24 h leptin concentration.138 Plasma leptin
levels respond to short-term increases in energy intake or energy defi-
cit, but there is a lag in returning to pretreatment levels which may
predict subsequent changes in energy intake.289
13.7.2 | Effect of overfeeding on adiponectin
Adiponectin is the most abundant protein produced in fat cells and is
secreted in sufficient quantities to produce circulating concentrations
in the ug/L range. Like leptin, it shows sexual dimorphism with higher
levels in females.346 It also varies inversely with changes in insulin
resistance. With overfeeding there are mixed reports of changes in
adiponectin. In short-term overfeeding studies causing small amounts
of weight gain, adiponectin was increased in some
reports139,157,205,221,252,293 but not in others.142,182,183,191 In one
study, 5 days of high-fat overfeeding which did not lead to detectable
changes in body weight did cause an increase in plasma adiponectin in
young males with normal or low birth weight.156 Plasma leptin levels
increased with short-term fat overfeeding293 but only in the normal
birth weight group.156
In studies lasting 8–28 days, adiponectin was increased in three
studies88,197,198 but not in a fourth study.112 Among the long-term
studies lasting more than 28 days, there was no change in adiponectin
in five studies,49,52,62,193,196 but there was an increase in two stud-
61,267 349,351
ies and a decrease in two others. Blood levels of total or
high molecular weight adiponectin in two long-term overfeeding stud-
ies were unchanged49,193 and only marginally augmented in a third
study in which subjects experienced a mean weight gain of 3.8 kg.267
Interestingly, overfeeding increased the ratio of leptin/adiponectin
from 0.91 to 1.33 in the latter study. The basis for the divergent
response of adiponectin with diet and length of exposure to excess
energy awaits further research.
13.8 | Effects of overfeeding on adipose tissue gene
expression profile
Two types of adipose tissue gene expression studies have been
reported over the last decade or so: targeted adipose tissue genes and
global gene expression profiling. Studies on the effects of chronic
overfeeding on specific adipose tissue genes have focused on the
extracellular matrix, lipolysis, lipogenesis and inflammatory responses.
Expression of the extracellular matrix gene COL6A3 is increased
in adipose tissue with overfeeding.43,330 In contrast, the expression of
genes related to the lipolytic pathways, HSL, ATGL and MGL, are not
176
affected by overfeeding. Expression of the adipose tissue LPL gene
was also unchanged with overfeeding176 and this is seen in both
BRAY AND BOUCHARD
insulin resistant and insulin sensitive subjects.87 Similarly, overfeeding
did not alter the expression of adiponectin in abdominal subcutaneous
adipose tissue193 whereas a strong increase in adipose tissue leptin
gene expression was observed.163
Among other specific gene transcripts targeted in response to
overfeeding, small increases in subcutaneous abdominal fat expres-
sion of peroxisome proliferator activated receptor gamma 1 (PPARG1),
peroxisome proliferator activated receptor gamma 2 (PPARG2), adap-
tor related protein complex 2 (AP2) and uncoupling protein 2 (UCP2)
were observed in 14 healthy females who gained an average of 1.5 kg
with overfeeding.348 In contrast, 5 days of high-fat overfeeding did
not result in changes in adipose tissue expression of the PPARGC1A
gene under basal and stimulated conditions in low or normal birth
weight adults.180 The same group also showed that short-term over-
feeding with a high-fat diet failed to increase FGF21 gene expression
levels in adipose tissue or skeletal muscle in normal or low birth
weight adults, suggesting that the liver is primarily responsible for the
observed increase in serum levels of FGF21.187 In 26 subjects
exposed to 56 days of overfeeding leading to a body weight gain of
2.3 kg, a comparison of the mRNA levels in abdominal adipose tissue
for 10 genes resulted in an overfeeding-induced expression in eight of
them: transferrin, ferroportin-1, hephaestin, ferritin light chain, mito-
chondrial membrane protein (mitoNEET), stearoyl coA desaturase
(SCD), diacylglycerol O-acyltransferase homologue-2, sterol regulatory
element binding transcription factor-1 (SREBF1).352 The pattern of
changes in adipose tissue transcript abundance led to the conclusion
that chronic overfeeding alters the expression of genes involved in
the metabolism of iron. In nine volunteers, including four women, who
were either fed an isocaloric diet or overfed a high-carbohydrate diet
for 4 days, the gene expression of sterol regulatory element-binding
protein 1c (SREBP1C), fatty acid synthase (FAS) and acetyl-CoA car-
boxylase (ACC) were all increased by the short-term overfeeding in
the gluteal adipose tissue depot127 and in lean and overweight sub-
jects in another study from the same laboratory.295 Following a
weight gain of 3 kg induced by 4 weeks of overfeeding, the abdominal
adipose tissue gene expression profile of several lipogenic genes (glu-
cose transporter type 4 (GLUT4), fatty acid binding protein 4 (FABP4),
phosphoenolpyruvate carboxykinase (PEPCK), fatty acid transport pro-
tein 1 (FATP1), adipose triglyceride lipase (ATGL) revealed a decrease
in expression among insulin sensitive subjects but not in those with
insulin resistance at baseline.87
One report based on 36 individuals overfed for 28 days, with a
mean weight gain of 2.7 kg, concluded that there were no changes
with overfeeding in the number of abdominal subcutaneous fat mac-
rophages or T-cells and inflammatory gene expression levels of C-C
motif chemokine ligand 2 (CCL2), cluster of differentiation 68 (CD68),
CD11c, also named integrin alpha X (ITGAX), CD206, also named man-
nose receptor C type 1 (MRC1), cluster of differentiation 40 (CD40),
IL6, IL10 and MCP1.193,218 The response of adipose tissue inflamma-
tory markers to chronic overfeeding is potentially related to the depot
adipocyte size in the preoverfeeding state.52
A limited number of studies have explored the changes with
overfeeding of the global adipose tissue gene expression profile
BRAY AND BOUCHARD
using various microarray technologies or RNA Seq. In the first
study on this topic, Shea et al investigated the changes in the
transcriptome of abdominal subcutaneous adipose tissue in eight
lean and eight obese men exposed +40% overfeeding for 7 days.226
The mean weight gain reached about 3 kg in both groups. A total
of 45 genes were shown to be differentially expressed with over-
feeding, seven of which were down-regulated. Six of these
45 genes were differentially regulated in response to overfeeding
between subjects who were lean or obese: transferrin (TF),
stearoyl-CoA desaturase (SCD), transaldolase 1 (TALDO1), cathepsin
C (CTSC), insulin receptor substrate 2 (IRS2) and pyruvate dehydro-
genase lipoamide kinase isozyme 4 (PDK4). These genes relate to
lipid and glucose metabolism, cell adhesion processes and immune
response. The top five pathways revealed by a Kyoto Encyclopedia
of Genes and Genomes (KEGG) analysis of the overfeeding-induced
changes in adipose tissue gene expression profile identified vitamin
B-6 metabolism, pyruvate metabolism, thiamine metabolism,
glycolysis/gluconeogenesis and alanine and aspartate metabolism as
the top overfeeding-responsive pathways. In 2011, Franck et al
reported on a fast food overfeeding experiment in which six lean
subjects, including two women, were overfed for 4 weeks resulting
in a mean weight gain of 7.3 kg.50 A total of 52 genes were up-
regulated and 50 were down-regulated by overfeeding (with caloric
restriction revealing changes of expression in the opposite direc-
tion). Most of these genes were qualified as “metabolic” genes and
related to lipogenesis, protein synthesis and insulin metabolism.
In a study of 20 subjects exposed to an overfeeding protocol for
56 days, biopsies of abdominal subcutaneous fat were obtained at base-
line, 14 and 56 days for microarray exploration of gene expression.43,176
The report confirms that adipose tissue lipid storage genes (SCD1,
DGAT2, FACL1 (fatty acid-Coenzyme A ligase, long-chain 1), CIDEA and
SREBP1C) were generally up-regulated with chronic overfeeding
resulting in a mean weight gain of 2.5 kg.176 Among the important find-
ings, 10 genes related to lipid metabolism, eight genes related to angio-
genesis, five transcripts related to extracellular matrix and three genes
related to inflammation in adipose tissue were uniquely regulated after
the first 14 days of exposure to overfeeding while there were 10, 29,
60 and 14 genes of the same pathways uniquely regulated after 56 days
of overfeeding.43 Early and late adaptation to chronic overfeeding seem
to entrain different responses in adipose tissue as there were only eight
(24% of all genes observed in the pathway) lipid metabolism genes, two
(5%) angiogenesis, one (1.5%) extracellular matrix and two (11%) inflam-
matory marker genes whose differential expression from baseline was
common between days 14 and 56 of the overfeeding protocol. The vast
majority of these genes revealed by the microarray assay were con-
firmed by RT-qPCR.
Comparing 40 participants for gene expression levels in abdomi-
nal subcutaneous adipose tissue after 5 days of high-fat overfeeding
with no detectable weight gain, 3276 genes were shown to be differ-
entially regulated which represented 16.5% of all genes expressed in
the adipose tissue depot.293 The most strongly up-regulated gene
transcripts were those involved in elongation of very long chain fatty
acids including elongase 6 (ELOVL6), fatty acid desaturase 2 (FADS2)
53
and neuronatin (NNAT) while insulin receptor (INSR), insulin receptor
substrate 2 (IRS2) and solute carrier family 27 member 2 (SLC27A2)
were the most profoundly down-regulated. There were no consistent
differences in the changes in gene expression pattern between normal
and low birth weight participants. The genome-wide transcriptomic
profile of abdominal subcutaneous adipose tissue was explored in a
study in which 17 subjects, including four women, ate a hypercaloric
diet enriched in saturated fatty acids and 14 subjects, including five
women, ate a hypercaloric diet enriched in polyunsaturated fatty
acids. This resulted in mean weight gains of about 1.6 kg.297 Combin-
ing the data of both groups showed that 1117 transcripts (3% of the
total) changed their expression levels in response to the 7-week high-
fat overfeeding protocols. A subtotal of 216 transcripts were up- or
down-regulated by more than 20%. A KEGG analysis of the 776 up-
regulated genes revealed that they were mainly involved in carbohy-
drate metabolism (citrate cycle, pyruvate metabolism, glycolysis and
gluconeogenesis), lipid metabolism (biosynthesis of unsaturated fatty
acids, glycerolipid metabolism, fatty acid metabolism) and oxidative
phosphorylation. As previously shown,293 ELOVL6, SLC27A2, ATP cit-
rate lyase (ACLY), glycogen synthase 2 (GYS2) and thrombospondin
1 (THBS1) were among the most regulated adipose tissue genes with
overfeeding even in the presence of a very small weight gain.
In summary, adipose cell size and adipose tissue metabolism are
remarkably similar in spontaneous obesity and experimental overfeed-
ing. With large weight gain, abdominal fat cell size increases more so
than in the femoral. The adipose cell matrix is remodelled in response
to overfeeding as a late event seen in long-term studies. Basal and
stimulated adipocyte lipolysis are not affected by experimental over-
feeding. Similarly, no changes in adipose tissue LPL activity is
observed with overfeeding. Overfeeding causes an inflammatory
response in adipose tissue which is influenced by the macronutrient
composition of the caloric surplus. Overfeeding leads to connective
tissue deposition in adipose tissue. Blood levels of leptin increase with
overfeeding, and leptin gene expression is increased in fat depots.
Studies of blood adiponectin response to overfeeding have yielded
divergent results. Exposure to overfeeding has a profound effect on
the adipose tissue transcriptome.
14 | E F F E C T S OF OV E R FE E D I N G O N
S K E LE T A L M U S C L E B I O L O G Y
The effects of overfeeding on the morphological and metabolic char-
acteristics of skeletal muscle and the role of muscle in modulating the
response to chronic overfeeding are examined in this section.
14.1 | Effects of overfeeding on fat free mass and
muscle mass
Short-term overfeeding protocols lasting 1 week or less do not cause
measurable changes in muscle mass or in whole-body lean mass (see
Table 3). Results are more heterogeneous for overfeeding periods
54
from about 10 to 28 days. For instance, Goran et al78 overfed six sub-
jects by 50% for 10 days and found only trivial changes in fat-free
mass (mean gain of 0.1 kg). In contrast, with 9 days of overfeeding,
Ravussin and collaborators reported that 45% of the weight gain was
recovered as fat-free mass.95 As the duration of the overfeeding
increases, more consistent results are observed. Thus, fat-free mass
increased 42% with 20 days of overfeeding in a study by Welle et al169
and 50% in the report by Forbes et al74 with 21 days of overfeeding.
In studies lasting more than 1 month, there are clear gains in fat-
free mass and muscle mass. For instance, with 100 days of overfeed-
ing, the gain in FFM in 24 young men reached 2.7 kg which was 33%
of the total weight gained.44 This percentage reached 39% in the
study by Diaz et al47 in which nine subjects were overfed for 42 days.
In another protocol lasting 56 days, 29 subjects overfed by an average
of 66 000 kcal with a 15% protein content gained 3.4 kg of fat-free
mass which represented 47% of the total weight gained52 A third
study lasting 56 days found that 16 participants gained 2.4 kg with
51% of their increase in weight in the form of fat-free mass.56 Finally,
58
Norgan and Durnin registered a fat-free mass gain of 39% in six
subjects overfed for 44 days.
Two reports have dealt with the potential changes in skeletal
muscle mass with long-term overfeeding. In the first, Deriaz and col-
leagues used computed tomography scans across nine body regions,
extending from the lower part of the leg to the sternoclavicular joints,
to quantify muscle mass in 22 young men (11 pairs of twins) exposed
to 100 days of overfeeding.353 The CT estimated skeletal muscle mass
was 27 kg at baseline and it increased by 1.7 kg (SD = 1.2) with the
84 000 kcal overfeeding protocol (about 6% of the baseline muscle
mass). There were no changes in bone, nonmuscular lean mass and
residual tissues. The gain in lean mass was accounted for entirely by
the gain in muscle mass whereas the total gain in body weight was
accounted for by the gain in muscle plus adipose tissue masses.
The second report dealt with the effects of long-term overfeeding
(56 days) on muscle mass in 25 participants.162 Subjects were ran-
domly assigned to a low-protein diet with 5% of calories from protein,
a normal 15% protein diet or a high-protein diet with 25% of calories
from protein, and overfed by a 40% increase in energy intake for the
duration of the protocol. At baseline and 56 days, DXA scans were
used to derive muscle mass. Overfeeding was associated with an
increase in muscle mass (of the order of about 8%) in the normal and
high-protein groups, but there was no increase of fat-free mass in the
participants eating the low-protein diet. No effect of overfeeding or
protein content was observed for brain or bone masses.
In summary, the gain in fat-free mass is generally undetectable
with overfeeding protocol lasting a week or less. When the overfeed-
ing period extends from about 10 days to 1 month, the increase in
fat-free mass is in the range of about 40 to 50% of the total weight
gained. With overfeeding exposure lasting from 1 month to about
3 months, with the caloric excess ranging from about 50 000 to
80 000 kcal, the proportion of fat-free mass gained relative to the
weight gained ranges from 30% to 50%. The latter values are
undoubtedly affected by the techniques used to quantify body com-
position and by the protein content of the overfeeding diets. The use
BRAY AND BOUCHARD
of exercise during an overfeeding protocol, particularly resistance
exercise, influences the gain in fat-free mass relative to the total
amount of weight gained.64 Finally, long-term overfeeding with a nor-
mal protein content is associated with a gain in skeletal muscle mass
estimated to reach about 6% to 8% of the baseline muscle mass.
14.2 | Effects of overfeeding on structure and
morphology of skeletal muscle
In one study on the effects of long-term overfeeding on fibre type dis-
tribution from biopsies obtained from the vastus lateralis, there was
no change in percentage of muscle fibre type 1, 2A and 2B in 24 young
men exposed to 100 days of overfeeding354 (Table S9). Similarly, in
five subjects in which needle biopsies of the vastus lateralis were
obtained at baseline and after gaining 10% of their weight with over-
feeding, no changes were observed in muscle fibre type distributions
or in capillary density around each type of fibers or in muscle glycogen
content.349
In 21 subjects exposed to 5 days of high-fat overfeeding, changes
in the gene expression profile in samples from the vastus lateralis mus-
cle were investigated, but no significant alterations in gene transcripts
could be detected with a protocol that did not measurably increase
body weight.294
14.3 | Muscle extracellular matrix
Extracellular matrix in skeletal muscle plays an important role in meta-
bolic functions, and this has been investigated in two studies. In a
28-day overfeeding protocol with a daily caloric excess of 1250 kcal,
the expression levels of seven genes considered markers of extracellu-
lar matrix remodelling were assessed in 19 participants who had mus-
cle biopsy samples at baseline and after 3 and 28 days of
overfeeding.337 Although there were no changes at Day 3, by Day
28 mRNA levels of muscle COL1, COL3 and MMP2 were increased in
contrast to MMP9, TIMP1, CD68 and integrin which were unchanged.
In a complementary microarray pathway exploration in muscle sam-
ples, pathways related to focal adhesion, adherens junction and extra-
cellular matrix receptors were altered by 28 days of overfeeding that
resulted in a 3% weight gain.
In the second study, 29 males were overfed by 40% of their base-
line calories for 56 days and muscle extracellular matrix remodelling
was investigated.193 Increases in mRNA expression levels in muscle
ranging from fourfold to 30 fold were seen in collagens (COL1, COL3,
COL4, COL5, COL6) and extracellular matrix genes (secreted protein
acidic and cysteine rich (SPARC) and Integrin aX, with a concomitant
upregulation of the transforming growth factor beta (TGFB) signalling
pathway, a canonical pathway of fibrosis. These changes were posi-
tively correlated with the overfeeding-induced gain in fat-free mass.
Inflammatory markers were also increased in muscle with overfeeding
as evidenced by two- to fourfold higher mRNA levels of CD68, CCL2,
CD40 and nuclear factor kappa-light-chain-enhancer of activated B
BRAY AND BOUCHARD
cells (NFkB), but there was no change in TNFA gene expression. Thus,
a 10% weight gain caused by chronic overfeeding leads a major
upregulation of muscle extracellular matrix, fibrosis and inflammatory
genes, which may contribute to the development of metabolic
dysfunction.
14.4 | Intramyocellular lipid
A few reports have dealt with the topic of intramyocellular lipid depo-
sition with overfeeding in order to understand whether chronic caloric
overload of variable nutrient composition was causing increases in
muscle ectopic fat deposition. In periods of overfeeding lasting 7 days
with one diet enriched in fructose and one enriched in glucose, intra-
myocellular lipid in 11 participants increased only with the high-
glucose diet.154 However, the same group showed that fructose over-
consumption for 7 days, in 16 males who were offspring of at least
one parent with diabetes, increased ectopic lipid deposition in mus-
183
cle. Nine subjects overfed for 2 weeks (4000 kcal/week) showed
179
no changes in intramyocellular lipids. Further, 56 days of overfeed-
ing by 40% more calories did not result in an increase of muscle
ectopic fat in 29 men.52 Variation in design and sample size may
account for some of the heterogeneity in findings, and it is thus still
not clear whether chronic overfeeding leads to increased intra-
myocellular lipid deposition.
