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Prostaglandins, Leukotrienes and Essential Fatty Acids 73 (2005) 9–15

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The adipose organ

S. Cinti
University of Ancona, Italy, European Union

Abstract

In mammals, the adipose tissues are contained in a multi-depot organ: the adipose organ. It consists of several subcutaneous and
visceral depots. Some areas of these depots are brown and correspond to brown adipose tissue, while many are white and
correspond to white adipose tissue. The organ is rich of vessels and parenchymal nerve fibers, but their density is higher in the brown
areas. White areas contain a variable amount of brown adipocytes and their number varies with age, strain and environmental
conditions. All adipocytes of the adipose organ express a specific adrenoceptor: X3AR. Recent data have stressed the plasticity of the
adipose organ in adult animals, and in parallel with the cytological variations there are also vascular as well as neural variations. Of
note, treatment of genetically and diet induced obese rats with X3 adrenoceptor agonists ameliorate their pathological condition and
this is accompanied by the appearance of brown adipocytes in white areas of the adipose organ. This drug-induced modification of
the anatomy of the organ is also obtained by the treatment with PPARg agonists in rats and dogs.
We have previously shown that the transformation of white adipose tissue into brown adipose tissue in rats treated with X3
adrenoceptor agonists is due to a direct transformation of differentiated unilocular adipocytes (transdifferentiation). We recently
also showed that the absence of X3 adrenoceptors strongly depress this type of plasticity in the adipose organ. All together these
experiments strongly suggest the possibility to modulate the plasticity of the adipose organ with therapeutic implications for obesity
and related disorders.
r 2005 Elsevier Ltd. All rights reserved.

1. Anatomy Both cell types share the unusual property of

It is well known that mammals have two distinct types
of adipocytes: white and brown. These cells have distinct
anatomy and function: white adipocytes are unilocular
(Fig. 1) and allow fatty acid accumulation after a meal
and they distribute fuel (fatty acids) to the organism
between meals. White adipocyte mass controls, by
secreting the hormone leptin [1], the individual beha-
viour concerning food research and intake.
Brown adipocytes are multilocular and rich of big
characteristic mitochondria (Fig. 2) containing the
protein UCP1 (uniquely expressed in these cells)
responsible for uncoupling of the oxidative phosphor-
ylation that in turn is responsible for the heat produc-
tion of this cell type.
Corresponding author. Tel.: +39 071 2206088;
fax: +39 071 2206088.
E-mail address: cinti@univpm.it.
accumulation and release of fatty acids that are at the
base of their activity, furthermore they both express the
rather specific adrenergic receptor b3 [2]. These two cell
types, organised in two tissues, the white adipose tissue
(WAT) and brown adipose tissue (BAT) are contained
into a multidepot organ: the adipose organ.
This organ is composed by several subcutaneous and
visceral depots (Fig. 3). Rodents have two main
subcutaneous fat depots, one anterior and one posterior,
lying in discrete anatomical locations. The anterior
depot is the more complex one; it lies at the base of the
forelimbs and mainly occupies the dorsal body region
between and under the scapulae, the axillary and
proximal regions of the forelimbs and the cervical area.
The interscapular area represents the more conspicuous
central portion of the depot (body) and extends laterally
(lateral wings) and ahead (anterior wings). The anterior
wings extend ahead among the superficial dorsal muscles
and reach the base of the neck.

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doi:10.1016/j.plefa.2005.04.010
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10 S. Cinti / Prostaglandins, Leukotrienes and Essential Fatty Acids 73 (2005) 9–15
Fig. 1. Light microscopy. Haematoxylin–eosin staining. Human white
adipose tissue. Objective magnification 20
.
Fig. 2. Scanning electron microscopy. Brown adipocyte mitochondria.
Scale bar: 333 nm.
The posterior depot is located at the base of the hind
legs. Its anatomy is simpler, consisting of a single tissue
band beginning from the dorsum at the lumbar level
(dorso-lumbar portion), extending ahead in the ingui-
no–crural region (inguinal portion) up to the pubic level
into the gluteal region (gluteal portion). At the pubic
level, this depot joins the contralateral depot.