14.5 | Effects of overfeeding on muscle
mitochondria and metabolism
Given the profound influence of chronic overfeeding has on metabo-
lism, it is reasonable to hypothesize that it will also have an impact on
skeletal muscle mitochondrial biology and enzymatic markers of inter-
mediary metabolism. Brons and co-workers investigated the effects of
5 days of a high-fat, high-calorie diet in 26 young healthy males on
in vivo mitochondrial function and on muscle gene expression of per-
oxisome proliferator-activated receptor gamma coactivator 1-alpha
(PGC1A) and oxidative phosphorylation marker genes.157 Compared
with their 5-day control diet period which provided 35% of calories
from fat, the high-fat, high-calorie diet consisted of a mean intake of
4230 kcal/day with a fat content of 60%. No mitochondrial indicator,
as evaluated by 31P-MRS (magnetic resonance spectroscopy) at rest
and after an energy depleting exercise, was affected by the high-fat,
high-calorie diet in a predominantly slow twitch fibre (tibialis anterior
of the leg) or fast twitch fibre (forearm flexors) muscle. Moreover,
there were no changes in muscle gene expression levels of PGC1A or
of genes encoding proteins implicated in mitochondrial oxidative
phosphorylation under basal and insulin stimulated conditions. The
same protocol was also used in 20 young healthy males who had a
low birth weight.156 There were no consistent differences between
normal and low birth weight males in muscle mitochondrial gene
expression or indicators of oxidative phosphorylation. Thus, these
data do not support the hypothesis that short-term overfeeding with
55
high-fat diets reduces the expression of nuclear genes encoding mito-
chondrial proteins and transcription factors involved in mitochondrial
biogenesis in healthy young men.355
Long-term overfeeding may lead to different results as suggested by
the observations reported by Samocha-Bonet and collaborators.199 In
their experimental overfeeding protocol, 26 subjects, including
13 women, completed a 28-day overfeeding protocol in which they were
overfed an average of 1040 kcal/day with 46% from fat. Muscle biopsies
were obtained at baseline and at Days 3 and 28. While there was evi-
dence of increases in the protein content of complexes I, II and V of mus-
cle mitochondria after 3 days of overfeeding, the increases had
disappeared after 28 days of overfeeding. Interestingly, muscle cartinine
palmitoyltransferase 1B (CPT1B) level increased with overfeeding, but
palmitate oxidation rate, citrate synthase and beta-hydroxyacyl CoA
dehydrogenase activities in muscle homogenates were unchanged. The
major conclusion from this experiment would be that while insulin resis-
tance is induced, muscle mitochondrial markers of oxidative phosphory-
lation were unaffected following exposure to 28 days of overfeeding by
more than 1000 kcal per day.199 Muscle 5' adenosine monophosphate-
activated protein kinase (AMPK), sirtuin 1 (SIRT1) and PGC1A are part of
an energy sensing network, and their protein levels were unaffected in
another 5 day study of overfeeding in 21 healthy volunteers.171 How-
ever a high-fat or a low-carbohydrate hypercaloric diet (40% caloric
excess) for 5 days was associated with increases in muscle AMPK phos-
phorylation and PGC1A deacetylation. Changes in signalling pathways
do not appear to be sufficient to translate into major adaptive changes in
mitochondrial function following exposure to overfeeding for modest
periods of time as shown by others.156,157,199
In a 56-day study of overfeeding with a high-fat diet at
760 kcal/day, both body fat and fatty acid oxidation increased and
there was a decrease in muscle pyruvate dehydrogenase kinase
4 (PDK4).186 This decrease in PDK4, a key regulator of energy par-
titioning, might result from the inhibition of the SIRT1-PGC1A path-
way accompanied by a lower NAD+ content in muscle. Muscle gene
expression of fatty acid binding protein 3 (FABP3), stearoyl-CoA desa-
turase (SCD), DGAT2, hydroxyacyl-CoA dehydrogenase trifunctional
multienzyme complex subunit alpha (HADHA), PDK4, UCP3, sirtuin
1 (SIRT1) and sirtuin 3 (SIRT3), which encode proteins participating in
lipid transport and storage, lipogenesis and regulation of energy par-
titioning were induced by long-term overfeeding. Globally 763 genes
were differentially expressed following the overfeeding protocol and
a subset of 80 of them were shown to be overrepresented in
established gene sets such as those previously identified as contribut-
ing to mitochondrial functions, including PGC1A, PPARG coactivator
1 alpha (PPARGC1A), OXPHOS), electron transport and fatty acid
metabolism. The ratio of mitochondrial DNA to nuclear DNA was
increased by overfeeding suggesting an increase in mitochondrial bio-
genesis. Mitochondrial state III and IV respiration were consistently
higher after overfeeding in the presence of various types of sub-
strates. The measured changes in state IV were accompanied by an
increase in UCP3 mRNA and protein levels, suggesting increased
uncoupling in muscle mitochondrial respiration with exposure to long-
term overfeeding.186
56
A global transcriptome exploration combined with the study of
primary myotubes was done on biopsy samples of the vastus lateralis
muscle obtained in 10 normal weight males, first-degree relatives of
patients with type 2 diabetes.211 Briefly the 10 young men were sub-
mitted to 7 days of a control diet and to 7 days of a high-fructose diet
(3.5 g/kg fat-free mass/d) in a crossover randomized design with 4 to
5 weeks of washout. The caloric overload reached +35% of caloric
intake. The microarray analysis on muscle samples revealed that the
high-fructose diet upregulated the expression of 177 genes and
down-regulated 230 transcripts. From the panel of down-regulated
genes, a coordinated reduction in the expression levels of genes
related to lipid metabolism, fatty acid oxidation and mitochondrial
function emerged. In contrast, the up-regulated genes reflected
increased lipid deposition in skeletal muscle and induction of lipogen-
esis. Studies on primary cell cultures of human myotubes were essen-
tially negative, indicating that fructose did not regulate the key genes
of lipid metabolism and mitochondrial function in vitro.211
Finally, skeletal muscle enzyme activities indicative of mitochon-
drial biology were assayed in two long-term overfeeding studies. In
the 100-day overfeeding experiment with 12 pairs of identical twins
who were overfed by 84 000 kcal and gained 13% of their initial
weight, the activity of the vastus lateralis muscle oxoglutarate dehy-
354
drogenase (OGDH) was shown to be decreased with overfeeding.
In contrast, the activity of phosphofructokinase (PFK) was unchanged,
which resulted in a major increase in the ratio of PFK to OGDH
suggesting a reduced reliance on lipid substrates (see Table S9).
Data from Goldsmith and co-workers differs somewhat from the
previous study. They investigated for the biological characteristics of
muscle in five subjects at baseline and after gaining 10% of their initial
weight.349 There were no significant changes in mitochondrial
enzymes such as citrate synthase, 2-hydroxylCoA dehydrogenase,
cytochrome c oxidase (COX), carnitine palmitoyl transferase or in PFK.
The ratio of PFK to COX was also unchanged. One concern with this
study is that it was underpowered due to the very small sample size.
14.6 | Effects of overfeeding on skeletal muscle
work efficiency
The concept that the efficiency of muscular activity might be altered
by overfeeding has been a recurring theme and has been examined in
several studies. In their second paper on overfeeding, Miller et al
(1967) argued that the potentiation of energy expenditure by exercise
after a meal might account for apparent metabolic inefficiency after
overfeeding.356
This concept was explored in the Vermont Overfeeding
145,357
Study. The four men in this study gained 19% above their base-
line weight, but, weight gain had no significant effect on energy
expenditure during cycle ergometry compared with that recorded at
baseline weight, even when extra weights were added to provide
comparable weights at both times. Thus the 19% weight gain in these
volunteers from the Vermont Study increased their energy expendi-
ture with cycle and treadmill exercise as predicted by the weight gain.
BRAY AND BOUCHARD
Significant elevations of oxygen utilization were observed during
walking on a treadmill, but again, these were strictly proportional to
weight gain or pack loading. Oxygen uptake per unit of work remained
constant at a given exercise level, and no metabolic inefficiency could
be demonstrated.145,357
Several other studies have considered the effects of chronic over-
feeding on the whole-body energy cost and work efficiency of sub-
maximal workloads. Apfelbaum and collaborators found that a caloric
surplus of 1500 kcal for 15 days increased the energy cost of muscu-
lar work.68 However, this finding was later found to be attributable to
technical issues in the estimates of baseline work efficiency.168
Whipp, Bray and Koyal investigated the same issue in four normal
weight subjects who gained weight by consuming extra calories for
periods of up to 6 weeks.168 Work efficiency was measured during
incremental submaximal workloads on a cycle ergometer in relative
steady state (fourth minute at each workload). Work efficiency ranged
from 27% to 30% and was not affected by weight gain. They con-
cluded that weight gain after overfeeding does not reduce whole-
body work efficiency as assessed under standardized steady state gas
exchange measurements. The latter observation was confirmed by
Welle et al in five volunteers who were overfed for 20 days and
gained 3 kg on average.358 Subjects were tested on a treadmill walk-
ing at 1.6 km/h for 20 min before and after the overfeeding, and
before and after a standardized breakfast meal at both time points.
No effects of overfeeding on exercise metabolic rate could be identi-
fied beyond the increase in caloric expenditure associated with the
weight gain. To evaluate the effects of the increased metabolic rate
and triiodothyronine (T3) on exercise efficiency, Acheson et al359
administered T3 to raise oxygen consumption by 6% but found no
effect on muscle efficiency or on the thermic effect of food.
This contrasts with the data from the overfeeding study that pro-
duced 10% weight gain above baseline weight.55 This study, in con-
trast to the ones described above, was associated with variability in
skeletal muscle contractile efficiency, which contributed to differ-
ences in nonresting EE. In their 1995 paper, Leibel et al observed that
nonresting EE is the metabolic component most affected by standard-
ized protocols of weight gain or weight loss.55 They commented that
the larger body mass with overfeeding cannot be the sole explanation
for the increase in nonresting EE adjusted for fat mass and fat-free
mass and that overfeeding could lead to decreases in the efficiency
with which skeletal muscle produces mechanical work resulting in var-
iability in EE, particularly during low intensity exercise.
The same group360 examined work efficiency at low power out-
put during cycle ergometry tests and rates of gastrocnemius muscle
ATP flux during plantar flexion contractions by magnetic resonance
spectroscopy in men and women who lost 10% or gained 10% of their
initial body weight. Of interest to the present review are the data per-
taining to the subjects who underwent a 10% weight gain which
included seven males and one female (seven of them were never
obese). Because they could not find changes in the time spent on
physical activity in subjects who were overfed to gain 10% of their ini-
tial weight, their hypothesis was that weight gain after overfeeding is
accompanied by decreased whole-body work efficiency during low-
BRAY AND BOUCHARD
intensity exercise and lower skeletal muscle contraction efficiency as
evaluated by ATP fluxes. The 10% gain of body weight was accompa-
nied by an increase in nonresting EE from 850 kcal to 1314 kcal over
24 h (15.8 to 23.2 kcal/fat-free mass). The oxygen uptake during cycle
ergometry at 10 W increased with the 10% weight gain (from 0.26 to
0.41 L) but not at 50 W of power output. There was no adjustment
for the heavier legs which have a linear relation to oxygen uptake dur-
ing cycling at zero external work load.361 When the low-intensity
exercise EE was adjusted for resting metabolic rate, the net energy
cost increased from 1.50 to 1.83 kcal/min. An increase in RQ from
0.80 to 0.86 was also observed during steady state exercise at 10 W.
No change in ATP flux or the energy cost of muscle contraction could
be detected by magnetic resonance spectroscopy following the 10%
increase in body weight even though there was some indication of an
increase in muscle contraction
The most recent study of this problem is by Vinales and col-
laborators who investigated the topic of ergometer cycling effi-
ciency under a series of acute overfeeding conditions.172 Twenty
healthy subjects were enrolled in a crossover design in which they
were studied in the fasted state and after five different diets that
contained twice the amount of calories they consumed during
weight stable conditions. The overfeeding diets included: control
(50% carbohydrate, 30% fat, 20% protein), high carbohydrate
(75%), high fat (60%), high protein (30%) and low-protein (3%). Net
cycling efficiency from calorimetric data defined as the energy cost
of cycling minus the resting EE did not change with the any of the
five acute overfeeding conditions.
Support for a lack of chronic overfeeding on the energy cost and
work efficiency during low- to moderate-intensity exercise also comes
from the twins overfed for 100 days165 and is summarized in
Table S10. There was no change in caloric expenditure and RQ mea-
sured 2 h after a standardized meal under any of the resting and exer-
cise conditions defined in the table.
The preponderance of evidence thus appears to show that experi-
mental overfeeding and the associated weight gain are not causing
detectable changes in whole-body work efficiency under standardized
ergometry conditions and are not causing alterations in ATP fluxes or
energy cost of muscle contraction as measured by magnetic reso-
nance spectroscopy. This is in contrast with the effects of caloric
restriction and weight loss where both whole-body calorimetry during
standardized ergometry conditions and magnetic resonance spectros-
copy assessments of ATP flux and energy cost of muscle contraction
360
reveal increased skeletal muscle efficiency.
14.7 | Skeletal muscle characteristics and the
response to overfeeding
The topic of skeletal muscle characteristics prior to being exposed to
chronic overfeeding and the role they play in the responsiveness to
the overfeeding challenge has not been addressed to a meaningful
extent. Only one study could be retrieved on the subject matter, and
it is based on the long-term overfeeding study of 12 pairs of identical
57
twins.354 Overall the study found that baseline skeletal muscle oxida-
tive capacity (from the vastus lateralis muscle) was protective against
adipose tissue accumulation in response to overfeeding. For instance,
muscle type 1 fibre proportion was negatively correlated with the
gains in fat mass (r = −0.43) whereas muscle type 2A fibers were posi-
tively associated with the gains in fat mass (r = 0.43). Similar observa-
tions were made for the changes in body fat percentage with
correlations of −0.49 and 0.47, respectively. Maximal activity in vitro
of baseline muscle OGDH enzyme was negatively correlated with the
gain in fat mass (r = −0.47), whereas the ratio of OGDH to PFK activ-
ity was positively associated with the gain in adiposity (r = 0.46). In
the same 100-day overfeeding protocol among a broad set of baseline
measurements, a low skeletal muscle oxidative potential was among
the best predictors of high gains in body weight, adiposity or body
energy.34
In summary, in studies lasting from about 1 to 3 months, 40% to
50% of the weight gained is in the form of fat-free mass and is mainly
accounted for by an increase in muscle mass. Overfeeding does not
affect skeletal muscle fibre type distribution, capillary density or gly-
cogen content. Experimental overfeeding can cause an upregulation
of muscle extracellular matrix, fibrosis and inflammatory genes. Mus-
cle gene expression studies indicate multiple genes are up-regulated
or down-regulated with overfeeding. Muscle glucose uptake is
unchanged with overfeeding but the response to insulin infusion is
blunted with the weight gain. Insulin resistance in muscle with
overfeeding-induced weight gain appears to be similar to what is
observed in spontaneous obesity. Overfeeding and associated weight
gain do not seem to cause changes in whole-body work efficiency
under standardized exercise conditions and do not affect the energy
cost of muscle contraction. Low baseline skeletal muscle oxidative
potential appears to predispose to larger weight gain in response to
experimental overfeeding.
15 | R E C O V E R Y F RO M E X P E RI M E NT A L
O V E RF E E D I N G
Overfeeding results in a positive energy balance and with time
adds to the stores of fat in adipose tissue and possibly ectopic
sites. Following the termination of overfeeding the individual is
free to choose the quantity and quality of foods to eat. This tran-
sition period has been evaluated in after both short periods of
overeating and after longer term follow-up of individuals who have
been overfed over many weeks to many months. The self-
experimentation of one of the authors of this paper has been
recently chronicled and analysed.362
15.1 | Follow-up of changes after overfeeding
Studies on the recovery from prolonged overfeeding began to
appear during the 1990s. After gaining 10–25% of body weight by
overfeeding, the volunteers in the Vermont Study were reported
58
to lose weight rapidly and to return to nearly baseline over several
weeks.63 This has been found with other overfeeding studies, too.
In one report the author gained 10% of his body weight over
3 months and then lost that weight within 2 months and has
remained at the initial weight subsequently.46 This rapid weight
loss after overfeeding has been documented in several stud-
58,96,164,288 58
ies. In the overfeeding study by Norgan and Durnin
five of the subjects returned to the laboratory 3 months later for
measurements of body weight, body fat content and skinfold thick-
nesses. Three of the participants were at or below their initial
body weight; two of them were 1.3 and 1.9 kg above baseline
body weight. None of the subjects had made a sustained conscious
effort to lose weight. Roberts et al96 overfed young men for
20 days and at 10 days after overfeeding, with only one individual
consciously trying to lose weight, they were within 0.005 kg of
baseline weight although there with a wide variability in amounts
of weight lost.96 In the Guru Walla overfeeding study in nine
young Cameroonian men, body weight 2.5 years after the massive
weight gain of 19 kg, had returned to baseline.149 Another 2.5-year
follow-up study of 18 individuals (12 M/6F) aged 26 who
increased fast food intake by 70% for 4 weeks and kept activity at
<5000 steps/day gained 6.4 kg. Six months post-overfeeding their
weight was 68.6 kg, an increase of 1.0 kg over baseline. Twelve
months after stopping overfeeding they weighted 69.7 kg, an
increase of 2.1 kg over baseline. By 2.5 years, body weight had
increased further to 72.9 kg, an increase of 5.3 kg compared with
baseline (p = 0.018) and only 1.1 kg below their peak weight. Fat-
free mass increased during overfeeding but returned to baseline at
12 months. Fat mass, on the other hand, increased from baseline
and remained elevated.363–365 After 2.5 years post-overfeeding,
there was considerable variability in weight change which ranged
between −2 kg and +3 kg for four women and −3 kg and +13 kg
for 11 men. In the control group variability among 10 men
extended from −4 and to +4 kg (Figure 8).
Recovery of body mass, body composition and blood chemistry
variables in monozygotic twins who gained 8 kg was examined
4 months and 5 years later.288 After 4 months, the twins had lost about
7 kg, but by the 5 year of follow-up, the men were on average 5 kg
heavier than when they started the study. Interestingly, based on a
representative sample of the US population, the 50th percentile for the
gain in body mass from 19 to 24 years reaches 1.9 kg/year or about
9.5 kg over 5 years.366 Thus the post-overfeeding increase in body
mass for these young men can be defined as normal for free-living indi-
viduals in this age class. Fluctuations in fat mass were greater than for
fat-free mass. All laboratory and clinical chemistry data had returned to
normal by 4 months and remained normal. The most recent study with
post-weight gain data was from an 8-week overfeeding study by
Johannsen et al.164 Follow-up was obtained in 30 of the 35 volunteers
at the 6-month follow-up visit. Mean weight was 82.9 ± 11.6 kg, com-
pared with a baseline weight of 79.8 ± 10.8 kg which meant that
43 ± 63% of the weight gain produced by overfeeding was retained.
Individuals who had a lower-than-predicted sleeping metabolic rate
prior to overfeeding lost less overall weight during the follow-up
BRAY AND BOUCHARD
(r = 0.35) and retained a significantly greater portion of the fat gained
during overfeeding (r = −0.38). Conversely, a greater than-predicted
24 h-EE in a metabolic chamber after overfeeding was associated with
significantly greater loss of body fat 6 months later (r = 0.45). These
associations were independent of fat mass at baseline and the amount
of fat gained during overfeeding.
In a second 5 year follow-up of overfeeding, Rynders et al367
found that the ability to adjust nocturnal fat oxidation in response
to overfeeding predicted 5-year weight gain in adults. Lower noc-
turnal fat oxidation and lower energy expenditure with overfeeding
were the strongest predictors of 5-year weight gain. When
adjusted for covariates, changes in nocturnal fat oxidation and EE
with overfeeding predicted 41% of the variance in weight change
over 5 years.367
15.2 | Food intake after termination of overfeeding
Overfeeding increases energy storage, and changes the feelings
towards food in general, or specific types of food which may also be
conditioned by individual behavioural characteristics. Subjective feel-
ings of hunger and the desire for food can change significantly after
only 1 day of overfeeding or underfeeding.368 In a 3-day study com-
paring thin men and women with men and women who had been
reduced in weight, overfeeding reduced the feelings of hunger and
satiety after eating more in the thin individuals when compared with
individuals who were reduced from an obese state. After terminating
the overfeeding, the thin women experienced a 30% reduction in daily
energy intake, but there was no difference in thin men or in either sex
of the group who were reduced-obese.109 Using the three-factor eat-
ing inventory of Stunkard and Messick,369 Halliday et al271 reported
that adults who were prone to obesity had greater dietary disinhibi-
tion than adults who were resistant to obesity. Moreover, this greater
baseline disinhibition was associated with greater weight gain over a
period of 5 years of follow-up.
Several studies have examined the effect of short-term overfeed-
ing on subsequent energy intake. In one study, adults were overfed
one of three diets: a high-fat, low-energy dense diet (1.05 kcal/g); a
high-fat, high-energy dense diet (1.60 kcal/g); or a high-carbohydrate,
low-energy dense diet (1.05 kcal/g) for 2 days at 40% excess energy
using a crossover design.103 Food intake was followed for 4 days after
termination of overfeeding. Energy intake was not suppressed until
the second day after termination of overeating and then accounted
for only 30% of the excess energy. Reductions in energy intake did
not differ among the three diets or across days, and overeating had
no effect on subsequent EE. However, physical activity, as measured
by the number of steps walked each day decreased after the high-car-
bohydrate, low-energy diet and the high-fat, high-energy diet, but not
after the high-fat, low-energy dense diet. Sleeping time increased
after the high-fat, high-energy diet compared with both low-energy
dense diets. After overeating a high-fat, high-energy density diet, car-
bohydrate cravings, hunger, prospective food consumption and sad-
ness increased whereas satisfaction, relaxation and tranquility
BRAY AND BOUCHARD
F I G U R E 8 Twelve men and six
women were overfed by 70% for 4 weeks
with a fast food diet. An age-sex matched
control group of 18 individuals was not
overfed. Both males and females
increased weight significantly. At
6 months post-overfeeding, females were
still significantly heavier than at baseline,
but not at 12 or 30 months. Men also
increased their weight significantly with
overfeeding but they were not different
from baseline at 6 months. At 12 months
their weight was “borderline” (p = 0.052)
increased and significantly increased at
30 months. Drawn from the data of
Ernersson et al.363
decreased compared with the high-fat/low-energy dense diet.103
These data suggest that short-term overeating is associated with
incomplete short-term compensation for excess energy.