Recently, we observed that subfascial depots are
present in the limbs. Two main limbs depots are located
one at the root of the limb and the second at the level of
the join in the middle of the limbs. The join depot at the
knee level (popliteal depot) contains a lymphnode.
The visceral depots are located into the thorax and
abdomen cavities. The former lies mainly among the
proximal tracts of the intercostal nerve-vascular bun-
dles, the heart and the aorta. The abdominal depots can
be retro- or intraperitoneal. The retroperitoneal depot
Fig. 3. The adipose organ of an adult Sv129 mouse maintained at
29 1C for 10 days. The organ has been dissected with the aid of a
surgical microscope and each depot has been placed on the mouse
profile mimicking its anatomical position. The organ is made up of two
subcutaneous and several visceral depots. The most representative
visceral depots are shown. Kidneys and testes were dissected together
with the depots. Names of the single depot: A ¼ deep cervical
B ¼ anterior subcutaneous (interscapular, subscapular, axillo-toracic,
superficial cervical) C ¼ visceral mediastinic D ¼ visceral mesenteric
E ¼ visceral retroperitoneal F ¼ visceral perirenal, periovaric, para-
metrial and perivescical G ¼ posterior subcutaneous (dorso-lumbar,
inguinal and gluteal). White areas made up of white adipose tissue and
brown areas composed of brown adipose tissue are indicated by the
scheme.
par excellence has an elongated conical shape and lies in
paravertebral position, i.e. on the border between the
spine and the posterior abdominal wall, and is generally
white. The perirenal depot is divided from the retro-
peritoneal by a peritoneal fold and can be dissected
separately.
The omental depot is small in rodents and the
mesenteric depot is outlined by the two peritoneal
leaflets holding the intestine against the posterior
abdominal wall.
The perigonadal depot is well circumscribed, envel-
oped and bound to the epididymis by the peritoneal
leaflets. In females, it surrounds ovaries, uterus and
bladder.
All the above-described depots of the adipose organ
are mixed: composed by WAT and BAT. The relative
amount of the two tissues is responsible for the colour,
i.e. when WAT is prevalent the colour is white, when
BAT is prevalent the colour is brown. The colour (and
therefore the cell composition) of each single depot, or
part of it, depends on several factors: age, strain, species,
environmental and nutritional conditions. Thus, in adult
mice and rats maintained in standard conditions, the
BAT component is macroscopically visible in the
anterior subcutaneous depot and in the mediastinic
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S. Cinti / Prostaglandins, Leukotrienes and Essential Fatty Acids 73 (2005) 9–15
and perirenal sites. However, light microscopic exam-
ination of WAT depots evidences the consistent
presence of BAT, especially in the inguinal region of
the posterior subcutaneous depot, in the periovarian
region of the gonadal depot and in the retroperitoneal
region. Importantly, the amount of brown adipocytes
found in the different depots is genetically determined,
since in similar environmental and feeding conditions it
is substantially species-related [3].
Subject age is also important under this respect. In a
systematic study of the anterior subcutaneous depot in
animals of different ages, we demonstrated that in
ageing rats the BAT component undergoes a progressive
substitution by WAT, albeit retaining a considerable
size in older (2 year) animals [4,5]. In general, the
number of brown adipocytes tends to decrease with age
in all depots.
Only microscopically detectable WAT depots are
found in bone marrow, parotid, parathyroid and
pancreas.
2. Vessels and nerves
Each depot is endowed with a well-organised vascular
and nerve network. Whereas the former aspect is well
explored and exhaustively described, studies addressing
the specific innervation of each depot are few and
scarcely informative.
The most widely investigated depot is the anterior
subcutaneous, which exhibits a dense vascularity and
innervation [6]. Two anterior vessels reach the wings of
the depot bilaterally from the axillary artery, accom-
panied by veins and nerves of different size. The thinner
nerves are less myelinated and contain a larger number
of noradrenergic fibres. The thicker and more myeli-
nated nerves also contain fibres immunoreactive for the
neuropeptide Y, that co-localise with noradrenaline in
noradrenergic fibres, and for neuropeptides, such as
substance P and calcitonin gene-related peptide, that are
usually found into sensory nerves. A peripheral cyto-
trophic and vasculotrophic action of these sensitive
fibres has been described [7]. Five nerves, prevalently
made up of noradrenergic fibres, run bilaterally and
symmetrically from the intercostal spaces into the body
of the deposit. A large solitary vein (Sulzer’s vein)
ensures a conspicuous outflow from the depth of the
interscapular depot and reaches the azygos vein.
Immunohistochemical and ultrastructural studies
from our and other laboratories have shown that the
fibres innervate the vessels up to the most peripheral
capillaries and also branch directly into the parenchyma,
giving rise to real, consistently noradrenergic, neuro-
adipocytic junctions [8]. The WAT component of the
depot is similarly, but not as densely vascularised and
innervated as the BAT one.
11
A recent immunohistochemical investigation by our
group has shown that the mediastinic BAT depot is
provided not only with sympathetic noradrenergic
nerves but also with cholinergic parenchymal nerves,
likely belonging to the parasympathetic system [9].
3. Adipose organ plasticity and medical implications
Different types of stimuli, such as nutritional stimuli
or modifications in environmental temperature, are able
to determine dramatic anatomical changes in the
adipose organ, that thus is endowed with striking plastic
properties.
4. Fasting
Fasting or food restriction produces organ modifica-
tions that vary based on their duration [8]. During
prolonged fasting, changes involve the sole white
component via a focal tissue response. Indeed, areas
can be observed where some adipocytes have shrunk
and are surrounded by still apparently unaffected
adipocytes. At extreme degrees of delipidation, white
adipocytes are elongated or star-shaped and are still
easily distinguishable from the other parenchymal cells,
essentially for the distinctive presence, on the surface of
these slimming cells, of invaginations (described by
some researchers as surface evaginations or villus-like
structures) characterised by abundant pinocytotic vesi-
cles. These invaginations arise early in the cell slimming
process, because nearly unaffected cells (exhibiting
prevalently unilocular lipid accumulations) show ‘‘tails’’
of slimmed-down cytoplasm rich in invaginations. The
tannic acid technique devised by Blanchette-Mackie and
Scow [10] has made it possible to follow at the electron
microscopic level the process of delipidation by allowing
electrondense myelinic figures (corresponding to migrat-
ing free fatty acids) to be detected in the adipose tissue.
These figures are found on the surface of lipid vacuoles;
in the hyaloplasm; in direct contact with mitochondria;
and in close relation to the cytoplasmic invaginations
rich in pinocytotic vesicles that are characteristic of this
slimmed-down state; they are also detectable free in the
interstitial space, in the cytoplasm of endothelial cells,
and in vessel lumina. In practice, they are found at all
the sites of fatty acid migration during lipolysis.
Although they are not exclusively observed during the
lipolytic process, they are significantly more numerous
during adipocyte delipidisation.
In conditions of prolonged fasting, brown adipocytes
with a complete lipid endowment can be observed side
by side with slimmed-down adipocytes, easily recognised
for the features described above [8]. We feel that this is
the most convincing evidence that, despite the organ’s
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12 S. Cinti / Prostaglandins, Leukotrienes and Essential Fatty Acids 73 (2005) 9–15
plasticity, a small component of either type of cells is
consistently found in the organ. Indeed, in extreme
fasting conditions a part of the energy contained in the
organ’s triglycerides is stored for the critical thermo-
genic requirements, obviously into the brown adipo-
cytes.
The fate of totally slimmed-down cells is unclear.
According to some researchers they may undergo
apoptosis. Still unpublished data from our laboratory
seem to indicate that this is not the case: after protracted
fasting, the retroperitoneal depot of three adult mice
was completely delipidised, but careful electron micro-
scopic examination (highly sensitive to signs of apopto-
sis) of about 500 delipidised cells/mouse showed no sign
of apoptosis in any slimmed-down cell.