In another study, energy intake during the 3 days of ad libitum
food intake that followed 3 days of overfeeding was significantly
reduced in the individuals resistant to obesity, but not in those prone
to obesity. This suggests that those resistant to obesity may be able
to compensate better for the energy excess of overfeeding than indi-
viduals who are prone to obesity.271
In a final short-term study, He et al117 found that overfeeding for
3 days compared with a 3-day weight maintenance period did not
result in a decrease in subsequent ad libitum food intake, core body
temperature or overall change in sedentary time.
Cognitive restraint is another mechanism for rebalancing energy
80
intake and expenditure after overfeeding. Jebb et al concluded that
after overfeeding, the appetite control mechanisms, on which we rely
to regulate energy balance, are inadequate, emphasizing the role of
intentional cognitive restraint as a mechanism for controlling food
intake. In their study, there were three periods of 3 weeks each with
overfeeding increasing from +20% to +40% to +60% separated by
1 week of ad libitum diet. There was considerable individual variability
of intake during ad libitum diet with some men called “compensators”
compensating with a significant drop of intake in each period and
other men called “noncompensators” who failed to reduce energy
intake sufficiently to make up for the previous excess. This variable
response of appetite control mechanisms after overfeeding leads the
authors to emphasize that some people need to adopt intentional cog-
nitive restraint when food is plentiful if they are to control body
weight.80
The possibility that different macronutrients might have differ-
ential effects on subsequent adaption to overfeeding has also been
tested.120 In a cross-over study, six men were studied on three
times using a 5-day protocol. On Days 1 and 2, they were fed a
59
medium-fat weight-maintenance diet with energy calculated at
1.6× resting metabolic rate. Subjects entered a room metabolic cal-
orimeter for 48 h at 08.00 on Day 3. On Day 3 they ate a
medium fat diet at 1.5× resting metabolic rate and received an
additional 0.6× resting metabolic rate as protein, or carbohydrate
or fat. On days 4 and 5, (outcome days), subjects had ad libitum
access to an isoenergetically dense medium-fat diet. Subjective
hunger and satiety were tracked hourly during waking hours
throughout Days 1−5. Subjects felt significantly fuller and less hun-
gry on the high-protein diet than the other two diets. By the end
of the test Day 3, each overfed nutrient led to a significant
increase in its own balance relative to the other two diets. These
effects did not influence the subsequent day's energy intake. These
authors concluded that higher protein diets were more satiating
than isoenergetically-dense higher carbohydrate or higher fat diets
on the day they are eaten. The higher carbohydrate diet was tran-
siently more satiating than the higher fat diet after each meal. This
study shows that in men relatively large changes in nutrient bal-
ance on 1 day appear to be poorly compensated by changes in
energy intake on a subsequent day.120
The relative lack of precision in control of food intake after over-
feeding was also reported when ad libitum intake of food was mea-
sured before and following 13 days of overfeeding by 35% which
produced a weight gain of 2.3 kg.370 In the ad libitum periods, individ-
uals were given a 1200 kcal/day diet with additional food eaten ad
libitum. Following overfeeding, mean daily caloric intake was not sig-
nificantly suppressed, returning only to baseline values. Despite nor-
mal energy intake, participants lost half the weight they had gained.
These results indicate that the physiological control of eating behav-
iour in humans is not the only mechanism responsible for the recovery
of body weight following a period of overfeeding as an increase in EE
of about 14% was required to account for the weight loss following
overfeeding.370
60
15.3 | Physical activity after termination of
overfeeding
Spontaneous physical activity might be one explanation for living in
an environment that promotes weight gain and yet not gaining
weight.133 To test this idea, a group of healthy men and women ages
25–35 years who were empirically classified as either resistant to obe-
sity or prone to obesity based on personal and family weight history
were recruited. Using a physical activity monitoring system that mea-
sured body position and movement combined with pedometers and
accelerometers while subjects were awake, either in a controlled
eucaloric condition or during 3 days of overfeeding (1.4 × basal
energy), and for the subsequent 3 days (ad libitum recovery period),
changes in physical activity were assessed. Although there were no
differences between groups when examined across all study periods,
3 days following overfeeding, subjects prone to obesity significantly
decreased the amount of time they spent walking, whereas subjects
who were resistant to obesity maintained their walking.133 Thus, there
may be a regulatory system for physical activity level over time similar
to the one suggested for regulation of food intake.371
In summary, almost all participants who are overfed return or
nearly return to their baseline weight over a matter of a few weeks to
months, but there is considerable variability in the response to
reduced caloric intake. The differences between individuals who are
prone to obesity and those who are resistant to obesity offer opportu-
nities to unravel some of the variability in this response. The period
after overfeeding could also provide a fruitful area for studies of brain
imaging.
1 6 | G E N E T I C A ND E P I G E NE T I C F A C T O R S
I N F L U E N C I N G TH E R E S P O N S E T O
OVERFEEDING
In this section, we review the evidence for the contribution of geno-
mic variation and epigenetic events in the adaptation to chronic over-
feeding. Even though the evidence available is limited, some lessons
can be derived from the research reported to date. With the excep-
tion of rare or low-frequency cases in which genetic mutations with
very large effect sizes could condition the responsiveness to over-
feeding, a high degree of predictability cannot be achieved regarding
the role of genetics and epigenetics. A probabilistic paradigm is more
likely and, as in all common human traits, the response to chronic
overfeeding is likely to be entrained by complex contributions of DNA
variants, changes in gene expression and epigenetic events.
For ethical reasons, the role of specific genes with large effect
sizes has not been investigated in the context of an experimental
overfeeding paradigm. Thus studies of chronic overfeeding on sub-
jects with leptin or melanocortin 4 receptor (MC4R) deficiencies have
not been undertaken. Most overfeeding studies involve adults with
normal body weight at recruitment, although some studies have
included individuals with obesity.70,71,82,196,212,223,226,295 In studies on
individuals of normal weight where DNA sequence variants with large
BRAY AND BOUCHARD
effect sizes are unlikely to be contributing to the response pattern.
Such studies favour by design genetic contributions resulting from
large numbers of genes and noncoding DNA variants, each with small
to very small effect size.
16.1 | Genetic epidemiology of responsiveness to
overfeeding
One condition needs to be met before launching complex and costly
genetic studies of the response to experimental overfeeding: human
heterogeneity in responsiveness must be established. We have dis-
cussed this topic earlier in Section 5. Using only the studies in which
the caloric overload reached at least 50 000 kcal with excellent con-
trol over caloric consumption and physical activity level during the
overfeeding protocol,33,44,47,56,58 we found, on average, a threefold
range between the lowest and the highest body weight gainers, with a
range extending from 1.8- to 5.1-fold (see Table 5). Thus, there is
human variability in response to experimental overfeeding, and this
variability cannot be accounted for by problems of compliance or
adherence to the intervention. Therefore, one is justified in asking
whether human genetic variation is playing a role in adaptation to
overfeeding.
To test whether there is a genetic component, one can rely on
the methods and designs of genetic epidemiology. For overfeeding,
there are only two studies conducted with pairs of identical twins and
none with nuclear family or adopted offspring with siblings and foster
parents. The first study was conducted with six pairs of young male
monozygotic twins who were overfed by 1000 kcal for
22 days.94,152,160,253,334 Significant within pair resemblance relative to
between pair differences was observed for body weight, body compo-
sition and for several physiological and metabolic indicators
suggesting that there may be a genetic component contributing to
human variability in adaptation to moderate overfeeding. However,
this study was limited by the fact that the caloric excess was moder-
ate and induced a weight gain of only about 2 kg in a very small num-
ber of twin pairs.
These problems were partly addressed in a second overfeeding
experiment performed with 12 pairs of monozygotic twins.44 The
24 young men were kept under a controlled environment for
120 consecutive days under constant supervision. They were studied
by groups of four pairs of twins at a time over a 2-year period to
allow for closer supervision. During the first 2 weeks, subjects were
asked to maintain a constant body weight, and all the food that they
ate was carefully quantified. Body weight was measured daily and
body composition was assessed several times during these 2 weeks.
Subjects were kept sedentary during the whole period of 120 days.
After the initial 2 weeks, the energy cost of weight maintenance was
calculated and the overfeeding protocol begun. Subjects consumed
an extra 1000 kcal per day, 6 days a week. On the 7th day, they con-
sumed the baseline weight maintenance diet. All prescribed diets had
the following composition: 50% carbohydrate, 35% lipid and 15%
protein. The total overfeeding load reached 84 000 kcal per subject.
BRAY AND BOUCHARD
Correlations between the total energy consumed and body weight
and adiposity were not different from zero,44 indicating that the
energy cost of weight maintenance had been appropriately
quantified.
Figure 9 depicts the changes in body weight (left panel) and com-
puted tomography-assessed visceral adipose tissue adjusted for the
changes in total fat mass (right panel). Each black dot in the figure is a
pair of monozygotic twins. There was 3.4 times more variability
between pairs of twins compared with the variability between
brothers of the same pair (intraclass correlation of 0.55). The twin
resemblance was even stronger for the changes in visceral adipose tis-
sue adjusted for the gains in fat mass with six times more variance
between pairs of twins compared with the within-pair variability
(intraclass correlation of 0.72). These findings are strongly suggestive
of a significant genetic component in the adaptation of body weight
and preferential fat deposition to long-term overfeeding.44 Similar
observations were made for the gains in total adiposity (F ratio = 3.0;
intraclass correlation = 0.50) and total body energy gain (F ratio = 3.12;
intraclass correlation = 0.51). In contrast, there was no twin pair
resemblance when changes in resting metabolic rate were reported
per unit of kg weight or kg fat-free mass or in net thermic effect of
food.3 No studies have attempted to replicate these findings.
From the same chronic overfeeding study, it was shown that
there was a significant within twin pair resemblance in response to
the caloric overload for fasting insulin (intraclass correlation = 0.71),
fasting glucagon (0.53) and fasting glucose (0.69). During an oral glu-
cose tolerance test, area under the curve was increased after over-
feeding for insulin, but not glucose. Following a 4-h standardized
1000 kcal meal test the area under the curve was increased for both
insulin and glucagon.269 The same overfeeding protocol generated sig-
nificant within-pair resemblance for the changes in plasma cholesterol,
triglycerides, VLDL-cholesterol, VLDL-triglycerides, LDL-triglycerides,
HDL-cholesterol, HDL-apo A1, HDL2-cholesterol, HDL3-cholesterol,
with intraclass correlations ranging from 0.48 to 0.85.300 Within twin
pair similarity in response to overfeeding was also observed for
changes in plasma T4 (intraclass correlation = 0.34) and free T4 (0.44)
thyroid hormone concentrations.250 In contrast, no twin resemblance
F I G U R E 9 Changes in body weight
(left panel) and computed tomography-
assessed abdominal visceral fat adjusted
for the gain in total fat mass (right panel)
in 12 pairs of monozygotic twins in
response to a 100-day overfeeding
protocol which delivered an excess caloric
intake of 84 000 kcal. Each dot represents
a pair of twin brothers. Reproduced from
Bouchard et al44 by permission of the
Massachusetts Medical Society
61
was found for the changes with overfeeding in plasma adiponectin
concentration,351 and there was no significant effect on plasma
ghrelin levels.350 Among a large panel of plasma steroids including
conjugated steroid as well as steroids originating from the adrenal
gland and gonad, the only overfeeding-induced changes characterized
by a within-pair resemblance were in sex hormone binding globulin
(intraclass correlation = 0.55) and dehydrotesterone (0.48).260 Isolated
abdominal fat cells were studied for their lipolytic rates in the same
twin overfeeding experiment.315 Both basal and epinephrine stimu-
lated lipolysis in response to the 84 000 kcal overload were character-
ized by significant twin resemblance (intraclass correlations ranging
from 0.60 to 0.69) but not isoproterenol stimulated lipolysis. Skeletal
muscle fibre type distribution of the vastus lateralis muscle was not
influenced by the long-term overfeeding protocol, but a significant
increase was seen in muscle creatine kinase activity, a decrease in the
activity of the muscle enzyme OGDH and an increase in the PFK to
OGDH ratio with no evidence of twin resemblance in the response of
these muscle traits.354
Four months after overfeeding stopped there was a very strong
twin resemblance in weight loss (F ratio = 8.4; intraclass correla-
tion = 0.79) which was still significant 5 years later (F ratio = 2.6;
intraclass correlation = 0.44). Similar trends were seen for the post
overfeeding changes in percentage body fat and visceral adipose tis-
sue as well as in plasma glucose, insulin, cholesterol, HDL-cholesterol
and trigycerides and blood pressure.
16.2 | Associations with specific DNA variants
Studies on the contribution of specific DNA variants to adaptation to
chronic overfeeding are challenging as the sample size of controlled
overfeeding experiments is typically small. Testing for the effects of
variants in candidate genes has been undertaken on the participants
of only one overfeeding study and the findings are summarized here.
As a hypothesis-generating exercise, DNA variants in 40 candidate
genes were genotyped in 12 pairs of monozygotic twins who were
exposed to an overfeeding protocol for 100 days. The findings were
62
reported in multiple papers301–305,307,372–375 and were summarized
by classes of adiposity and metabolic traits in a review.376 A marker in
adrenergic beta 2 receptor (Gln27Glu) was associated with higher
body weight and higher gains in adiposity. Polymorphisms in the
glucorticoid receptor and adipsin gene loci were also consistently
associated with changes in weight and adiposity traits (fat mass and
visceral adipose tissue). The adipsin (complement factor D) locus
encodes a protease catalysing the rate limiting step of the alterna-
tive pathway of the complement activation and is as an adipokine
secreted by the adipose tissue. An adipsin polymorphism was asso-
ciated with visceral adipose tissue changes in response to overfeed-
ing. In a study of UCP-2 and UCP-3, the A55VAV genotype of UCP2
and the RsaI CC genotype of UCP3 had a greater increase in TSH in
response to a challenge with TRH than those with the AA or CC
375
genotypes.
Variants in IGF1, IGF2 and two binding protein genes, IGFBP1 and
IGFBP3, were also genotyped. Among those with the ApaI GG geno-
type for IGF2, fasting plasma insulin was higher and increased more
during an oral glucose challenge than the other genotypes. After over-
feeding, men with the IGFBP1 Bgl II AA variant had a greater increase
in visceral fat, insulin response during a glucose tolerance test and
total cholesterol than the other genotypes for IGFBP1.373 Insulin sen-
sitivity decreased in the subjects with IGF2 Apa I GG variant, and the
subjects with IGFBP1 Bgl II AA variant showed an accumulation of vis-
ceral adipose tissue. Two variants of the glucocorticoid receptor gene
influenced weight gain in the twin study.307 Overfeeding induced a
greater increase in body weight in the 12 men with the Bcl I cleavage
2.3/2.3 kb variant than in the 12 men with the 4.5/2.3 kb variant. In
addition, plasma levels of total cholesterol, LDL cholesterol and sys-
tolic blood pressure increased more in the 2.3/2.3 kb men than in the
4.5/2.3 kb men. Visceral adipose tissue also increased more in the
2.3/2.3 kb genotype than in the 4.5/2.3 kb genotype.
Variants in the cholesteryl ester transfer protein (CETP) gene
locus were associated with more adiposity and visceral fat gains
301
and with larger decreases in HDL, its fractions and apoA1. Vari-
ants at the LPL locus might affect body composition and lipid and
lipoprotein changes with overfeeding. Those with the LPL Hind III
polymorphism H2H2 had higher levels of VLDL-triglycerides and
VLDL-cholesterol before and after overfeeding as well as higher
HDL-triglycerides but lower HDL-cholesterol levels after overfeed-
ing. In addition, HDL2-cholesterol levels were lower in these indi-
viduals and their HDL2-cholesterol as well as the HDL2-C/HDL3-C
ratio decreased with overfeeding. In the twins with the other
genotypes (H1H1 or H1H2) HDL2-cholesterol increased slightly
together with a decrease in HDL3-cholesterol resulting in an
increase in the HDL2-C/HDL3-C ratio. These observations support
the notion that the men with the H2H2 genetic variant had
reduced catabolism of VLDL in response to overfeeding. A DNA
variant in intron 8 of the LPL gene was found to be associated
with the changes in adipose tissue LPL activity.304 Twins with the
GlnGln genotype at the leptin receptor (LEPR) Gln223Arg polymor-
phism experienced more profound adverse plasma lipid changes
than the other genotypes.376
BRAY AND BOUCHARD
In summary, these findings provide a series of candidates that
would benefit greatly from being explored further in experimental
overfeeding but with much larger sample sizes. They are catalysts for
hypothesis-driven studies.377 However, the pursuit of research on the
genetics of adaptation to sustained overfeeding should not be limited
to candidate genes of adaptation to excess nutrients and calories but
should rather be driven by unbiased genome-wide screens and
transcriptome-wide and other omics explorations in appropriate
tissues.
16.3 | Overfeeding and gene expression
The effects of chronic overfeeding on the gene expression profile of
peripheral blood mononuclear (lymphocytes and monocytes) cells, adi-
pocytes, skeletal muscle or on the microbiome population distribution
have been investigated in a few studies. The findings on gene expres-
sion in adipose tissue and skeletal muscle have been summarized in
previous sections. Here, we comment briefly on the findings related
to lymphocytes and monocytes and to the microbiome.
Blood mononuclear cells were isolated in 23 healthy adults before
and after a 30-day overfeeding protocol with an excess intake of
880 kcal/day, on average, leading to a mean weight gain of 2.8 kg.231
Despite the modest weight gain, there were 318 blood mononuclear
cell transcripts whose expression levels were significantly altered with
overfeeding; notably, this set of transcripts included genes associated
with lipid metabolism (e.g., LPL) and inflammatory response but also
with dilated cardiomyopathy. In the same study, changes in the micro-
biome were explored using 16S and whole-metagenome sequencing
from stool samples acquired before and after overfeeding. A signifi-
cant increase in the population of Akkermansia muciniphilia was
observed in response to overfeeding but only in the subsample of
subjects who were classified as insulin sensitive.
To-date, there is a dearth of data on the potential role of adipose
tissue and skeletal gene expression profile at baseline during the weight
maintenance stage on the response level to experimental overfeeding.
16.4 | DNA methylation and the response to
overfeeding
Only one report has dealt with the impact of the preoverfeeding
methylome on the response to overfeeding.297 From adipose tissue
biopsies obtained in 31 normal weight adults who were overfed for
7 weeks by 750 kcal/day with diets rich in either saturated fatty acids
(N = 17) or polyunsaturated fatty acids (N = 14), DNA methylation at
461 450 CpG sites, annotated to 20 899 gene loci, were studied for
their impact on abdominal adipose tissue gene expression. A total of
12 CpG sites (annotated to nine genes) were identified where the
methylation status was associated with the percentage increase in
body weight when both dietary groups were combined (mean weight
gain of about 1.6 kg or about 2.5%). These methylated sites were
related to the following loci: CUB and Sushi multiple domains
BRAY AND BOUCHARD
1 (CSMD1), melanin concentrating hormone receptor 1 (MCHR1),
mitogen-activated protein kinase 7 (MAPK7), FCH and double SH3
domains 1 (FCHSD1), transient receptor potential cation channel sub-
family C member 4 (TRPC4), PMS1 homolog 2, mismatch repair sys-
tem component pseudogene 5 (PMS2L5), stromal antigen 3-like
2 (pseudogene) (STAG3L2), replication initiator 1 (REPIN1) and splicing
factor SWAP (SFRS8). All methylated CpG sites were positively related
to weight gain except the site in the proximity of SFRS8 which had a
negative association pattern.
This is clearly a promising area that warrants further research
incorporating DNA methylation as well as histone modifications.
16.5 | Effects of overfeeding on DNA methylation
events
A few reports have dealt with the effects of chronic overfeeding on
the DNA methylation profile in cells from adipose tissue or skeletal
muscle. Abdominal fat cell DNA methylation was altered at 652 CpG
sites after 5 days of high-fat overfeeding resulting in no detectable
weight gain in 40 participants.293 Loci potentially impacted by these
methylation events included cyclin dependent kinase 5 (CDK5), insulin
like growth factor binding protein 5 (IGFBP5) and solute carrier family
2 member 4 (SLC2A4). In 31 subjects exposed for 7 weeks to either a
dietary excess of saturated or polyunsaturated fatty acids, the DNA
methylation response profile of abdominal fat cells differed between
297
the two hypercaloric diets. Whereas the methylation profile at
1797 genes was changed by the hypercaloric polyunsaturated fatty
acid diet, it was altered at only 125 genes with the saturated fatty acid
diet. Moreover, while the polyunsaturated fatty acid diet did not influ-
ence gene expression level in abdominal adipose cells, the saturated
fatty acid diet changed significantly the expression levels of
28 transcripts.