5. Warm exposure
Warm exposure entails a reduction of the sympathetic
drive to BAT, resulting in the reduction of heat
production by brown adipocytes. Morphologically, this
corresponds to a transformation of brown fat cells into
cells similar to white adipocytes. During this transfor-
mation, we have observed a reduced gene expression of
UCP1 and increased gene expression of leptin [11]. This
suggests that such morphological transformation is
accompanied by a new functional situation, with
adipocytes losing their thermogenic ability and acquir-
ing the properties of white fat cells, including production
of such an important hormone as leptin. This is in line
with the observation that classic multilocular brown
adipocytes subjected to adrenergic stimulation express
UCP1 but not leptin [11,12], whereas cold exposure and
adrenergic drugs reduce leptinaemia [13] and induce the
transformation of white into brown adipocytes [14] (see
below: transdifferentiation).
It is interesting to note that the energy required for
basic metabolic activities is considerably reduced in
animals maintained in thermoneutral conditions. In-
deed, in an adult mouse subjected to prolonged fasting
(48 h), the whole WAT shrinks almost to disappearance
at around 10 1C, whereas it is nearly unchanged if the
animal is kept at 28 1C [8].
6. Cold exposure
This condition involves the immediate activation of
the sympathetic nervous system, with the consequent,
immediate functional activation of brown adipocytes via
the release of noradrenaline. In a few hours, over-
stimulated brown adipocytes adapt their thermogenic
ability to increased requirements by synthesising new
mitochondria and higher amount of UCP1; in the
course of a few days, new cells develop, giving rise to a
new tissue organisation characterised by an increased
number of vessels and nerves [15,16]. In the framework
of the new concept of adipose organ, it is easy to
understand that the arising of new brown cells does not
only involve some areas long considered as pure BAT,
but the whole organ. It follows that its macroscopic
appearance as well as the microscopic features of the
different depots turn towards a more brown appearance.
It is important to note that this process does not
necessarily entail the arising of brown adipocytes in
white depots—since, as mentioned above, brown cells
are normally present in the various depots (whose
amount depending on age, strain, etc., see above)—but
rather an increase in their number [17,18].
Transmission electron microscopic (TEM) studies
(which are very sensitive in detecting the different
stages of cell development) have shown that the adipose
organ of adult mice and rats kept in thermoneutral
conditions and fed on a normal diet contains neither
totally undifferentiated nor scarcely differentiated
cells. By contrast, in cold-exposed animals undifferen-
tiated pericytes and brown preadipocytes (i.e. cells
with minimal differentiation features that allow their
morphological identification as cells committed to
differentiate into brown adipocytes) arise both in
BAT and in WAT. Preadipocytes may be found in
pericytic and/or perivascular position, suggesting that
cold exposure induces the tissue conditions that lead to
the arising of brown preadipocytes in all WAT and
BAT areas of the organ. Therefore, irrespectively
of the existence of distinct white and brown cell
precursors, their arising in different depots of adult
animals clearly depends on microenvironmental tissue
conditions. Cold exposure induces an increased activity
of noradrenergic parenchymal fibres and simultaneous
branching of nerve fibres both in WAT [19] and in BAT
[6]. In turn, this induces an overall increase in
noradrenaline tissue concentration followed by the
development of brown cells in the various depots of
the organ.
In contrast, development of white preadipocytes in
the various areas appears to be triggered by different
stimuli (see under: positive energy balance) directed at
increasing the energy storage capacity.
7. Transdifferentiation
Transdifferentiation is a biological phenomenon by
which a differentiated cell turns phenotypically and
functionally into a differentiated cell of another type
without undergoing dedifferentiation [20]. We believe
that brown and white adipocytes can transdifferentiate
into one another in the adipose organ. We here report
some data providing evidence for physiological trans-
differentiation of white into brown fat cells.
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It has been known since Ashwell’s pioneering work
(1984) that after cold exposure brown adipocytes
expressing UCP1 arise in WAT areas of the adipose
organ [17]. Our group has demonstrated that the
precursors developing in the periovarian depot have
ultrastructural and genetic expression features typical of
brown precursors [18]. A recent study of WAT tissue in
different mice strains exposed to cold for varying
periods of time has suggested that a significant number
of multilocular adipocytes arise in a matter of days in
visceral as well as subcutaneous WAT depots [21].