One group has reported three papers on DNA methylation at
targeted sites from muscle biopsy samples following exposure to
5 days of high-fat overfeeding and 5 days of a control diet (in a ran-
dom order) interspaced by a washout period of 6 to 8 weeks.104,181,294
In the first study, high-fat overfeeding reduced muscle PPARGC1A
and OXPHOS gene expression by about 25% in low birth weight
(N = 20) compared with normal birth weight (N = 26) participants.104
DNA methylation in the promoter region of the PPARGC1A was inves-
tigated at four CpG sites. Methylation increased with fat overfeeding
but only in the normal birth weight subjects. The high-fat
overfeeding-induced changes in the methylation profile of the
PPARGC1A promoter were shown to be reversible in the normal birth
weight subjects when those who were initially assigned to the high-
fat overfeeding condition shifted to the control diet after the washout
period. In the next study, DNA methylation was assessed at 27 578
CpG sites covering 14 475 genes in 21 young men using muscle sam-
ples obtained under the same protocol.294 High-fat overfeeding
induced methylation changes at more than 6500 genes (45%). The
alterations in the methylation profile with high-fat overfeeding
impacted genes enriched in inflammation, the reproductive system
63
and cancer. Muscle gene expression levels were assayed for 13 candi-
date genes for type 2 diabetes. No change in gene expression was
observed with either condition, and there was no consistent associa-
tion between the DNA methylation signatures at these candidate
genes and their expression levels. Interestingly the changes in short-
term overfeeding methylation profile were only partially reversed
after 6 to 8 weeks of recovery. In their last report, the same group
compared the DNA methylation profile between low and normal birth
weight groups under the same protocol.181 While widespread methyl-
ation changes were identified in response to high-fat overfeeding in
normal birth weight subjects, only a few methylated sites were identi-
fied in the low birth weight group. It was suggested that men with low
birth weight may have less plasticity in muscle adaptation to short-
term (5 days) high-fat overfeeding compared with normal birth weight
individuals.
In summary, from limited experimental overfeeding studies per-
formed with pairs of identical twins, it is strongly suggested that there
is a genetic component accounting for some of the response variabil-
ity in weight, fat mass, fat-free mass, visceral fat gains. The same stud-
ies that the metabolic alterations observed in response to
experimental overfeeding were also influenced by genetic differences.
Very limited data have been reported on the potential role of a few
candidate genes. The preoverfeeding global methylome seems to
associate with the propensity to gain more or less weight. Even
though the evidence is still limited, experimental overfeeding seem to
exert an influence on the methylation profile at multiple genes in adi-
pose tissue and skeletal muscle.
17 | D I S C U S S I O N
This paper has examined the response to short- and long-term over-
feeding from a number of perspectives driven by the findings from
more than 150 human studies of experimental overfeeding. Figure 10
provides an overview of the various aspects of the biology of human
overfeeding protocols that provided a foundation for the present
review. We now offer a brief discussion, draw several conclusions and
identify areas where additional studies are needed.
We have organized the numerous overfeeding studies into
three categories based on the duration of the overfeeding proto-
cols. There are clear differences on what can be accomplished by
short term and long-term studies. For instance, studies lasting
7 days or less are better suited to investigate issues related to
acute exposure to excess calories or excess of a given macronutri-
ent composition. In contrast, studies of long durations have the
potential to generate useful information on the effects of both
excess nutrients and calories plus body weight and body composi-
tion changes when the post overfeeding measurements are per-
formed a day or two post overfeeding. A third option, which is
less frequently used, incorporates the post overfeeding measure-
ment panel only after weight stability has been achieved at the
new body weight. This design allows focusing on the true conse-
quences of weight gain.
64
The relationship between the number of calories eaten and
weight gain is very strong (r2 of 0.82) across 19 studies meeting sev-
eral inclusion criteria. During the early phase of overfeeding, most of
the energy surplus consumed is recovered as body energy gain, mainly
as fat, whereas gains in body mass represented only about 60% of the
overfed calories with long-term overfeeding. The weight gain tends to
slow over time under an exponential model with a half-time of
86 days. The difference between the lowest and highest weight
gainers across several well-controlled experimental overfeeding
groups reaches threefold, with reduced heterogeneity in response to
differing protein content in the caloric overload. The gain in lean tis-
sues is generally less than the increase observed for fat mass, rep-
resenting about 30% to 40% of the weight gain depending on the
overfeeding protocol. Importantly, low-protein overfeeding does not
lead to significant increases in fat-free mass.
The potential contributions of age, gender and ethnicity to the
variability in body weight and changes in body composition with
sustained overfeeding has had limited investigation. In one report,34
the most consistent correlates of weight, adiposity and body energy
gains were baseline low fat-free mass, low skeletal oxidative potential,
low cardiorespiratory fitness, low plasma androgen levels and high
plasma leptin levels. Less powerful predictors included large mean
abdominal fat cell size, low postprandial EE, high plasma oestrogen
levels, a poor response of TSH to a TRH challenge and low plasma cor-
tisol and dehydroepiandrosterone. Although preliminary, these obser-
vations support the hypothesis that there is a biology underlying the
variability in response to human overfeeding.
Metabolic rates, total daily EE and substrate oxidation are of para-
mount importance for our understanding of the response to experi-
mental overfeeding. Although, there are still several questions to be
addressed, a number of generalizations emerge from this systematic
review. There is a strong trend for overfeeding lasting about 3 weeks
and more, with a substantial caloric excess, to result in an increase in
resting EE when expressed in kcal/day but not when expressed by kg
BRAY AND BOUCHARD
of weight or kg of fat-free mass. One topic that deserves attention is
whether the increase in resting metabolic rate is entrained purely by
the overfeeding-induced increase in body mass or whether there are
tissue- and organ-specific increases in metabolic rate caused by the
excess caloric intake. In addition the role of exercise needs more eval-
uation. With advancing age, the rise in resting metabolic rate in the
older men during overfeeding was marginally lower in response to
overfeeding than in younger men. There are no comparative studies
of overfeeding in women of different ages who have different propor-
tions of fat mass to fat-free mass, as well as differences in body fat
distribution, and such data could provide valuable insights into the
biology of overfeeding. Of interest would be further study on the
effects of variable macronutrient composition, including high levels of
fructose, on sleeping metabolic rate given recent data on elevation of
sleeping EE when overfed by high fructose corn syrup.209
Postprandial EE over several hours tends to increase in response
to a standardized meal of mixed composition but the increase is
driven by the gain in resting metabolic rate primarily entrained by the
gain in body mass resulting from overfeeding. When the net thermic
effect of food is calculated, the changes induced by overfeeding are
generally nonsignificant. There is no consistent evidence for a role of
exercise as an enhancer of the thermic effect of food. Additional
research is needed on the possibility that the macronutrient composi-
tion of the test meal may lead to variation in food-induced thermo-
genesis in response to overfeeding.120 The differences in response to
overfeeding with saturated fats versus polyunsaturated fats clearly
deserves further exploration as well.
The EE associated with physical activity level when subjects are
challenged by an overfeeding protocol is difficult to investigate. On
the one hand, if subjects are studied in an inpatient setting, their level
of activity is constrained and no change in activity would be expected
by design. On the other hand, when participants are studied in an out-
patient context, the control over intake and expenditure is less strin-
gent and small changes in physical activity level can go undetected
F I G U R E 1 0 The increased energy
intake from overfeeding is partially
buffered by associated changes in energy
expenditure. Overfeeding has a variety of
effects on energy stores and adipose
tissue, endocrine and metabolic
responses, expression of genes and tissue
and plasma levels of proteins, lipids and
other metabolites. DNA sequence
variants and epigenetic events exert
downstream effects on tissue levels of
gene expression. The protein content of
the caloric overload plays an important
role in the response to experimental
overfeeding. See text for additional
information
BRAY AND BOUCHARD
with current technologies. Short-term overfeeding studies may not be
adequate for addressing the question of whether chronic overfeeding
167
has a direct impact on the level of physical activity. The energy cost
of various postures and low to moderate intensity exercise increases
slightly with exposure to chronic overfeeding when the EE is reported
in kcal per unit of time. However, the small increasing effect of over-
feeding disappears when the energy costs are expressed per unit of
body mass. Total daily EE increases, particularly with exposure to
long-term overfeeding. Increases of the order of 300 to 500 kcal per
day have been found based on doubly-labelled water measurements.
The increase is substantially less when EE is quantified in the confines
of a metabolic chamber. The increase in daily EE is attenuated when
the overfeeding protocol includes a low-protein diet. There is evi-
dence that combining an exercise regimen with a defined overfeeding
protocol reduces the impact of the caloric overload on body weight,
fat mass, fat-free mass and metabolic changes. This is an area deserv-
ing further research with a focus not only on various forms of exercise
regimens but also deprivation of exercise when exposed to experi-
mental overfeeding.
One of the most discussed topics over the last century is whether
there are energy dissipation mechanisms in the presence of “luxus
consumption” or excessive food intake. A review of the topic by one
of us (GAB) more than 20 years ago asked whether luxus consumption
was myth or reality and the conclusion was that the notion of energy
dissipation mechanisms when humans were challenged by overfeed-
ing was a myth.4 The view that such mechanisms exist in humans have
been fuelled by studies indicating that these mechanisms are found in
rodents.378 The major site of the overfeeding-induced extra thermo-
genesis in rodents resides in activated brown adipose tissue via the
stimulation of the sympathetic nervous system and an uncoupling
36
protein in this tissue. However, in humans, the research on a role
for overfeeding-induced stimulation of the sympathetic nervous sys-
tem is generally not supportive of a role of brown adipose tissue in
combatting the weight gain caused by the excessive consumption of
calories. For instance, the increase in resting metabolic rate caused by
overfeeding is similar between subjects who were lean or obese in
response to norepinephrine infusion.82 In an experiment in which five
men were subjected to an excess of 1600 kcal per day, treatment with
propranolol, a drug which blocks sympathetic stimulation did not pre-
vent an increase in resting metabolic rate of about 15%.100 These data
support the notion that the overfeeding-induced increases in resting
metabolic rate in humans are not mediated by augmented beta-
adrenergic activity but are rather predominantly the consequence of a
higher body weight and more metabolically active tissue.
Other relevant observations include the fact that the increase in
EE is substantially larger when adult participants (normal weight or
overweight) gain 10% of their baseline weight than the lowering of EE
seen when they lose 10% of their baseline weight.55 The defence of
body weight engages more powerful counter regulatory mechanisms
with weight loss than with weight gain. While it is important to recog-
nize that a strong defence against body weight gain under conditions
of standardized experimental overfeeding is not supported by solid
evidence at this time, it is also useful to appreciate that there are
65
individual differences in weight gain even when the overfeeding pro-
tocol is conducted in an inpatient setting and under highly standard-
ized conditions.33,44 Our inability to identify energy wasting
mechanisms triggered by human overfeeding could be in part caused
by challenges associated with measurement errors and day-to-day
subject variability at multiple levels. It could also be a consequence of
such mechanisms being characterized by small effect sizes poorly cap-
tured by current measurement programs and technologies.
What could be the sites of variability in energy dissipation mecha-
nisms in response to chronic overfeeding, if they exist? One potential
site resides in the coupling efficiency of mitochondrial oxidative phos-
phorylation to generate ATP. With less than perfect coupling, protons
leak across the mitochondrial membrane resulting in energy dissipa-
tion.379 Even in the absence of UCP1, coupling efficiency reaches only
about 90% for stages I and III of mitochondrial respiration. Efficiency
is lower for stage IV with values around 75%. Another mechanism
could be energy losses associated with substrate cycling when there
is an excess of ATP consumed compared with the expected energy
cost of the cycle, the so-called “futile cycles.”380 Recently, it has been
reported that there are more than 100 000 substrate cycles operating
in mammalian cells that consume ATP with a potential for energy dis-
sipation.381 Even though estimates are that energy losses for any
given substrate cycle would be quite small, in the aggregate the
potential for energy dissipation and variable efficiency via this mecha-
nism cannot be discarded. One cannot also exclude the possibility that
subtle differences in brown adipose tissue activation or in beige adi-
pose tissue could lead to small but biologically meaningful energy dis-
sipation, particularly with long term overfeeding when the rate of
body mass gain begins to slow down. However, at this time, there is
no evidence that these various mechanisms or any other energy dissi-
pation pathways are activated in response to chronic overfeeding in
order to increase metabolic rates and attenuate body energy gains to
compensate the extra calories consumed. This discussion is consistent
with the fact that blood levels of leptin change in overfeeding studies
as a function of change in body fat.
The bulk of the evidence suggests that chronic overfeeding
increases the rate and the absolute amount of carbohydrate oxida-
tion whereas the rate and absolute amount of lipid oxidation
decreases. This is observed under resting conditions as well as in
postprandial state after ingestion of a meal test. The contribution
of protein to the mixture of substrates oxidized in response to
overfeeding is less clear and seems to be quite variable. There is
also evidence for the presence of lipogenesis from glucose in the
fasted state following exposure to overfeeding, especially carbohy-
drate overfeeding.
A low-protein diet increases the percentage of energy stored as
fat. The change in visceral adipose tissue with overfeeding is related
to fat cell size at baseline, with individuals with larger fat cells storing
more visceral adipose tissue.177 The quality of the fat in the diet also
plays a role in the response to overfeeding. Eating a diet enriched with
saturated fatty acids markedly increased liver fat compared with a diet
supplemented with polyunsaturated fatty acids. The saturated fatty
acid diet also caused a twofold larger increase in visceral adipose
66
tissue compared with one enriched with polyunsaturated fatty acids
which also caused a nearly threefold larger increase in lean tissue than
seen in those eating the saturated fatty acid SFA diet. The increase in
liver fat is directly correlated with changes in plasma saturated fatty
acids and inversely with polyunsaturated fatty acids.62 Individuals
overeating in frequent episodes also showed a greater increase in
190
ectopic fat deposits than those overeating less frequently.
A family history of diabetes is recognized as a major risk factor
for weight gain. Concordant with this observation, insulin resistance
affects the response to overfeeding. Individuals who are insulin resis-
tant or are defined as metabolically abnormal or have a family history
of diabetes, show undesirable metabolic responses to overfeeding
including in adipose tissue gene expression profiling. Individuals with
low birth weight are known to have an increased risk of diabetes. The
altered metabolic profile in these individuals in response to overfeed-
ing, in comparison with individuals with normal birth, provides some
insights into the mechanisms for this increased risk level. These differ-
ences expand the hypothesis of Dulloo et al9 that a low protein diet is
a useful tool to unmask susceptibility to obesity suggesting that the
metabolic response to overfeeding can be a tool to identify suscepti-
bly to future weight gain.367
Many studies of overfeeding have focused on changes in the
endocrine system. A common observation is that weight gain pro-
duced by overfeeding recapitulates the endocrine changes seen in
spontaneous obesity.63 Suppression of growth hormone occurs early
in adaptation to overfeeding and it is accompanied by hyperlipidaemia
and insulin resistance. The thyroid hormone profile is modified with
overfeeding. Metabolic clearance of norepinephrine and neural activ-
ity of sympathetic nerves suggest that overfeeding has a stimulatory
effect on sympathetic nervous system activity. Contrary to expecta-
tions, overfeeding does not alter the gastrointestinal hormone profile.
Little has been reported on the effects of overfeeding on the brain.
This is an area in need of research with the most advanced technolo-
gies of neuroscience.
Changes in cardiovascular and haemodynamic traits are seldom
observed with a few days of overfeeding and remain generally small
with weeks of overnutrition. Alterations in heart rate and blood pres-
sure are not consistently found. People with above average cardiore-
spiratory fitness tend to prevent the effects of overfeeding on the
cardiovascular system better than those with lower cardiorespiratory
fitness. Weight gain caused by experimental overfeeding is associated
with a decrease in the flow-mediated dilatation of the brachial artery,
which is in turn correlated with the gains in visceral fat.
The effects of overfeeding on the plasma lipid and lipoprotein
profile are well documented. Cholesterol and triglyceride levels usu-
ally increase with weight gain but the values remain generally within
the normal ranges. Similarly, the lipoprotein profile is not substantially
altered with exposure to overfeeding. However, plasma triglyceride
concentrations are sensitive to the amounts and types of carbohy-
drate and fat in the diet as they influence both triglyceride formation
and removal.
The profile of adipose cell size and the metabolism of adipose tis-
sue is similar in spontaneous obesity and experimental overfeeding.
BRAY AND BOUCHARD
No consistent measurable change in fat cell size is seen with a weight
gain of less than 3 kg. With larger weight gain, an increase in abdomi-
nal fat cell size is observed but less so in the femoral depot where the
increased demand for lipid storage is accommodated by an increase in
fat cell number. Beige adipose cells, UCP1-dependent and indepen-
dent pathways could potentially play a role in adaptation to overfeed-
ing but they have not been tested systematically yet. The adipose cell
matrix is remodelled in response to overfeeding but this appears to be
a late event observed in long-term studies. Overfeeding does not
seem to alter basal and stimulated adipocyte lipolytic levels. This con-
clusion is also supported by the observation that overfeeding does
not change the adipose tissue expression of genes involved in lipolytic
pathways. Likewise no changes in adipose tissue LPL activity or LPL
gene expression have been found after overfeeding. Experimental
overfeeding entrains an inflammatory response in adipose tissue but
the response is variable and is partly influenced by the macronutrient
composition of the caloric surplus. With the increase in fat cell size,
overfeeding also leads to connective tissue deposition in adipose tis-
sue with no consistent increase in the number of macrophages. Short
duration and long-term overfeeding protocols have shown that blood
levels of leptin increase with overfeeding. Leptin gene expression is
increased in abdominal and femoral fat depots. Adiponectin is
secreted by the adipose tissue and low levels are associated with insu-
lin resistance. For reasons that are not clear yet, many studies have
been conducted on the adiponectin response to overfeeding but the
findings are heterogeneous and divergent. Global gene expression
profiling in adipose tissue samples shows that exposure to overfeed-
ing has a profound effect on the transcriptome.
With short-term overfeeding, no detectable changes in fat-free
mass or skeletal muscle mass are reported. However with overfeeding
protocols lasting from about 1 to 3 months, 40% to 50% of the weight
gained is in the form of fat-free mass and is mainly accounted for by
an increase in muscle mass with a smaller increase in nonmuscle pro-
tein residual mass.162 When the caloric surplus has a normal protein
content, the gain in muscle mass represents as much as 6% to 8% of
the baseline muscle mass. Overfeeding does not cause changes in
muscle fibre type distribution, muscle capillary density or glycogen
content. Studies have shown that chronic overfeeding can cause an
upregulation of muscle extracellular matrix, fibrosis and inflammatory
genes, all of which are associated with metabolic dysfunction. It is not
clear whether overfeeding increases intramyocellular lipid deposition.
Markers of muscle energy sensing networks are unchanged with short
duration overfeeding as are mitochondrial markers of oxidative phos-
phorylation. Gene expression studies reveal that large number of
genes are up-regulated or down-regulated in skeletal muscle in
response to overfeeding. The up-regulated genes reflect increased
lipid deposition in muscle and augmented lipogenesis whereas the
down-regulated genes suggest attenuation of muscle lipid metabo-
lism. Mitochondrial DNA variability with overfeeding suggest
increases in mitochondrial biogenesis. Basal glucose uptake by skele-
tal muscle is unchanged with overfeeding but the response to insu-
lin infusion is blunted with the weight gain. The development of
insulin resistance in muscle with weight gain in normal weight
BRAY AND BOUCHARD 67

subjects is similar to what is observed in spontaneous obesity. Even
though there is still controversy, the preponderance of the evidence
suggests that overfeeding and associated weight gain do not cause
detectable changes in whole-body work efficiency under standard-
ized ergometry conditions and do not cause alterations in ATP
fluxes or energy cost of muscle contraction. In contrast, caloric
restriction and major weight loss have been shown to increase
skeletal muscle efficiency. Finally, a low baseline low skeletal muscle
oxidative potential is associated with a larger weight gain in
response to experimental overfeeding.
Even when subjects make no specific effort to lose weight,
rapid weight loss after overfeeding has been documented in
several studies. However, inter-individual differences are commonly
observed in body weight recovery when overfeeding stops. In the
recovery phase from overfeeding, participants typically resume a
normal diet with no evidence of caloric restriction. This suggests
that there are persisting elevations in energy expenditure and pos-
sibly activity levels that explain the weight recovery in addition to
the normalization of caloric intake.
It has been suggested that adaptation to chronic overfeeding is
influenced by genetic differences. Unbiased genome- and
transcriptome-wide technologies together with proper overfeeding
study designs will be required to uncover the complete genotype
responsible for the variability in responsiveness to experimental over-
feeding. A number of attempts aimed at understanding whether DNA
methylation plays a role in human variability when confronted by
hyperphagia have provided indications that this may well be the case.
Research on the global DNA methylome but also the chemical profil-
ing of histones in relevant tissues will be necessary to shed light on
this fundamental issue.
The major elements of contemporary human overfeeding
research paradigm are depicted in Figure 11. One limitation is that
even when volunteers are confined to an in-patient setting for the
duration of the overfeeding experiment, there is no practical way
to clamp variability in metabolic rates (e.g., thermic effect of meals)
and in energy expenditure of spontaneous physical activities and
fidgeting. A major strength of experimental overfeeding is that the
caloric overload and its nutrient composition can be clamped rea-
sonably well, particularly when the experimental subjects are under
experimental control 20 h a day in a clinical setting. If one is inter-
ested in a mean treatment effect, a combination of in-patient con-
ditions on key days at the onset of the intervention and the last

F I G U R E 1 1 Schematic depiction of the major components of the human experimental overfeeding paradigm. Even though excellent control
can be imposed on the number of calories overfed and its macronutrient composition, the control of micronutrient intake, metabolic rates and
energy expenditure of physical activity is typically less rigorous. The arrows of variable size connecting the box on innate and acquired biological
individuality to the box on the level of positive energy balance and nutrient partitioning illustrate the range of response potentially accounted for
by genetic and epigenetic factors. The conditions under which post overfeeding measurements are performed make it possible to address
different goals related to the overfeeding per se and/or the body weight gain. See text for more explanation
68
few days preceding the post overfeeding measurement period, in
combination with prepackaged meals for the food to be consumed
away from the clinic, could be sufficient. However, when one is
interested in human variability in response to experimental over-
feeding, it would be critical to keep all volunteers as inpatients at
all times.
1 8 | CONC LU SIONS
Research on the biology of experimental overfeeding began with
single-subject studies in which investigators altered their own diets to
evaluate the consequences on body weight over long periods of time.
Beginning in the 1950s, we began to see studies in which experimen-
tal overfeeding was explored with multiple volunteers and more
robust experimental designs. Since then, studies have grown in
sophistication and have been asking more refined questions utilizing
the most advanced techniques. We believe that experimental over-
feeding offers a unique paradigm to gain insight not only into the biol-
ogy of human adaptability but also into the aetiology and
consequences of obesity and other metabolic disorders.
Important distinctions have emerged that have implications for
future studies in this field. Short-term overfeeding protocols are par-
ticularly useful for the investigation of the acute or repeated exposure
to a caloric surplus or a given nutrient combination in the presence of
no meaningful changes in body weight or adiposity. Long-term over-
feeding studies, say 1 month and more, offer not only the ability to
evaluate the effects of experimental hyperphagia and of specific nutri-
ent overload or imbalance but also the changes in body weight and
body composition. Moreover, when the post overfeeding protocol
measurements are performed on subjects who have become weight
stable at their new body weight level, the design makes it possible to
assess the consequences of the gains in body weight and adiposity.
Experimental overfeeding is a powerful research design that can be
helpful in elucidating the determinants of energy balance, the meta-
bolic drivers of energy partitioning, the molecular basis of metabolic
dysfunction and the mechanisms of body weight regulation as well as
the basic biology underlying a predisposition to gain weight or to
resist weight gain.
Considering the multiple advantages of rigorously implemented
experimental overfeeding protocols performed with the collaboration
of volunteers who are under in-patient conditions for the whole dura-
tion of the study, we believe that the time has come to launch multi-
centre collaborative studies aimed at uncovering the physiological,
metabolic and molecular mechanisms underlying adaptation to hyper-
phagia. Collaborative efforts in experimental overfeeding protocols
would make it possible to attain sufficiently large sample sizes to jus-
tify incorporating genomic, epigenomic, transcriptomic, proteomic,
lipidomic and metabolomics panels aimed at illuminating the biology
of adaptation to hyperphagia and the predisposition to gain weight
and adiposity or to defend body weight.
A substantial body of knowledge has accumulated over the last
70 years of research as this review makes clear. However, there are
BRAY AND BOUCHARD
still many questions that have been only partially answered and gap-
ing holes remain in our understanding of the biology of adaptation to
experimental overfeeding and on the prediction of the responsiveness
to a given dose of overfeeding. Hopefully the next generation of
experimental overfeeding studies will make it possible to close
these gaps.
AC KNOWLEDG EME NT S
The authors would like to express their gratitude to Melanie Peterson
for her assistance with editing of the text, tables, figures and refer-
ences. Thanks are also expressed to Lori Steib for her input in the sea-
rch for the human overfeeding related literature. We also thank
Christian Couture for his help with new analysis performed on the
data of the overfeeding study conducted on pairs of identical twins.
GAB is Boyd Professor Emeritus of the Pennington Biomedical
Research Center, a campus of the Louisiana State University System.
CB is Boyd Professor and the John W, Barton Sr. Chair in Genetics
and Nutrition at the same institution. CB is partially supported by the
National Institute of General Medical Sciences (NIGMS)-funded
COBRE Grant 8-P30-GM-118430-01.
CONFLICTS OF INTEREST
The authors have no conflicts of interest to declare.
OR CID
George A. Bray https://orcid.org/0000-0001-9945-8772
Claude Bouchard https://orcid.org/0000-0002-0048-491X
RE FE RE NCE S
1. Gulick A. A study of weight regulation in the adult human body dur-
ing over-nutrition. Am J Physiol. 1922;60:371-395.
2. Neumann RO. Experimentelle Beiträge zur Lehre von dem täglichen
Nahrungsbedarf des Menschen unter besonderer Berücksichtigung
der notwendigen Eiweissmenge. Arch Hyg Bakteriol. 1902;45:1-87.
3. Bouchard C, Tremblay A. Genetic influences on the response
of body fat and fat distribution to positive and negative
energy balances in human identical twins. J Nutr. 1997;127:943S-
947S.
4. Bray GA. Luxuskonsumption—myth or reality? Obes Res. 1995;3:
491-495.
5. Bray GA. The Battle of the Bulge. Pittsburg, PA: Dorrance Publishing
Co.; 2007.
6. Danforth E Jr, Horton ES, O'Connell M, et al. Dietary-induced alter-
ations in thyroid hormone metabolism during overnutrition. J Clin
Invest. 1979;64(5):1336-1347.
7. Danforth EJ, Sims EAH. Thermic effects of overfeeding: role of
thyroid hormones, catecholamines, and insulin resistance. In:
Angel A, Hollenberg CH, Roncari DAK, eds. The Adipocyte and Obe-
sity: Cellular and Molecular Mechanisms. New York: Raven Press;
1983:271-282.
8. Dhurandhar EJ, Kaiser KA, Dawson JA, Alcorn AS, Keating KD,
Allison DB. Predicting adult weight change in the real world: a sys-
tematic review and meta-analysis accounting for compensatory
changes in energy intake or expenditure. Int J Obes (Lond). 2015;
39(8):1181-1187.
9. Dulloo AG, Jacquet J. Low-protein overfeeding: a tool to unmask
susceptibility to obesity in humans. Int J Obes Relat Metab Disord.
1999;23:1118-1121.
BRAY AND BOUCHARD
10. Garrow JS. Chronic effects of over- and under-nutrition on thermo-
genesis. Int J Vitam Nutr Res. 1986;56:201-204.
11. Horton ES, Danforth E, Jr., Sims EAH, Salans LB. Endocrine and
metabolic alterations in spontaneous and experimental obesity. In:
Bray GA, John E. Fogarty International Center for Advanced Study
in the Health Sciences (eds.). Obesity in Perspective: Proceedings of
the Conference. U.S. Govt. Printing Office: Washington DC 1975;
323-334.
12. James WP, McNeill G, Ralph A. Metabolism and nutritional adapta-
tion to altered intakes of energy substrates. Am J Clin Nutr. 1990;51:
264-269.
13. Muller MJ, Enderle J, Bosy-Westphal A. Changes in energy expendi-
ture with weight gain and weight loss in humans. Curr Obes Rep.
2016;5:413-423.
14. Norgan NG. Thermogenesis above maintenance in humans. Proc
Nutr Soc. 1990;49:217-226.
15. Olena A. How gaining and losing weight affects the body. In: The Sci-
entist. LabX Media Group; 2018.
16. Rosenbaum M. Metabolic propensity for weight gain unmasked by
overfeeding. Obesity (Silver Spring). 2018;26:633-634.
17. Saltzman E, Roberts SB. Effects of energy imbalance on energy
expenditure and respiratory quotient in young and older men: a
summary of data from two metabolic studies. Aging (Milano). 1996;8:
370-378.
18. Schutz Y. Human overfeeding experiments: potentials and limita-
tions in obesity research. Br J Nutr. 2000;84:135-137.
19. Stock MJ. Gluttony and thermogenesis revisited. Int J Obes Relat
Metab Disord. 1999;23:1105-1117.
20. St-Pierre DH, George V, Rabasa-Lhoret R, Poehlman ET. Genetic
variation and statistical considerations in relation to overfeeding and
underfeeding in humans. Nutrition. 2004;20:145-154.
21. Tappy L. Metabolic consequences of overfeeding in humans. Curr
Opin Clin Nutr Metab Care. 2004;7:623-628.
22. Westerterp KR. Metabolic adaptations to over—and underfeeding—
still a matter of debate? Eur J Clin Nutr. 2013;67:443-445.
23. Cuthbertson DJ, Steele T, Wilding JP, et al. What have human exper-
imental overfeeding studies taught us about adipose tissue expan-
sion and susceptibility to obesity and metabolic complications? Int J
Obes (Lond). 2017;41(6):853-865.
24. Engin A. Eat and death: chronic over-eating. Adv Exp Med Biol. 2017;
960:53-80.
25. Joosen AM, Westerterp KR. Energy expenditure during overfeeding.
Nutr Metab (Lond). 2006;3:25.
26. Schutz Y. Dietary fat, lipogenesis and energy balance. Physiol Behav.
2004;83:557-564.
27. Leaf A, Antonio J. The effects of overfeeding on body composition:
the role of macronutrient composition—a narrative review. Int J
Exerc Sci. 2017;10:1275-1296.
28. Ng M, Fleming T, Robinson M, et al. Global, regional, and national
prevalence of overweight and obesity in children and adults during
1980-2013: a systematic analysis for the Global Burden of Disease
Study 2013. Lancet. 2014;384(9945):766-781.
29. Forbes GB. Energy intake and body weight: a reexamination of two
“classic” studies. Am J Clin Nutr. 1984;39:349-350.
30. Wiley FH, Newburgh LH. The doubtful nature of
“luxuskonsumption”. J Clin Invest. 1931;10:733-744.
31. Horton TJ, Drougas H, Brachey A, Reed GW, Peters JC, Hill JO. Fat
and carbohydrate overfeeding in humans: different effects on
energy storage. Am J Clin Nutr. 1995;62(1):19-29.
32. Kovacs EM, Westerterp-Plantenga MS. Effects of (-)-hydroxycitrate
on net fat synthesis as de novo lipogenesis. Physiol Behav. 2006;88:
371-381.
33. Bray GA, Smith SR, de Jonge L, et al. Effect of dietary protein content
on weight gain, energy expenditure, and body composition during
overeating: a randomized controlled trial. JAMA. 2012;307(1):47-55.
69
34. Bouchard C, Tchernof A, Tremblay A. Predictors of body composi-
tion and body energy changes in response to chronic overfeeding.
Int J Obes (Lond). 2014;38:236-242.
35. Shamseer L, Moher D, Clarke M, et al. Preferred reporting items for
systematic review and meta-analysis protocols (PRISMA-P) 2015:
elaboration and explanation. BMJ. 2015;350:g7647.
36. Stock M, Rothwell N. Obesity and Leanness: Basic Aspects. New York:
John Wiley & Sons; 1982.
37. Schoeller DA, van Santen E. Measurement of energy expenditure in
humans by doubly labeled water method. J Appl Physiol Respir Envi-
ron Exerc Physiol. 1982;53:955-959.
38. FAO. Food and Nutrition Paper 77: Food energy - methods of analy-
sis and conversion factors. In: Food and Agriculture Organization of
the United Nations (FAO). Rome: Food and Agriculture Organization
of the United Nations; 2003.
39. Flatt JP. The biochemistry of energy expenditure. In: Bray GA,
ed. Recent advances in obesity research II. London: Newman; 1978:
1-28.
40. Goranzon H, Forsum E, Thilen M. Calculation and determination of
metabolizable energy in mixed diets to humans. Am J Clin Nutr.
1983;38:954-963.
41. Hwaung P, Bosy-Westphal A, Muller MJ, et al. Obesity tissue: com-
position, energy expenditure, and energy content in adult humans.
Obesity (Silver Spring). 2019;27(9):1472-1481.
42. Schutz Y, Dulloo AG. Resting metabolic rate, thermic effect of food,
and obesity. In: Bray GA, Bouchard C, eds. Handbook of Obesity—
Volume 1: Epidemiology, Etiology, and Pathophysiology. Boca Raton:
CRC Press; 2014:267-279.
43. Alligier M, Meugnier E, Debard C, et al. Subcutaneous adipose tissue
remodeling during the initial phase of weight gain induced by over-
feeding in humans. J Clin Endocrinol Metab. 2012;97:E183-E192.
44. Bouchard C, Tremblay A, Despres JP, et al. The response to long-
term overfeeding in identical twins. N Engl J Med. 1990;322:1477-
1482.
45. Brands M, Swat M, Lammers NM, et al. Effects of a hypercaloric diet
on beta-cell responsivity in lean healthy men. Clin Endocrinol (Oxf).
2013;78:217-225.
46. Bray GA, Whipp BJ, Koyal SN. The acute effects of food intake on
energy expenditure during cycle ergometry. Am J Clin Nutr. 1974;27:
254-259.
47. Diaz EO, Prentice AM, Goldberg GR, Murgatroyd PR, Coward WA.
Metabolic response to experimental overfeeding in lean
and overweight healthy volunteers. Am J Clin Nutr. 1992;56:
641-655.
48. Erdmann J, Kallabis B, Oppel U, et al. Development of
hyperinsulinemia and insulin resistance during the early stage of
weight gain. Am J Physiol Endocrinol Metab. 2008;294:E568-E575.
49. Fabbrini E, Yoshino J, Yoshino M, et al. Metabolically normal obese
people are protected from adverse effects following weight gain.
J Clin Invest. 2015;125:787-795.
50. Franck N, Gummesson A, Jernas M, et al. Identification of adipocyte
genes regulated by caloric intake. J Clin Endocrinol Metab. 2011;96:
E413-E418.
51. Gentile CL, Orr JS, Davy BM, Davy KP. Cardiorespiratory fitness
influences the blood pressure response to experimental weight gain.
Obesity (Silver Spring). 2007;15:3005-3012.
52. Johannsen DL, Tchoukalova Y, Tam CS, et al. Effect of 8 weeks of
overfeeding on ectopic fat deposition and insulin sensitivity: testing
the “adipose tissue expandability” hypothesis. Diabetes Care. 2014;
37:2789-2797.
53. Kolaczynski JW, Ohannesian JP, Considine RV, Marco CC, Caro JF.
Response of leptin to short-term and prolonged overfeeding in
humans. J Clin Endocrinol Metab. 1996;81:4162-4165.
54. Koopman KE, Booij J, Fliers E, Serlie MJ, la Fleur SE. Diet-induced
changes in the Lean Brain: Hypercaloric high-fat-high-sugar snacking
70
decreases serotonin transporters in the human hypothalamic region.
Mol Metab. 2013;2:417-422.
55. Leibel RL, Rosenbaum M, Hirsch J. Changes in energy expenditure
resulting from altered body weight. N Engl J Med. 1995;332:
621-628.
56. Levine JA, Eberhardt NL, Jensen MD. Role of nonexercise activity
thermogenesis in resistance to fat gain in humans. Science. 1999;
283:212-214.
57. Miller DS, Mumford P. Gluttony. 1. An experimental study of overeat-
ing low- or high-protein diets. Am J Clin Nutr. 1967;20:1212-1222.
58. Norgan NG, Durnin JV. The effect of 6 weeks of overfeeding on the
body weight, body composition, and energy metabolism of young
men. Am J Clin Nutr. 1980;33:978-988.
59. Pasquet P, Brigant L, Froment A, et al. Massive overfeeding and
energy balance in men: the Guru Walla model. Am J Clin Nutr. 1992;
56:483-490.
60. Riumallo JA, Schoeller D, Barrera G, Gattas V, Uauy R. Energy
expenditure in underweight free-living adults: impact of energy sup-
plementation as determined by doubly labeled water and indirect
calorimetry. Am J Clin Nutr. 1989;49:239-246.
61. Romero-Corral A, Sert-Kuniyoshi FH, Sierra-Johnson J, et al. Modest
visceral fat gain causes endothelial dysfunction in healthy humans.
J Am Coll Cardiol. 2010;56:662-666.
62. Rosqvist F, Iggman D, Kullberg J, et al. Overfeeding polyunsaturated
and saturated fat causes distinct effects on liver and visceral fat
accumulation in humans. Diabetes. 2014;63:2356-2368.
63. Sims EA, Goldman RF, Gluck CM, et al. Experimental obesity in man.
Trans Assoc Am Physicians. 1968;81:153-170.
64. Spillane M, Willoughby DS. Daily overfeeding from protein and/or
carbohydrate supplementation for eight weeks in conjunction with
resistance training does not improve body composition and muscle
strength or increase markers indicative of muscle protein synthesis
and myogenesis in resistance-trained males. J Sports Sci Med. 2016;
15:17-25.
65. Tchoukalova YD, Votruba SB, Tchkonia T, et al. Regional differences
in cellular mechanisms of adipose tissue gain with overfeeding. Proc
Natl Acad Sci U S A. 2010;107:18226-18231.
66. Webb P, Annis JF. Adaptation to overeating in lean and overweight
men and women. Hum Nutr Clin Nutr. 1983;37:117-131.
67. Acheson KJ, Schutz Y, Bessard T, et al. Glycogen storage capacity
and de novo lipogenesis during massive carbohydrate overfeeding in
man. Am J Clin Nutr. 1988;48:240-247.
68. Apfelbaum M, Bostsarron J, Lacatis D. Effect of caloric restriction
and excessive caloric intake on energy expenditure. Am J Clin Nutr.
1971;24:1405-1409.
69. Baba NH, Sultan R, Cortas N, Habbal Z. Diet composition affects
weight gain, adiposity and blood parameters in healthy human vol-
unteers. Nutr Res. 1999;19:1313-1326.
70. Bandini LG, Schoeller DA, Edwards J, et al. Energy expenditure dur-
ing carbohydrate overfeeding in obese and nonobese adolescents.
Am J Physiol. 1989;256:E357-E367.
71. Bray GA. Lipogenesis in human adipose tissue: some effects of nib-
bling and gorging. J Clin Invest. 1972;51:537-548.
72. Claesson AL, Holm G, Ernersson A, Lindstrom T, Nystrom FH.
Two weeks of overfeeding with candy, but not peanuts, increases
insulin levels and body weight. Scand J Clin Lab Invest. 2009;69:
598-605.
73. Cornford AS, Barkan AL, Hinko A, Horowitz JF. Suppression in
growth hormone during overeating ameliorates the increase in insu-
lin resistance and cardiovascular disease risk. Am J Physiol Endocrinol
Metab. 2012;303:E1264-E1272.
74. Forbes GB, Brown MR, Welle SL, Lipinski BA. Deliberate overfeed-
ing in women and men: energy cost and composition of the weight
gain. Br J Nutr. 1986;56:1-9.
BRAY AND BOUCHARD
75. French SJ, Murray B, Rumsey RD, Fadzlin R, Read NW. Adaptation
to high-fat diets: effects on eating behaviour and plasma cholecysto-
kinin. Br J Nutr. 1995;73:179-189.
76. Germain N, Galusca B, Caron-Dorval D, et al. Specific appetite, ener-
getic and metabolomics responses to fat overfeeding in resistant-to-
bodyweight-gain constitutional thinness. Nutr Diabetes. 2014;4:e126.
77. Goldrick RB, Havenstein N, Carroll KF, Reardon M. Effects of over-
feeding on lipid and carbohydrate metabolism in lean young adults.
Metabolism. 1972;21:761-770.
78. Goran MI, Calles-Escandon J, Poehlman ET, O'Connell M,
Danforth E Jr. Effects of increased energy intake and/or physical
activity on energy expenditure in young healthy men. J Appl Physiol
(1985). 1994;77:366-372.
79. Jebb SA, Prentice AM, Goldberg GR, et al. Changes in macronutrient
balance during over- and underfeeding assessed by 12-d continuous
whole-body calorimetry. Am J Clin Nutr. 1996;64:259-266.
80. Jebb SA, Siervo M, Fruhbeck G, et al. Variability of appetite control
mechanisms in response to 9 weeks of progressive overfeeding in
humans. Int J Obes (Lond). 2006;30:1160-1162.
81. Joosen AM, Bakker AH, Westerterp KR. Metabolic efficiency and
energy expenditure during short-term overfeeding. Physiol Behav.
2005;85:593-597.
82. Katzeff HL, O'Connell M, Horton ES, et al. Metabolic studies in
human obesity during overnutrition and undernutrition: thermogenic
and hormonal responses to norepinephrine. Metabolism. 1986;35:
166-175.
83. Klein S, Goran M. Energy metabolism in response to overfeeding in
young adult men. Metabolism. 1993;42:1201-1205.