Analysis of the gene expression of these tissues after
acute cold exposure (2 days) showed a significant
increase in UCP1mRNA from baseline, suggesting that
the new multilocular cells express UCP1mRNA1. Inter-
estingly, these cells do not express UCP1 protein (as
demonstrated by immunohistochemical and Western
blotting techniques), indicating that after 2 days of cold
exposure a significant number of well-differentiated
multilocular adipocytes express UCP1 gene but not the
protein. Considering that in the course of foetal organ
development, brown adipocyte precursors express
UCP1 quite early with respect to the overall stage of
cell differentiation (and, signally, of the amount of
cytoplasmic lipids stored), it seems reasonable to
hypothesise that also the arising of the multilocular
adipocytes in WAT depots after cold exposure is a
phenomenon quite distinct from mere preadipocyte
differentiation. In the course of the same experiments,
we also noted that the number of multilocular adipo-
cytes increased further after a longer exposure to cold
(10 days), and that, unlike those arising after acute
exposure, these cells also expressed UCP1 protein. It
thus appears that the continuous adrenergic stimulus
prompted the multilocular cells already expressing
UCP1mRNA to express also the protein and thus to
acquire the functional thermogenic properties of brown
adipocytes. In transgenic animals not expressing X 3
adrenergic receptor, the same experiments evidenced a
1
Whereas in the opinion of several researchers the presence of UCP1
is a brown adipocyte hallmark, in our view it is merely a cellular
feature that subserves the main function of BAT; however, brown
adipocytes do not consistently express it, and in some conditions in
which they do not do so, they assume the appearance of white fat cells.
For instance, in an animal maintained above its thermoneutral
temperature (see above), the BAT-activating adrenergic stimulus is
off; brown adipocytes thus undergo a morphological transformation:
they become unilocular and lose the typical mitochondrial features,
thereby becoming similar to white adipocytes. This morphological
transformation is accompanied by inhibition of UCP1 gene and by ob
gene (leptin) activation [11]. The same process takes place in mice
lacking all X -adrenergic receptor subtypes, demonstrating that this
phenomenon is mediated by the X -adrenergic stimulus [22]. Although
these data seem to be consistent with the concept that a brown
adipocyte lacking UCP1 is a more or less typical white fat cell, this is
denied by the fact that multilocular brown adipocytes quite different
from white fat cells are found in the adipose organ of UCP1 knock-out
mice (our unpublished data).
13
nearly complete inhibition of this phenomenon, suggest-
ing an important role for this receptor in its mediation.
In addition, given that brown adipocyte precursors do
not appear to express X 3AR receptor, the hypothesis
may be advanced that brown preadipocytes do not play
a critical role in determining the presence of the new
population of multilocular or brown adipocytes.
Immunohistochemical studies demonstrate that
UCP1 protein is expressed by varying proportions of
multilocular cells in the different depots. In this context,
the most interesting observation, made in our laboratory
in collaboration with the group of J.P. Giacobino
(Geneve), was that in the subcutaneous depot of cold-
exposed (6 1C for 10 days) SV129 mice, about 35% of
adipocytes were multilocular cells, 85% of which
expressed UCP1. Also in this condition, then, the
absence of X 3AR receptor significantly reduced the
number of multilocular adipocytes (our unpublished
data). These findings seem to substantiate the hypoth-
esis, advanced in a previous work by our group, that
chronic (7 days) stimulation with the X 3AR agonist CL
316,243 could induce direct transformation of white into
multilocular adipocytes, around 8% of which would
express UCP1 (14). Prolonged stimulation (7 more days)
with the same drug induced a considerable increase in
the proportion of UCP1-expressing cells (33%) without
a concurrent increase in the number of multilocular cells
(our unpublished data). This suggests that only a
subpopulation white adipocytes would be committed
to this transformation, confirming that white-to-brown
transdifferentiation passes through successive steps
beginning with the transformation of the lipid vacuole
from unilocular into multilocular, followed by the
expression of UCP1.