84. Lammert O, Hansen ES. Effects of excessive caloric intake and calo-
ric restriction on body weight and energy expenditure at rest and
light exercise. Acta Physiol Scand. 1982;114:135-141.
85. Lammert O, Grunnet N, Faber P, et al. Effects of isoenergetic over-
feeding of either carbohydrate or fat in young men. Br J Nutr. 2000;
84:233-245.
86. Mann GV, Teel K, Hayes O, Mc NA, Bruno D. Exercise in the disposi-
tion of dietary calories; regulation of serum lipoprotein and cholesterol
levels in human subjects. N Engl J Med. 1955;253:349-355.
87. McLaughlin T, Craig C, Liu LF, et al. Adipose cell size and regional fat
deposition as predictors of metabolic response to overfeeding in
insulin-resistant and insulin-sensitive humans. Diabetes. 2016;65:
1245-1254.
88. Meugnier E, Bossu C, Oliel M, et al. Changes in gene expression in
skeletal muscle in response to fat overfeeding in lean men. Obesity
(Silver Spring). 2007;15:2583-2594.
89. O'Dea K, Esler M, Leonard P, Stockigt JR, Nestel P. Noradrenaline
turnover during under- and over-eating in normal weight subjects.
Metabolism. 1982;31:896-899.
90. Olefsky J, Crapo PA, Ginsberg H, Reaven GM. Metabolic effects of
increased caloric intake in man. Metabolism. 1975;24:495-503.
91. Passmore R, Meiklejohn AP, Dewar AD, Thow RK. Energy utilization
in overfed thin young men. Br J Nutr. 1955;9:20-27.
92. Passmore R, Meiklejohn AP, Dewar AD, Thow RK. An analysis of the
gain in weight of overfed thin young men. Br J Nutr. 1955;9:27-37.
93. Passmore R, Strong JA, Swindells YE, Eldin N. The effect of over-
feeding on two fat young women. Br J Nutr. 1963;17:373-383.
94. Poehlman ET, Tremblay A, Fontaine E, et al. Genotype dependency
of the thermic effect of a meal and associated hormonal changes fol-
lowing short-term overfeeding. Metabolism. 1986;35:30-36.
95. Ravussin E, Schutz Y, Acheson KJ, et al. Short-term, mixed-diet over-
feeding in man: no evidence for “luxuskonsumption”. Am J Physiol.
1985;249:E470-E477.
96. Roberts SB, Young VR, Fuss P, et al. Energy-expenditure and subse-
quent nutrient intakes in overfed young men. Am J Physiol. 1990;
259:R461-R469.
BRAY AND BOUCHARD
97. Robertson MD, Henderson RA, Vist GE, Rumsey RD. Plasma ghrelin
response following a period of acute overfeeding in normal weight
men. Int J Obes Relat Metab Disord. 2004;28:727-733.
98. Samocha-Bonet D, Campbell LV, Viardot A, et al. A family history of
type 2 diabetes increases risk factors associated with overfeeding.
Diabetologia. 2010;53:1700-1708.
99. Welle S, Campbell RG. Stimulation of thermogenesis by carbohy-
drate overfeeding. Evidence against sympathetic nervous system
mediation. J Clin Invest. 1983;71:916-925.
100. Welle SL, Nair KS, Campbell RG. Failure of chronic beta-adrenergic
blockade to inhibit overfeeding-induced thermogenesis in humans.
Am J Physiol. 1989;256:R653-R658.
101. Aarsland A, Chinkes D, Wolfe RR. Hepatic and whole-body fat syn-
thesis in humans during carbohydrate overfeeding. Am J Clin Nutr.
1997;65:1774-1782.
102. Adochio RL, Leitner JW, Gray K, Draznin B, Cornier MA. Early
responses of insulin signaling to high-carbohydrate and high-fat
overfeeding. Nutr Metab (Lond). 2009;6:37.
103. Apolzan JW, Bray GA, Hamilton MT, et al. Short-term overeating
results in incomplete energy intake compensation regardless of
energy density or macronutrient composition. Obesity (Silver Spring).
2014;22:119-130.
104. Brons C, Jacobsen S, Nilsson E, et al. Deoxyribonucleic acid methyla-
tion and gene expression of PPARGC1A in human muscle is
influenced by high-fat overfeeding in a birth-weight-dependent
manner. J Clin Endocrinol Metab. 2010;95:3048-3056.
105. Casper K, Matthews DE, Heymsfield SB. Overfeeding: cardiovascu-
lar and metabolic response during continuous formula infusion in
adult humans. Am J Clin Nutr. 1990;52:602-609.
106. Chen M, Wu L, Zhao J, et al. Altered glucose metabolism in mouse
and humans conceived by IVF. Diabetes. 2014;63:3189-3198.
107. Clore JN, Helm ST, Blackard WG. Loss of hepatic autoregulation
after carbohydrate overfeeding in normal man. J Clin Invest. 1995;
96:1967-1972.
108. Cooper JA, Polonsky KS, Schoeller DA. Serum leptin levels in obese
males during over- and underfeeding. Obesity (Silver Spring). 2009;
17:2149-2154.
109. Cornier MA, Grunwald GK, Johnson SL, Bessesen DH. Effects of
short-term overfeeding on hunger, satiety, and energy intake in thin
and reduced-obese individuals. Appetite. 2004;43:253-259.
110. Dallosso HM, James WP. Whole-body calorimetry studies in adult
men. 2. The interaction of exercise and over-feeding on the thermic
effect of a meal. Br J Nutr. 1984;52:65-72.
111. Dirlewanger M, di Vetta V, Guenat E, et al. Effects of short-term car-
bohydrate or fat overfeeding on energy expenditure and plasma lep-
tin concentrations in healthy female subjects. Int J Obes Relat Metab
Disord. 2000;24:1413-1418.
112. Faeh D, Minehira K, Schwarz JM, et al. Effect of fructose overfeed-
ing and fish oil administration on hepatic de novo lipogenesis and
insulin sensitivity in healthy men. Diabetes. 2005;54:1907-1913.
113. Fagerberg B, Herlitz H, Jonsson O, et al. Increased erythrocyte
sodium efflux during overfeeding without evidence of mediation by
circulating catecholamines or thyroid hormone. Metabolism. 1984;
33:994-998.
114. Freymond D, Larson K, Bogardus C, Ravussin E. Energy expenditure
during normo- and overfeeding in peripubertal children of lean and
obese Pima Indians. Am J Physiol. 1989;257:E647-E653.
115. Glick Z, Shvartz E, Magazanik A, Modan M. Absence of increased
thermogenesis during short-term overfeeding in normal and over-
weight women. Am J Clin Nutr. 1977;30:1026-1035.
116. Hagobian TA, Braun B. Interactions between energy surplus and
short-term exercise on glucose and insulin responses in healthy peo-
ple with induced, mild insulin insensitivity. Metabolism. 2006;55:
402-408.
71
117. He J, Votruba S, Pomeroy J, Bonfiglio S, Krakoff J. Measurement of
ad libitum food intake, physical activity, and sedentary time in
response to overfeeding. PLoS ONE. 2012;7:e36225.
118. Hill JO, Peters JC, Yang D, et al. Thermogenesis in humans during
overfeeding with medium-chain triglycerides. Metabolism. 1989;38:
641-648.
119. Hulston CJ, Churnside AA, Venables MC. Probiotic supplementation
prevents high-fat, overfeeding-induced insulin resistance in human
subjects. Br J Nutr. 2015;113:596-602.
120. Johnstone AM, Stubbs RJ, Harbron CG. Effect of overfeeding mac-
ronutrients on day-to-day food intake in man. Eur J Clin Nutr. 1996;
50:418-430.
121. Kosmiski LA, Bessesen DH, Stotz SA, Koeppe JR, Horton TJ. Short-
term overfeeding increases resting energy expenditure in patients
with HIV lipodystrophy. Am J Clin Nutr. 2007;86:1009-1015.
122. Lagerpusch M, Enderle J, Later W, et al. Impact of glycaemic index
and dietary fibre on insulin sensitivity during the refeeding phase of
a weight cycle in young healthy men. Br J Nutr. 2013;109:1606-
1616.
123. Lecoultre V, Egli L, Carrel G, et al. Effects of fructose and glucose
overfeeding on hepatic insulin sensitivity and intrahepatic lipids in
healthy humans. Obesity (Silver Spring). 2013;21:782-785.
124. Lundsgaard AM, Fritzen AM, Sjoberg KA, et al. Circulating FGF21 in
humans is potently induced by short term overfeeding of carbohy-
drates. Mol Metab. 2017;6:22-29.
125. Magkos F, Smith GI, Reeds DN, et al. One day of overfeeding
impairs nocturnal glucose but not fatty acid homeostasis in over-
weight men. Obesity (Silver Spring). 2014;22:435-440.
126. McDevitt RM, Poppitt SD, Murgatroyd PR, Prentice AM. Macronu-
trient disposal during controlled overfeeding with glucose, fructose,
sucrose, or fat in lean and obese women. Am J Clin Nutr. 2000;72:
369-377.
127. Minehira K, Bettschart V, Vidal H, et al. Effect of carbohydrate over-
feeding on whole body and adipose tissue metabolism in humans.
Obes Res. 2003;11:1096-1103.
128. Murgatroyd PR, Goldberg GR, Leahy FE, Gilsenan MB, Prentice AM.
Effects of inactivity and diet composition on human energy balance.
Int J Obes Relat Metab Disord. 1999;23:1269-1275.
129. Nijhuis J, van Dielen FM, Schaper NC, et al. Short-term overfeeding
induces insulin resistance in weight-stable patients after bariatric
surgery. Obes Surg. 2008;18:300-305.
130. Parry SA, Smith JR, Corbett TR, Woods RM, Hulston CJ. Short-term,
high-fat overfeeding impairs glycaemic control but does not alter
gut hormone responses to a mixed meal tolerance test in healthy,
normal-weight individuals. Br J Nutr. 2017;117:48-55.
131. Peterson CM, Lecoultre V, Frost EA, et al. The thermogenic
responses to overfeeding and cold are differentially regulated. Obe-
sity (Silver Spring). 2016;24:96-101.
132. Sagayama H, Jikumaru Y, Hirata A, et al. Measurement of body com-
position in response to a short period of overfeeding. J Physiol
Anthropol. 2014;33:29.
133. Schmidt SL, Harmon KA, Sharp TA, Kealey EH, Bessesen DH. The
effects of overfeeding on spontaneous physical activity in obesity
prone and obesity resistant humans. Obesity (Silver Spring). 2012;20:
2186-2193.
134. Schwarz JM, Neese RA, Turner S, Dare D, Hellerstein MK. Short-
term alterations in carbohydrate energy intake in humans. Striking
effects on hepatic glucose production, de novo lipogenesis, lipolysis,
and whole-body fuel selection. J Clin Invest. 1995;96:2735-2743.
135. Sobrecases H, Le KA, Bortolotti M, et al. Effects of short-term over-
feeding with fructose, fat and fructose plus fat on plasma and
hepatic lipids in healthy men. Diabetes Metab. 2010;36:244-246.
136. Sun G, Bishop J, Khalili S, et al. Serum visfatin concentrations are
positively correlated with serum triacylglycerols and down-regulated
72
by overfeeding in healthy young men. Am J Clin Nutr. 2007;85:
399-404.
137. Utiger RD. Differing thyrotropin responses to increased serum triio-
dothyronine concentrations produced by overfeeding and by triio-
dothyronine administration. Metabolism. 1982;31:180-183.
138. van Aggel-Leijssen DP, van Baak MA, Tenenbaum R, Campfield LA,
Saris WH. Regulation of average 24h human plasma leptin level; the
influence of exercise and physiological changes in energy balance.
Int J Obes Relat Metab Disord. 1999;23:151-158.
139. Walhin JP, Richardson JD, Betts JA, Thompson D. Exercise counter-
acts the effects of short-term overfeeding and reduced physical
activity independent of energy imbalance in healthy young men.
J Physiol. 2013;591:6231-6243.
140. Weststrate JA, Hautvast JG. The effects of short-term carbohydrate
overfeeding and prior exercise on resting metabolic rate and diet-
induced thermogenesis. Metabolism. 1990;39:1232-1239.
141. Weyer C, Vozarova B, Ravussin E, Tataranni PA. Changes in energy
metabolism in response to 48 h of overfeeding and fasting in Cauca-
sians and Pima Indians. Int J Obes Relat Metab Disord. 2001;25:
593-600.
142. Wijers SL, Saris WH, van Marken Lichtenbelt WD. Individual ther-
mogenic responses to mild cold and overfeeding are closely related.
J Clin Endocrinol Metab. 2007;92:4299-4305.
143. Wulan SN, Westerterp KR, Plasqui G. Metabolic profile before and
after short-term overfeeding with a high-fat diet: a comparison
between South Asian and White men. Br J Nutr. 2014;111:1853-
1861.
144. Zed C, James WP. Dietary thermogenesis in obesity: fat feeding at
different energy intakes. Int J Obes (Lond). 1986;10:375-390.
145. Goldman RF, Haisman MF, Bynum G, Horton ES, Sims EAH. Experi-
mental obesity in man. metabolic rate in relation to dietary intake.
In: Bray GA, ed. Obesity in Perspective: a conference. Washington, D.
C: U.S. Government Printing Office; 1975:165-188.
146. Bouchard C, Tremblay A. Genetic effects in human energy expendi-
ture components. Int J Obes (Lond). 1990;14(Suppl 1):49-55. discus-
sion 55-8
147. Gentile CL, Orr JS, Davy BM, Davy KP. Modest weight gain is
associated with sympathetic neural activation in nonobese
humans. Am J Physiol Regul Integr Comp Physiol. 2007;292:R1834-
R1838.
148. Rosenbaum M, Ravussin E, Matthews DE, et al. A comparative study
of different means of assessing long-term energy expenditure in
humans. Am J Physiol. 1996;270:R496-R504.
149. Pasquet P, Apfelbaum M. Recovery of initial body weight and com-
position after long-term massive overfeeding in men. Am J Clin Nutr.
1994;60:861-863.
150. Roberts SB, Fuss P, Heyman MB, Dallal GE, Young VR. Effects of
age on energy expenditure and substrate oxidation during experi-
mental underfeeding in healthy men. J Gerontol A Biol Sci Med Sci.
1996;51:B158-B166.
151. Ling Y, Galusca B, Hager J, et al. Rational and design of an overfeed-
ing protocol in constitutional thinness: Understanding the physiol-
ogy, metabolism and genetic background of resistance to weight
gain. Ann Endocrinol (Paris). 2016;77:563-569.
152. Poehlman ET, Despres JP, Marcotte M, et al. Genotype dependency
of adaptation in adipose tissue metabolism after short-term over-
feeding. Am J Physiol. 1986;250:E480-E485.
153. Welle S, Matthews DE, Campbell RG, Nair KS. Stimulation of protein
turnover by carbohydrate overfeeding in men. Am J Physiol. 1989;
257:E413-E417.
154. Sock ETN, Le KA, Ith M, et al. Effects of a short-term overfeeding
with fructose or glucose in healthy young males. Brit J Nutr. 2010;
103:939-943.
155. Schmidt SL, Kealey EH, Horton TJ, VonKaenel S, Bessesen DH. The
effects of short-term overfeeding on energy expenditure and
BRAY AND BOUCHARD
nutrient oxidation in obesity-prone and obesity-resistant individuals.
Int J Obes (Lond). 2013;37:1192-1197.
156. Brons C, Jacobsen S, Hiscock N, et al. Effects of high-fat overfeed-
ing on mitochondrial function, glucose and fat metabolism, and
adipokine levels in low-birth-weight subjects. Am J Physiol Endocrinol
Metab. 2012;302:E43-E51.
157. Brons C, Jensen CB, Storgaard H, et al. Impact of short-term high-
fat feeding on glucose and insulin metabolism in young healthy men.
J Physiol. 2009;587:2387-2397.
158. Thearle MS, Pannacciulli N, Bonfiglio S, Pacak K, Krakoff J. Extent
and determinants of thermogenic responses to 24 hours of fasting,
energy balance, and five different overfeeding diets in humans.
J Clin Endocrinol Metab. 2013;98:2791-2799.
159. Vinales KL, Schlogl M, Piaggi P, et al. The consistency in
macronutrient oxidation and the role for epinephrine in the
response to fasting and overfeeding. J Clin Endocrinol Metab. 2017;
102:279-289.
160. Poehlman ET, Tremblay A, Despres JP, et al. Genotype-controlled
changes in body composition and fat morphology following over-
feeding in twins. Am J Clin Nutr. 1986;43:723-731.
161. Deriaz O, Tremblay A, Bouchard C. Non linear weight gain with long
term overfeeding in man. Obes Res. 1993;1:179-185.
162. Bray GA, Redman LM, de Jonge L, et al. Effect of protein overfeed-
ing on energy expenditure measured in a metabolic chamber.
Am J Clin Nutr. 2015;101:496-505.
163. Levine JA, Eberhardt NL, Jensen MD. Leptin responses to overfeed-
ing: relationship with body fat and nonexercise activity thermogene-
sis. J Clin Endocrinol Metab. 1999;84:2751-2754.
164. Johannsen DL, Marlatt KL, Conley KE, Smith SR, Ravussin E. Meta-
bolic adaptation is not observed after 8 weeks of overfeeding but
energy expenditure variability is associated with weight recovery.
Am J Clin Nutr. 2019.
165. Tremblay A, Despres JP, Theriault G, Fournier G, Bouchard C. Over-
feeding and energy expenditure in humans. Am J Clin Nutr. 1992;56:
857-862.
166. Sutton EF, Bray GA, Burton JH, Smith SR, Redman LM. No evidence
for metabolic adaptation in thermic effect of food by dietary protein.
Obesity (Silver Spring). 2016;24:1639-1642.
167. Apolzan JW, Bray GA, Smith SR, et al. Effects of weight gain induced
by controlled overfeeding on physical activity. Am J Physiol
Endocrinol Metab. 2014;307:E1030-E1037.
168. Whipp BJ, Bray GA, Koyal SN. Exercise energetics in normal man
following acute weight gain. Am J Clin Nutr. 1973;26:1284-1286.
169. Welle SL, Campbell RG. Improved carbohydrate tolerance and stim-
ulation of carbohydrate oxidation and lipogenesis during short-term
carbohydrate overfeeding. Metabolism. 1983;32:889-893.
170. Acheson KJ, Schutz Y, Bessard T, et al. Nutritional Influences on
Lipogenesis and Thermogenesis after a Carbohydrate Meal.
Am J Physiol. 1984;246:E62-E70.
171. Draznin B, Wang C, Adochio R, Leitner JW, Cornier MA. Effect of
dietary macronutrient composition on AMPK and SIRT1 expression
and activity in human skeletal muscle. Horm Metab Res. 2012;44:
650-655.
172. Vinales KL, Schlogl M, Reinhardt M, et al. Cycling efficiency during
incremental cycle ergometry after 24 hours of overfeeding or
fasting. Obesity (Silver Spring). 2018;26:368-377.
173. Schlogl M, Piaggi P, Pannacciuli N, et al. Energy expenditure
responses to fasting and overfeeding identify phenotypes associated
with weight change. Diabetes. 2015;64:3680-3689.
174. Bray GA, Glennon JA, Salans LB, et al. Spontaneous and experimen-
tal human obesity: effects of diet and adipose cell size on lipolysis
and lipogenesis. Metabolism. 1977;26:739-747.
175. Schlogl M, Piaggi P, Thiyyagura P, et al. Overfeeding over 24 hours
does not activate brown adipose tissue in humans. J Clin Endocrinol
Metab. 2013;98:E1956-E1960.
BRAY AND BOUCHARD
176. Alligier M, Gabert L, Meugnier E, et al. Visceral fat accumulation dur-
ing lipid overfeeding is related to subcutaneous adipose tissue charac-
teristics in healthy men. J Clin Endocrinol Metab. 2013;98:802-810.
177. Bray GA, Redman LM, de Jonge L, Rood J, Smith SR. Effect of three
levels of dietary protein on metabolic phenotype of healthy individ-
uals with 8 weeks of overfeeding. J Clin Endocrinol Metab. 2016;101:
2836-2843.
178. Cornford AS, Barkan AL, Horowitz JF. Rapid suppression of growth
hormone concentration by overeating: potential mediation by
hyperinsulinemia. J Clin Endocrinol Metab. 2011;96:824-830.
179. Cornford AS, Hinko A, Nelson RK, Barkan AL, Horowitz JF. Rapid
development of systemic insulin resistance with overeating is not
accompanied by robust changes in skeletal muscle glucose and lipid
metabolism. Appl Physiol Nutr Metab. 2013;38:512-519.
180. Gillberg L, Jacobsen SC, Ronn T, Brons C, Vaag A. PPARGC1A DNA
methylation in subcutaneous adipose tissue in low birth weight
subjects—impact of 5 days of high-fat overfeeding. Metabolism.
2014;63:263-271.
181. Jacobsen SC, Gillberg L, Bork-Jensen J, et al. Young men with low
birthweight exhibit decreased plasticity of genome-wide muscle
DNA methylation by high-fat overfeeding. Diabetologia. 2014;57:
1154-1158.
182. Lagerpusch M, Bosy-Westphal A, Kehden B, Peters A, Muller MJ.
Effects of brief perturbations in energy balance on indices of glu-
cose homeostasis in healthy lean men. Int J Obes (Lond). 2012;36:
1094-1101.
183. Le KA, Ith M, Kreis R, et al. Fructose overconsumption causes dys-
lipidemia and ectopic lipid deposition in healthy subjects with and
without a family history of type 2 diabetes. Am J Clin Nutr. 2009;89:
1760-1765.
184. Nestel PJ, Carroll KF, Havenstein N. Plasma triglyceride response
to carbohydrates, fats and caloric intake. Metabolism. 1970;19:
1-18.
185. Seematter G, Dirlewanger M, Rey V, Schneiter P, Tappy L. Metabolic
effects of mental stress during over- and underfeeding in healthy
women. Obes Res. 2002;10:49-55.
186. Seyssel K, Alligier M, Meugnier E, et al. Regulation of energy metab-
olism and mitochondrial function in skeletal muscle during lipid
overfeeding in healthy men. J Clin Endocrinol Metab. 2014;99:
E1254-E1262.
187. Vienberg SG, Brons C, Nilsson E, et al. Impact of short-term high-fat
feeding and insulin-stimulated FGF21 levels in subjects with low
birth weight and controls. Eur J Endocrinol. 2012;167:49-57.
188. Ribel-Madsen A, Hellgren LI, Brons C, et al. Plasma amino acid levels
are elevated in young, healthy low birth weight men exposed to
short-term high-fat overfeeding. Physiol Rep. 2016;4.
189. Heilbronn LK, Coster AC, Campbell LV, et al. The effect of short-
term overfeeding on serum lipids in healthy humans. Obesity (Silver
Spring). 2013;21:E649-E659.
190. Koopman KE, Caan MW, Nederveen AJ, et al. Hypercaloric diets
with increased meal frequency, but not meal size, increase
intrahepatic triglycerides: a randomized controlled trial. Hepatology.
2014;60:545-553.
191. Savage DB, Murgatroyd PR, Chatterjee VK, O'Rahilly S. Energy
expenditure and adaptive responses to an acute hypercaloric fat
load in humans with lipodystrophy. J Clin Endocrinol Metab. 2005;
90:1446-1452.
192. Sevastianova K, Santos A, Kotronen A, et al. Effect of short-term
carbohydrate overfeeding and long-term weight loss on liver fat in
overweight humans. Am J Clin Nutr. 2012;96:727-734.
193. Tam CS, Covington JD, Bajpeyi S, et al. Weight gain reveals dramatic
increases in skeletal muscle extracellular matrix remodeling. J Clin
Endocrinol Metab. 2014;99:1749-1757.
194. Theytaz F, Noguchi Y, Egli L, et al. Effects of supplementation with
essential amino acids on intrahepatic lipid concentrations during
73
fructose overfeeding in humans. Am J Clin Nutr. 2012;96:1008-
1016.
195. Fabbrini E, Tiemann Luecking C, Love-Gregory L, et al. Physiological
mechanisms of weight gain-induced steatosis in people with obesity.
Gastroenterology. 2016;150:79-81. e2
196. Morio B, Comte B, Martin JF, et al. Metabolomics reveals differen-
tial metabolic adjustments of normal and overweight subjects during
overfeeding. Metabolomics. 2015;11:920-938.
197. Knudsen SH, Hansen LS, Pedersen M, et al. Changes in insulin sensi-
tivity precede changes in body composition during 14 days of step
reduction combined with overfeeding in healthy young men. J Appl
Physiol (1985). 2012;113:7-15.
198. Heilbronn LK, Campbell LV, Xu A, Samocha-Bonet D. Metabolically
protective cytokines adiponectin and fibroblast growth factor-21
are increased by acute overfeeding in healthy humans. PLoS ONE.
2013;8:e78864.
199. Samocha-Bonet D, Campbell LV, Mori TA, et al. Overfeeding
reduces insulin sensitivity and increases oxidative stress, without
altering markers of mitochondrial content and function in humans.
PLoS ONE. 2012;7:e36320.
200. van der Meer RW, Hammer S, Lamb HJ, et al. Effects of short-term
high-fat, high-energy diet on hepatic and myocardial triglyceride con-
tent in healthy men. J Clin Endocrinol Metab. 2008;93:2702-2708.
201. Iggman D, Rosqvist F, Larsson A, et al. Role of dietary fats in modu-
lating cardiometabolic risk during moderate weight gain: a random-
ized double-blind overfeeding trial (LIPOGAIN study). J Am Heart
Assoc. 2014;3:e001095.
202. Hill JO, Peters JC, Swift LL, et al. Changes in blood lipids during six
days of overfeeding with medium or long chain triglycerides. J Lipid
Res. 1990;31:407-416.
203. Lagerpusch M, Enderle J, Eggeling B, et al. Carbohydrate quality and
quantity affect glucose and lipid metabolism during weight regain in
healthy men. J Nutr. 2013;143:1593-1601.
204. Kahlhofer J, Lagerpusch M, Enderle J, et al. Carbohydrate intake and
glycemic index affect substrate oxidation during a controlled weight
cycle in healthy men. Eur J Clin Nutr. 2014;68:1060-1066.
205. Karschin J, Lagerpusch M, Enderle J, et al. Endocrine determinants
of changes in insulin sensitivity and insulin secretion during a weight
cycle in healthy men. PLoS ONE. 2015;10:e0117865.
206. Breusing N, Lagerpusch M, Engstler AJ, et al. Influence of energy
balance and glycemic index on metabolic endotoxemia in healthy
men. J Am Coll Nutr. 2017;36:72-79.
207. Stanhope KL, Goran MI, Bosy-Westphal A, et al. Pathways and
mechanisms linking dietary components to cardiometabolic disease:
thinking beyond calories. Obes Rev. 2018;19:1205-1235.
208. Couchepin C, Le KA, Bortolotti M, et al. Markedly blunted metabolic
effects of fructose in healthy young female subjects compared with
male subjects. Diabetes Care. 2008;31:1254-1256.
209. Ibrahim M, Bonfiglio S, Schlogl M, et al. Energy expenditure and hor-
mone responses in humans after overeating high-fructose corn
syrup versus whole-wheat foods. Obesity (Silver Spring). 2018;26:
141-149.
210. Le KA, Faeh D, Stettler R, et al. A 4-wk high-fructose diet alters lipid
metabolism without affecting insulin sensitivity or ectopic lipids in
healthy humans. Am J Clin Nutr. 2006;84:1374-1379.
211. Seyssel K, Meugnier E, Le KA, et al. Fructose overfeeding in first-
degree relatives of type 2 diabetic patients impacts energy metabo-
lism and mitochondrial functions in skeletal muscle. Mol Nutr Food
Res. 2016;60:2691-2699.
212. McDevitt RM, Bott SJ, Harding M, et al. De novo lipogenesis during
controlled overfeeding with sucrose or glucose in lean and obese
women. Am J Clin Nutr. 2001;74:737-746.
213. Silbernagel G, Machann J, Unmuth S, et al. Effects of 4-week very-
high-fructose/glucose diets on insulin sensitivity, visceral fat and
intrahepatic lipids: an exploratory trial. Br J Nutr. 2011;106:79-86.
74
214. Lecoultre V, Carrel G, Egli L, et al. Coffee consumption attenuates
short-term fructose-induced liver insulin resistance in healthy men.
Am J Clin Nutr. 2014;99:268-275.
215. Bray GA, Redman LM, de Jonge L, et al. Plasma amino acids during
8 weeks of overfeeding: relation to diet body composition and
fat cell size in the proof study. Obesity (Silver Spring). 2018;26:
324-331.
216. Bray GA, Redman LM, de Jonge L, et al. Plasma fatty acyl-carnitines
during 8Weeks of overfeeding: relation to diet energy expenditure
and body composition: the PROOF study. Metabolism. 2018.
217. Roberts SB, Fuss P, Dallal GE, et al. Effects of age on energy expen-
diture and substrate oxidation during experimental overfeeding in
healthy men. J Gerontol A Biol Sci Med Sci. 1996;51:B148-B157.
218. Tam CS, Viardot A, Clement K, et al. Short-term overfeeding may
induce peripheral insulin resistance without altering subcutaneous adi-
pose tissue macrophages in humans. Diabetes. 2010;59:2164-2170.
219. Heymsfield SB, Peterson CM, Thomas DM, et al. Establishing energy
requirements for body weight maintenance: validation of an intake-
balance method. BMC Res Notes. 2017;10:220.
220. Chen M, Liu B, Thompson CH, Wittert GA, Heilbronn LK. Acute
overfeeding does not alter liver or adipose tissue-derived cytokines
in healthy humans. Ann Nutr Metab. 2016;69:165-170.
221. Cahill F, Amini P, Wadden D, et al. Short-term overfeeding increases
circulating adiponectin independent of obesity status. PLoS ONE.
2013;8:e74215.
222. Cahill F, Shea JL, Randell E, Vasdev S, Sun G. Serum peptide YY in
response to short-term overfeeding in young men. Am J Clin Nutr.
2011;93:741-747.
223. Rosenbaum M, Hirsch J, Murphy E, Leibel RL. Effects of changes in
body weight on carbohydrate metabolism, catecholamine excretion,
and thyroid function. Am J Clin Nutr. 2000;71:1421-1432.
224. Shea J, Randell E, Vasdev S, et al. Serum retinol-binding protein
4 concentrations in response to short-term overfeeding in normal-
weight, overweight, and obese men. Am J Clin Nutr. 2007;86:1310-
1315.
225. Smith GI, Magkos F, Reeds DN, et al. One day of mixed meal over-
feeding reduces hepatic insulin sensitivity and increases VLDL parti-
cle but not VLDL-triglyceride secretion in overweight and obese
men. J Clin Endocrinol Metab. 2013;98:3454-3462.
226. Shea J, French CR, Bishop J, et al. Changes in the transcriptome of
abdominal subcutaneous adipose tissue in response to short-term
overfeeding in lean and obese men. Am J Clin Nutr. 2009;89:
407-415.
227. Wadden D, Cahill F, Amini P, et al. Serum acylated ghrelin concen-
trations in response to short-term overfeeding in normal weight,
overweight, and obese men. PLoS ONE. 2012;7:e45748.
228. Cornier MA, Bergman BC, Bessesen DH. The effects of short-term
overfeeding on insulin action in lean and reduced-obese individuals.
Metabolism. 2006;55:1207-1214.
229. Reinhardt M, Thearle MS, Ibrahim M, et al. A human thrifty pheno-
type associated with less weight loss during caloric restriction. Dia-
betes. 2015;64:2859-2867.
230. Reinhardt M, Schlogl M, Bonfiglio S, et al. Lower core body tempera-
ture and greater body fat are components of a human thrifty pheno-
type. Int J Obes (Lond). 2016;40:754-760.
231. Piening BD, Zhou W, Contrepois K, et al. Integrative personal omics
profiles during periods of weight gain and loss. Cell Syst. 2018;6:
1-14.
232. Jenkins AB, Batterham M, Samocha-Bonet D, et al. Segregation of a
latent high adiposity phenotype in families with a history of type
2 diabetes mellitus implicates rare obesity-susceptibility genetic var-
iants with large effects in diabetes-related obesity. PLoS ONE. 2013;
8:e70435.
233. Sato K, Samocha-Bonet D, Handelsman DJ, et al. Serum sex steroids
and steroidogenesis-related enzyme expression in skeletal muscle
BRAY AND BOUCHARD
during experimental weight gain in men. Diabetes Metab. 2014;40:
439-444.
234. DeLany JP, Bray GA, Harsha DW, Volaufova J. Energy expenditure
in African American and white boys and girls in a 2-y follow-up of
the Baton Rouge Children's Study. Am J Clin Nutr. 2004;79:
268-273.
235. Ravussin E, Lillioja S, Knowler WC, et al. Reduced rate of energy
expenditure as a risk factor for body-weight gain. N Engl J Med.
1988;318:467-472.
236. Roberts SB, Savage J, Coward WA, Chew B, Lucas A. Energy expen-
diture and intake in infants born to lean and overweight mothers. N
Engl J Med. 1988;318:461-466.
237. Barker DJ, Gluckman PD, Godfrey KM, et al. Fetal nutrition and car-
diovascular disease in adult life. Lancet. 1993;341:938-941.
238. Wadhwa PD, Buss C, Entringer S, Swanson JM. Developmental ori-
gins of health and disease: brief history of the approach and current
focus on epigenetic mechanisms. Semin Reprod Med. 2009;27:
358-368.
239. Ribel-Madsen A, Ribel-Madsen R, Brons C, et al. Plasma
acylcarnitine profiling indicates increased fatty acid oxidation rela-
tive to tricarboxylic acid cycle capacity in young, healthy low birth
weight men. Physiol Rep. 2016;4.
240. Calles-Escandon J, Goran MI, O'Connell M, Nair KS, Danforth E Jr.
Exercise increases fat oxidation at rest unrelated to changes in energy
balance or lipolysis. Am J Physiol. 1996;270:E1009-E1014.
241. Goran MI. Variation in total energy expenditure in humans. Obes
Res. 1995;3(Suppl 1):59-66.
242. Wulan SN, Schrauwen-Hinderling VB, Westerterp KR, Plasqui G.
Liver fat accumulation in response to overfeeding with a high-fat
diet: a comparison between South Asian and Caucasian men. Nutr
Metab (Lond). 2015;12:18.
243. Sims EA, Danforth E Jr, Horton ES, et al. Endocrine and metabolic
effects of experimental obesity in man. Recent Prog Horm Res. 1973;
29:457-496.
244. Salans LB, Zarnowski MJ, Segal R. Effect of insulin upon the cellular
character of rat adipose tissue. J Lipid Res. 1972;13:616-623.
245. Sims EA, Danforth E Jr, Horton ES, et al. Experimental obesity in
man. A progress report. Isr J Med Sci. 1972;8:813-814.
246. Sims EA, Horton ES. Endocrine and metabolic adaptation to obesity
and starvation. Am J Clin Nutr. 1968;21:1455-1470.
247. Sims EA, Horton ES, Salans LB. Inducible metabolic abnormalities
during development of obesity. Annu Rev Med. 1971;22:235-250.
248. Votruba SB, Jensen MD. Insulin sensitivity and regional fat gain
in response to overfeeding. Obesity (Silver Spring). 2011;19:
269-275.
249. Davidson MB, Chopra IJ. Effect of carbohydrate and noncarbohydrate
sources of calories on plasma 3,5,3'-triiodothyronine concentrations in
man. J Clin Endocrinol Metab. 1979;48:577-581.
250. Oppert JM, Dussault JH, Tremblay A, et al. Thyroid hormones and
thyrotropin variations during long term overfeeding in identical
twins. J Clin Endocrinol Metab. 1994;79:547-553.
251. Bray GA, Fisher DA, Chopra IJ. Relation of thyroid hormones to
body-weight. Lancet. 1976;1(June 5):1026-1208.
252. Muller MJ, Enderle J, Pourhassan M, et al. Metabolic adaptation
to caloric restriction and subsequent refeeding: the Minnesota
Starvation Experiment revisited. Am J Clin Nutr. 2015;102:
807-819.
253. Tremblay A, Poehlman ET, Nadeau A, Dussault J, Bouchard C.
Heredity and overfeeding-induced changes in submaximal exercise
VO2. J Appl Physiol (1985). 1987;62:539-544.
254. Welle S, Campbell R. Effect of overeating on plasma and urinary
concentrations of norepinephrine. J Clin Endocrinol Metab. 1984;59:
531-534.
255. Forbes GB, Welle SL. Lean body mass in obesity. Int J Obes (Lond).
1983;7:99-107.
BRAY AND BOUCHARD
256. Scherrer U, Randin D, Tappy L, et al. Body fat and sympathetic nerve
activity in healthy subjects. Circulation. 1994;89:2634-2640.
257. Arrone LJ, Mackintosh R, Rosenbaum M, Leibel RL, Hirsch J. Cardiac
autonomic nervous system activity in obese and never-obese young
men. Obes Res. 1997;5:354-359.
258. Arone LJ, Mackintosh R, Rosenbaum M, Leibel RL, Hirsch J. Auto-
nomic nervous system activity in weight gain and weight loss.
Am J Physiol. 1995;269:R222-R225.
259. O'Connell M, Danforth E Jr, Horton ES, Salans L, Sims EA. Experi-
mental obesity in man. 3. Adrenocortical function. J Clin Endocrinol
Metab. 1973;36:323-329.
260. Pritchard J, Despres JP, Gagnon J, et al. Plasma adrenal, gonadal,
and conjugated steroids before and after long-term overfeeding in
identical twins. J Clin Endocrinol Metab. 1998;83:3277-3284.
261. Parry SA, Woods RM, Hodson L, Hulston CJ. A single day of exces-
sive dietary fat intake reduces whole-body insulin sensitivity: the
metabolic consequence of binge eating. Nutrients. 2017;9.
262. Felig P, Horton ES, Runge CF, Sims EA. Experimental obeisty in man:
hyperaminoacidemia and diminished effectiveness of insulin in regu-
lating peripheral amino acid release. Pgm Endocr Soc. 1971;(Abstract
29):A57.
263. Rabinowitz D, Zierler KL. Forearm metabolism in obesity and its
response to intra-arterial insulin. Characterization of insulin resis-
tance and evidence for adaptive hyperinsulinism. J Clin Invest. 1962;
41(12):2173-2181.
264. Cornier MA, Bessesen DH, Gurevich I, Leitner JW, Draznin B. Nutri-
tional upregulation of p85alpha expression is an early mole-
cular manifestation of insulin resistance. Diabetologia. 2006;49:
748-754.
265. Laeger T, Henagan TM, Albarado DC, et al. FGF21 is an endocrine
signal of protein restriction. J Clin Invest. 2014;124:3913-3922.
266. Davidson MB, Forsythe AB. Dietary and anticholinergic effects on
intravenous glucose tolerance and insulin secretion. Int J Obes
(Lond). 1978;2:309-320.
267. Singh P, Sharma P, Sahakyan KR, et al. Differential effects of leptin
on adiponectin expression with weight gain versus obesity. Int J
Obes (Lond). 2016;40:266-274.
268. Cobelli C, Toffolo GM, Dalla Man C, et al. Assessment of beta-cell
function in humans, simultaneously with insulin sensitivity and
hepatic extraction, from intravenous and oral glucose tests.
Am J Physiol Endocrinol Metab. 2007;293:E1-E15.
269. Oppert JM, Nadeau A, Tremblay A, et al. Plasma glucose, insulin,
and glucagon before and after long-term overfeeding in identical
twins. Metabolism. 1995;44:96-105.
270. Hagobian TA, Sharoff CG, Braun B. Effects of short-term exercise
and energy surplus on hormones related to regulation of energy bal-
ance. Metabolism. 2008;57:393-398.
271. Halliday TM, Rynders CA, Thomas E, et al. Appetite-related
responses to overfeeding and longitudinal weight change in obesity-
prone and obesity-resistant adults. Obesity (Silver Spring). 2020;28:
259-267.
272. Wadden D, Cahill F, Amini P, et al. Circulating glucagon-like peptide-
1 increases in response to short-term overfeeding in men. Nutr
Metab (Lond). 2013;10:33.
273. Schwartz MW, Seeley RJ, Zeltser LM, et al. Obesity pathogenesis:
an endocrine society scientific statement. Endocr Rev. 2017;38:
267-296.
274. Cornier MA, McFadden KL, Thomas EA, et al. Differences in the
neuronal response to food in obesity-resistant as compared to
obesity-prone individuals. Physiol Behav. 2013;110-111:122-128.
275. Cornier MA, Von Kaenel SS, Bessesen DH, Tregellas JR. Effects of
overfeeding on the neuronal response to visual food cues. Am J Clin
Nutr. 2007;86:965-971.
75
276. Cornier MA, Salzberg AK, Endly DC, et al. The effects of overfeeding
on the neuronal response to visual food cues in thin and reduced-
obese individuals. PLoS ONE. 2009;4:e6310.
277. Cornier MA, McFadden KL, Thomas EA, et al. Propensity to obesity
impacts the neuronal response to energy imbalance. Front Behav
Neurosci. 2015;9:52.
278. Bray GA, Heisel WE, Afshin A, et al. The science of obesity manage-
ment: an endocrine society scientific statement. Endocr Rev. 2018;
39:79-132.
279. Poirier P, Giles TD, Bray GA, et al. Obesity and cardiovascular dis-
ease: pathophysiology, evaluation, and effect of weight loss: an
update of the 1997 American Heart Association Scientific State-
ment on Obesity and Heart Disease from the Obesity Committee of
the Council on Nutrition, Physical Activity, and Metabolism. Circula-
tion. 2006;113:898-918.