X 3AR receptor agonists administered to rats with
genetic or diet-induced obesity are known to activate
BAT, to induce the transdifferentiation of white into
brown adipocytes [14], and to cure both their obesity
and the consequent diabetes. Unfortunately, although
some recent data indicate the presence of the receptor
also in the human adipose organ [23], a drug inducing
the same effects in man is still not available.
In a recent paper, Toseland [24] demonstrates that
other drugs could be involved in the plastic modulation
of the adipose organ, particularly the agonists of
PPARg—a nuclear receptor acting as a transcription
factor and implicated in the process of development of
fat cells [25]—which induces the transformation of the
adipose organ of treated dogs and rats into BAT.
8. Positive energy balance: overweight and obesity
When the energy balance is positive, the adipose
organ prevalently undergoes an increment in its white
component. White adipocytes become hypertrophic and
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14 S. Cinti / Prostaglandins, Leukotrienes and Essential Fatty Acids 73 (2005) 9–15
subsequently (likely due to a close causal relationship),
hyperplastic. In fact, adipocytes have been suggested to
be unable to expand beyond a given maximum volume,
or ‘‘critical size’’, which is genetically determined and
specific for each depot [26]. Adipocytes that have
reached the critical size trigger an increase in cell
number [27–29]. In a recent review, Hausman et al.
[30], after considering the evidence for this theory,
conclude that not only paracrine factors, but also
circulating factors as well as neural influences may play
a large role in regulating adipose tissue development and
growth. They suggest that in the development of obesity
enlarged fat cells produce and release proliferative
paracrine factors as internal controllers of preadipocyte
proliferation, and that their proliferative response is
modulated by neural inputs to fat tissue and/or serum
factors. In any case, paracrine factors appear to play a
pivotal role. Adipose tissue expresses numerous factors
that could be implicated in the modulation of adipogen-
esis: IGF I, TGF-b, TNF-a, macrophage colony-
stimulating factor (MCSF), angiotensin II, autotaxin-
lysophosphatidic acid (ATX-LPA), leptin, resistin,
etc. [31].
Of note, in genetically obese ob/ob mice (lacking
leptin) and in other types of genetic and diet-induced
obesity, the fat mass is hypertrophic and hyperplastic,
while in genetically obese db/db mice (lacking leptin
receptor) the fat mass is increased only by a hyper-
trophic mechanism ([32], and our unpublished observa-
tions). Therefore, the presence of leptin receptor seems
to be essential to induce hyperplasia. In the subcuta-
neous adipose tissue of a massively obese patient lacking
leptin receptor [33], we recently observed that mean
adipocyte volume was about half the one usually seen in
the same depot of patients with similar BMI due to
‘‘primitive obesity’’.
A recent study showed that obesity induced by a high-
fat diet in mice is hypertrophic, while the one induced by
hypothalamic lesion due to administration of mono-
sodium glutamate is hyperplastic [34].
A positive energy balance also affects the organ’s
brown component. The brown adipocytes of obese
animals are generally similar to white cells. Prevalently
unilocular cells are observed at the sites where brown
adipocytes are normally found, but these cells often
exhibit typical mitochondria distinct from the ‘‘normal’’
organelles of white fat cells. They usually—though
modestly—express UCP1 as well as the typical protein
of white adipocytes: leptin [12]. Interestingly, the large
amount of TNFa found in obese mice induces an
increased rate of apoptosis of brown fat cells [35].
Transgenic animals lacking X 1, 2 and 3 receptors
become massively obese eating the same amount of food
as lean ones [22]. In such animals, brown adipocytes are
identical to those found in the genetically obese (ob/ob)
and db/db mice described above: they are unilocular and
show leptin expression and modest UCP1 activity in
mitochondria with the typical morphology. This is in
line with the fact that obesity due to lack of leptin (ob/
ob) or its receptor (db/db) is accompanied by reduced
BAT adrenergic stimulation owing to the absence of the
stimulus normally exerted by the hormone on hypotha-
lamic sympathetic centres.
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