280. Gupta N, Jensen MD. Clinical effects of high-fat meals and weight
gain due to high-fat feeding. Int J Obes Suppl. 2012;2:S51-S55.
281. Peterson CM, Zhang B, Johannsen DL, Ravussin E. Eight weeks of
overfeeding alters substrate partitioning without affecting metabolic
flexibility in men. Int J Obes (Lond). 2017;41:887-893.
282. Covington JD, Bray GA, Redman LM, Johannsen DL, Ravussin E.
Eight weeks of dietary overfeeding increases renal filtration rates in
humans: implications for the pathogenesis of diabetic hyper-
filtration. J Intern Med. 2015;278:396-400.
283. Gupta AK, Johnson WD, Johannsen D, Ravussin E. Cardiovascular
risk escalation with caloric excess: a prospective demonstration of
the mechanics in healthy adults. Cardiovasc Diabetol. 2013;12:23.
284. Sacks FM, Lichtenstein AH, Wu JHY, et al. Dietary fats and cardio-
vascular disease: a presidential advisory from the american heart
association. Circulation. 2017;136:e1-e23.
285. Siri-Tarino P, Kraus RM. The evolving role of saturated fatty acids. In:
Bergeron N, Siri-Tarino PW, Bray GA, Krauss RM, eds. Nutrition and
Cardiometabolic Health. Boca Raton, FL: CRC Press; 2017:209-222.
286. Bellou E, Maraki M, Magkos F, et al. Effect of acute negative and
positive energy balance on basal very-low density lipoprotein tri-
glyceride metabolism in women. PLoS ONE. 2013;8:e60251.
287. Bian H, Hakkarainen A, Lundbom N, Yki-Jarvinen H. Effects of die-
tary interventions on liver volume in humans. Obesity (Silver Spring).
2014;22:989-995.
288. Bouchard C, Tremblay A, Despres JP, et al. Overfeeding in identical
twins: 5-year postoverfeeding results. Metabolism. 1996;45:1042-
1050.
289. Chin-Chance C, Polonsky KS, Schoeller DA. Twenty-four-hour leptin
levels respond to cumulative short-term energy imbalance and pre-
dict subsequent intake. J Clin Endocrinol Metab. 2000;85:2685-
2691.
290. Covington JD, Johannsen DL, Coen PM, et al. Intramyocellular lipid
droplet size rather than total lipid content is related to insulin sensi-
tivity after 8 weeks of overfeeding. Obesity (Silver Spring). 2017;25:
2079-2087.
291. Despres JP, Poehlman ET, Tremblay A, et al. Genotype-influenced
changes in serum HDL cholesterol after short-term overfeeding in
man: association with plasma insulin and triglyceride levels. Metabo-
lism. 1987;36:363-368.
292. Elshorbagy AK, Jerneren F, Samocha-Bonet D, Refsum H,
Heilbronn LK. Serum S-adenosylmethionine, but not methionine,
increases in response to overfeeding in humans. Nutr Diabetes.
2016;6:e192.
293. Gillberg L, Perfilyev A, Brons C, et al. Adipose tissue transcriptomics
and epigenomics in low birthweight men and controls: role of high-
fat overfeeding. Diabetologia. 2016;59:799-812.
294. Jacobsen SC, Brons C, Bork-Jensen J, et al. Effects of short-term
high-fat overfeeding on genome-wide DNA methylation in the
76
skeletal muscle of healthy young men. Diabetologia. 2012;55:3341-
3349.
295. Minehira K, Vega N, Vidal H, Acheson K, Tappy L. Effect of carbohy-
drate overfeeding on whole body macronutrient metabolism and
expression of lipogenic enzymes in adipose tissue of lean and
overweight humans. Int J Obes Relat Metab Disord. 2004;28:1291-
1298.
296. Orr JS, Gentile CL, Davy BM, Davy KP. Large artery stiffening with
weight gain in humans: role of visceral fat accumulation. Hyperten-
sion. 2008;51:1519-1524.
297. Perfilyev A, Dahlman I, Gillberg L, et al. Impact of polyunsaturated
and saturated fat overfeeding on the DNA-methylation pattern in
human adipose tissue: a randomized controlled trial. Am J Clin Nutr.
2017;105:991-1000.
298. Peterson CM, Orooji M, Johnson DN, Naraghi-Pour M, Ravussin E.
Brown adipose tissue does not seem to mediate metabolic adapta-
tion to overfeeding in men. Obesity (Silver Spring). 2017;25:502-505.
299. Ribel-Madsen R, Brons C, Friedrichsen M, Poulsen P, Vaag A. Reti-
nol-binding protein 4 in young men with low versus normal birth
weight: physiological response to short-term overfeeding. Obesity
(Silver Spring). 2011;19:1304-1306.
300. Teran-Garcia M, Despres JP, Couillard C, Tremblay A, Bouchard C.
Effects of long-term overfeeding on plasma lipoprotein levels in
identical twins. Atherosclerosis. 2004;173:277-283.
301. Teran-Garcia M, Despres JP, Tremblay A, Bouchard C. Effects of cho-
lesterol ester transfer protein (CETP) gene on adiposity in response to
long-term overfeeding. Atherosclerosis. 2008;196:455-460.
302. Ukkola O, Chagnon M, Tremblay A, Bouchard C. Genetic variation
at the adipsin locus and response to long-term overfeeding. Eur J
Clin Nutr. 2003;57:1073-1078.
303. Ukkola O, Joanisse DR, Tremblay A, Bouchard C. Na+-K+-ATPase
alpha 2-gene and skeletal muscle characteristics in response to long-
term overfeeding. J Appl Physiol (1985). 2003;94:1870-1874.
304. Ukkola O, Tremblay A, Bouchard C. Lipoprotein lipase polymor-
phisms and responses to long-term overfeeding. J Intern Med. 2002;
251:429-436.
305. Ukkola O, Tremblay A, Despres JP, et al. Leptin receptor Gln223Arg
variant is associated with a cluster of metabolic abnormalities in
response to long-term overfeeding. J Intern Med. 2000;248:
435-439.
306. Fantino M, Baigts F, Cabanac M, Apfelbaum M. Effects of an over-
feeding regimen—the affective component of the sweet sensation.
Appetite. 1983;4:155-164.
307. Ukkola O, Rosmond R, Tremblay A, Bouchard C. Glucocorticoid
receptor Bcl I variant is associated with an increased atherogenic
profile in response to long-term overfeeding. Atherosclerosis. 2001;
157:221-224.
308. Johnston RD, Stephenson MC, Crossland H, et al. No difference
between high-fructose and high-glucose diets on liver triacylglycerol
or biochemistry in healthy overweight men. Gastroenterology. 2013;
145:1016-1025. e2
309. Acheson KJ, Schutz Y, Bessard T, Flatt JP, Jequier E. Carbohydrate-
metabolism and denovo lipogenesis in human obesity. Am J Clin
Nutr. 1987;45:78-85.
310. Angel A, Bray GA. Synthesis of fatty acids and cholesterol by liver,
adipose tissue and intestinal mucosa from obese and control
patients. Eur J Clin Invest. 1979;9:355-362.
311. Birkenfeld AL, Shulman GI. Nonalcoholic fatty liver disease, hepatic
insulin resistance, and type 2 diabetes. Hepatology. 2014;59:
713-723.
312. Fabbrini E, Magkos F, Mohammed BS, et al. Intrahepatic fat, not vis-
ceral fat, is linked with metabolic complications of obesity. Proc Natl
Acad Sci U S A. 2009;106:15430-15435.
313. Spalding KL, Arner E, Westermark PO, et al. Dynamics of fat cell
turnover in humans. Nature. 2008;453:783-787.
BRAY AND BOUCHARD
314. Kashiwagi A, Mott D, Bogardus C, et al. The effects of short-term
overfeeding on adipocyte metabolism in Pima Indians. Metabolism.
1985;34:364-370.
315. Mauriege P, Despres JP, Marcotte M, et al. Adipose tissue lipolysis
after long-term overfeeding in identical twins. Int J Obes Relat Metab
Disord. 1992;16:219-225.
316. Salans LB, Bray GA, Cushman SW, et al. Glucose metabolism and
the response to insulin by human adipose tissue in spontaneous and
experimental obesity. Effects of dietary composition and adipose
cell size. J Clin Invest. 1974;53:848-856.
317. Salans LB, Horton ES, Sims EA. Experimental obesity in man:
cellular character of the adipose tissue. J Clin Invest. 1971;50:1005-
1011.
318. Hammarstedt A, Gogg S, Hedjazifar S, Nerstedt A, Smith U. Impaired
adipogenesis and dysfunctional adipose tissue in human hypertro-
phic obesity. Physiol Rev. 2018;98:1911-1941.
319. Levine JA, Jensen MD, Eberhardt NL, O'Brien T. Adipocyte macro-
phage colony-stimulating factor is a mediator of adipose tissue
growth. J Clin Invest. 1998;101:1557-1564.
320. Stock MJ, Rothwell NJ. Role of brown adipose tissue thermogenesis
in overfeeding: a review. J R Soc Med. 1983;76:71-73.
321. Cypess AM, Lehman S, Williams G, et al. Identification and impor-
tance of brown adipose tissue in adult humans. N Engl J Med. 2009;
360:1509-1517.
322. Saito M, Okamatsu-Ogura Y, Matsushita M, et al. High incidence of
metabolically active brown adipose tissue in healthy adult humans:
effects of cold exposure and adiposity. Diabetes. 2009;58:1526-
1531.
323. van Marken Lichtenbelt WD, Vanhommerig JW, Smulders NM, et al.
Cold-activated brown adipose tissue in healthy men. N Engl J Med.
2009;360:1500-1508.
324. Wu J, Bostrom P, Sparks LM, et al. Beige adipocytes are a distinct type
of thermogenic fat cell in mouse and human. Cell. 2012;150:366-376.
325. Ikeda K, Maretich P, Kajimura S. The common and distinct features
of brown and beige adipocytes. Trends Endocrinol Metab. 2018;29:
191-200.
326. Datta R, Podolsky MJ, Atabai K. Fat fibrosis: friend or foe? JCI
Insight. 2018;3.
327. Mariman EC, Wang P. Adipocyte extracellular matrix composition,
dynamics and role in obesity. Cell Mol Life Sci. 2010;67:1277-
1292.
328. Lin D, Chun TH, Kang L. Adipose extracellular matrix remodelling in
obesity and insulin resistance. Biochem Pharmacol. 2016;119:8-16.
329. Spencer M, Unal R, Zhu B, et al. Adipose tissue extracellular matrix
and vascular abnormalities in obesity and insulin resistance. J Clin
Endocrinol Metab. 2011;96:E1990-E1998.
330. Pasarica M, Gowronska-Kozak B, Burk D, et al. Adipose tissue
collagen VI in obesity. J Clin Endocrinol Metab. 2009;94:5155-5162.
331. Rutkowski JM, Stern JH, Scherer PE. The cell biology of fat expan-
sion. J Cell Biol. 2015;208:501-512.
332. Nielsen TS, Jessen N, Jorgensen JO, Moller N, Lund S. Dissecting
adipose tissue lipolysis: molecular regulation and implications for
metabolic disease. J Mol Endocrinol. 2014;52:R199-R222.
333. Wang H, Eckel RH. Lipoprotein lipase: from gene to obesity.
Am J Physiol Endocrinol Metab. 2009;297:E271-E288.
334. Bouchard C, Tremblay A, Despres JP, et al. Sensitivity to overfeed-
ing: the Quebec experiment with identical twins. Prog Food Nutr Sci.
1988;12:45-72.
335. Saltiel AR, Olefsky JM. Inflammatory mechanisms linking obesity
and metabolic disease. J Clin Invest. 2017;127:1-4.
336. Samocha-Bonet D, Tam CS, Campbell LV, Heilbronn LK. Raised cir-
culating fetuin-a after 28-day overfeeding in healthy humans. Diabe-
tes Care. 2014;37:e15-e16.
337. Tam CS, Chaudhuri R, Hutchison AT, Samocha-Bonet D,
Heilbronn LK. Skeletal muscle extracellular matrix remodeling after
BRAY AND BOUCHARD
short-term overfeeding in healthy humans. Metabolism. 2017;67:
26-30.
338. Cook KS, Groves DL, Min HY, Spiegelman BM. A developmentally
regulated mRNA from 3T3 adipocytes encodes a novel serine prote-
ase homologue. Proc Natl Acad Sci U S A. 1985;82:6480-6484.
339. Zhang Y, Proenca R, Maffei M, et al. Positional cloning of the
mouse obese gene and its human homologue. Nature. 1994;372:
425-432.
340. Lehr S, Hartwig S, Lamers D, et al. Identification and validation of
novel adipokines released from primary human adipocytes. Mol Cell
Proteomics. 2012;11:M111.010504.
341. Antuna-Puente B, Feve B, Fellahi S, Bastard JP. Adipokines: the
missing link between insulin resistance and obesity. Diabetes Metab.
2008;34:2-11.
342. Balistreri CR, Caruso C, Candore G. The role of adipose tissue and
adipokines in obesity-related inflammatory diseases. Mediators
Inflamm. 2010;2010:802078.
343. Nicholson T, Church C, Baker DJ, Jones SW. The role of adipokines
in skeletal muscle inflammation and insulin sensitivity. J Inflamm-
Lond. 2018;15.
344. Farooqi IS, O'Rahilly S. 20 years of leptin: human disorders of leptin
action. J Endocrinol. 2014;223:T63-T70.
345. Rosenbaum M, Nicolson M, Hirsch J, et al. Effects of weight change
on plasma leptin concentrations and energy expenditure. J Clin
Endocrinol Metab. 1997;82:3647-3654.
346. Considine RV, Premkumar A, Reynolds JC, et al. Adiponectin
and leptin in African Americans. Obesity (Silver Spring). 2008;16:
428-434.
347. Rosenbaum M, Pietrobelli A, Vasselli JR, Heymsfield SB, Leibel RL.
Sexual dimorphism in circulating leptin concentrations is not
accounted for by differences in adipose tissue distribution. Int J
Obes Relat Metab Disord. 2001;25:1365-1371.
348. Joosen AM, Bakker AH, Zorenc AH, et al. PPARgamma activity in
subcutaneous abdominal fat tissue and fat mass gain during short-
term overfeeding. Int J Obes (Lond). 2006;30:302-307.
349. Goldsmith R, Joanisse DR, Gallagher D, et al. Effects of experimental
weight perturbation on skeletal muscle work efficiency, fuel utiliza-
tion, and biochemistry in human subjects. Am J Physiol Regul Integr
Comp Physiol. 2010;298:R79-R88.
350. Ravussin E, Tschop M, Morales S, Bouchard C, Heiman ML. Plasma
ghrelin concentration and energy balance: overfeeding and negative
energy balance studies in twins. J Clin Endocrinol Metab. 2001;86:
4547-4551.
351. Ukkola O, Teran-Garcia M, Tremblay A, Despres JP, Bouchard C.
Adiponectin concentration and insulin indicators following over-
feeding in identical twins. J Endocrinol Invest. 2008;31:132-137.
352. Segrestin B, Moreno-Navarrete JM, Seyssel K, et al. Adipose tissue
expansion by overfeeding healthy men alters iron gene expression.
J Clin Endocrinol Metab. 2018.
353. Deriaz O, Fournier G, Tremblay A, Despres JP, Bouchard C.
Lean-body-mass composition and resting energy expenditure
before and after long-term overfeeding. Am J Clin Nutr. 1992;56:
840-847.
354. Sun G, Ukkola O, Rankinen T, Joanisse DR, Bouchard C. Skeletal
muscle characteristics predict body fat gain in response to overfeed-
ing in never-obese young men. Metabolism. 2002;51:451-456.
355. Sparks LM, Xie H, Koza RA, et al. A high-fat diet coordinately down-
regulates genes required for mitochondrial oxidative phosphoryla-
tion in skeletal muscle. Diabetes. 2005;54:1926-1933.
356. Miller DS, Mumford P, Stock MJ. Gluttony. 2. Thermogenesis in
overeating man. Am J Clin Nutr. 1967;20:1223-1229.
357. Hanson JS. Exercise responses following production of experimental
obesity. J Appl Physiol. 1973;35:587-591.
358. Welle SL, Seaton TB, Campbell RG. Some metabolic effects of over-
eating in man. Am J Clin Nutr. 1986;44:718-724.
77
359. Acheson K, Jequier E, Burger A, Danforth E. Thyroid-Hormones and
Thermogenesis - the Metabolic Cost of Food and Exercise. Metabol
Clin Exp. 1984;33:262-265.
360. Rosenbaum M, Vandenborne K, Goldsmith R, et al. Effects of experi-
mental weight perturbation on skeletal muscle work efficiency in
human subjects. Am J Physiol Regul Integr Comp Physiol. 2003;285:
R183-R192.
361. Bray GA. The energetics of obesity. Med Sci Sports Exerc. 1983;15:
32-40.
362. Bray GA. The pain of weight gain: self-experimentation with over-
feeding. Am J Clin Nutr. 2019.
363. Ernersson A, Nystrom FH, Lindstrom T. Long-term increase of fat
mass after a four week intervention with fast food based hyper-
alimentation and limitation of physical activity. Nutr Metab. 2010;7:
68-76.
364. Erlingsson S, Herard S, Dahlqvist Leinhard O, et al. Men develop
more intraabdominal obesity and signs of the metabolic syndrome
after hyperalimentation than women. Metabolism. 2009;58:995-
1001.
365. Kechagias S, Ernersson A, Dahlqvist O, et al. Fast-food-based hyper-
alimentation can induce rapid and profound elevation of serum ala-
nine aminotransferase in healthy subjects. Gut. 2008;57:649-654.
366. Hamill PV, Drizd TA, Johnson CL, Reed RB, Roche AF. NCHS growth
curves for children birth-18 years. United States. Vital Health Stat.
1977: i-iv;11:1-74.
367. Rynders CA, Bergouignan A, Kealey E, Bessesen DH. Ability to adjust
nocturnal fat oxidation in response to overfeeding predicts 5-year
weight gain in adults. Obesity (Silver Spring). 2017;25:873-880.
368. Thomas EA, Bechtell JL, Vestal BE, et al. Eating-related behaviors
and appetite during energy imbalance in obese-prone and obese-
resistant individuals. Appetite. 2013;65:96-102.
369. Stunkard AJ, Messick S. The three-factor eating questionnaire to
measure dietary restraint, disinhibition and hunger. J Psychosom Res.
1985;29:71-83.
370. Levitsky DA, Obarzanek E, Mrdjenovic G, Strupp BJ. Imprecise con-
trol of energy intake: absence of a reduction in food intake following
overfeeding in young adults. Physiol Behav. 2005;84:669-675.
371. Bray GA, Flatt JP, Volaufova J, Delany JP, Champagne CM. Correc-
tive responses in human food intake identified from an analysis of
7-d food-intake records. Am J Clin Nutr. 2008;88:1504-1510.
372. Ukkola O, Kesaniemi YA, Tremblay A, Bouchard C. Two variants in
the resistin gene and the response to long-term overfeeding. Eur J
Clin Nutr. 2004;58:654-659.
373. Ukkola O, Sun G, Bouchard C. Insulin-like growth factor 2 (IGF2)
and IGF-binding protein 1 (IGFBP1) gene variants are associated
with overfeeding-induced metabolic changes. Diabetologia. 2001;44:
2231-2236.
374. Ukkola O, Tremblay A, Bouchard C. Beta-2 adrenergic receptor vari-
ants are associated with subcutaneous fat accumulation in response
to long-term overfeeding. Int J Obes Relat Metab Disord. 2001;25:
1604-1608.
375. Ukkola O, Tremblay A, Sun G, Chagnon YC, Bouchard C. Genetic
variation at the uncoupling protein 1, 2 and 3 loci and the response
to long-term overfeeding. Eur J Clin Nutr. 2001;55:1008-1015.
376. Ukkola O, Bouchard C. Role of candidate genes in the responses to
long-term overfeeding: review of findings. Obes Rev. 2004;5:3-12.
377. Redden DT, Allison DB. The Quebec overfeeding study: a catalyst
for new hypothesis generation. Obes Rev. 2004;5:1-2.
378. Young JB, Landsberg L. Stimulation of the sympathetic nervous sys-
tem during sucrose feeding. Nature. 1977;269:615-617.
379. Harper ME, Green K, Brand MD. The efficiency of cellular energy
transduction and its implications for obesity. Annu Rev Nutr. 2008;
28:13-33.
380. Newsholme EA. Sounding board. A possible metabolic basis for the
control of body weight. N Engl J Med. 1980;302:400-405.
78 BRAY AND BOUCHARD

381. Gebauer J, Schuster S, de Figueiredo LF, Kaleta C. Detecting and

investigating substrate cycles in a genome-scale human metabolic How to cite this article: Bray GA, Bouchard C. The biology of
network. FEBS J. 2012;279:3192-3202.
human overfeeding: A systematic review. Obesity Reviews.
2020;1–78. https://doi.org/10.1111/obr.13040
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