Archives of Physiology and Biochemistry

The Journal of Metabolic Diseases

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Spirulina reduces diet-induced obesity through

downregulation of lipogenic genes expression in
Psammomys obesus

Hamza Saidi, Abdenour Bounihi, Asma Bouazza, Aziz Hichami, El Hadj

Ahmed Koceir & Naim Akhtar Khan

To cite this article: Hamza Saidi, Abdenour Bounihi, Asma Bouazza, Aziz Hichami, El Hadj Ahmed
Koceir & Naim Akhtar Khan (2020): Spirulina reduces diet-induced obesity through downregulation
of lipogenic genes expression in Psammomys�obesus, Archives of Physiology and Biochemistry,
DOI: 10.1080/13813455.2020.1743724

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Published online: 24 Mar 2020.

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ARCHIVES OF PHYSIOLOGY AND BIOCHEMISTRY
https://doi.org/10.1080/13813455.2020.1743724

ORIGINAL ARTICLE

Spirulina reduces diet-induced obesity through downregulation

of lipogenic genes expression in Psammomys obesus
Hamza Saidia , Abdenour Bounihia, Asma Bouazzaa, Aziz Hichamib, El Hadj Ahmed Koceira and
Naim Akhtar Khanb
a
Bioenergetics and Intermediary Metabolism team, Laboratory of Biology and Organism Physiology, University of Sciences and Technology
Houari Boumediene, Algiers, Algeria; bINSERM U1231, University of Burgundy Franche-Comt
e, Dijon, France

ABSTRACT ARTICLE HISTORY

The present study evaluates the protective effect of spirulina against diet-induced obesity and meta- Received 20 April 2019
bolic disorders in Psammomys obesus, an animal model of metabolic syndrome. Psammomys obesus Accepted 12 March 2020
lives on a low-energy diet, in order to remain healthy. However, under a standard laboratory chow Published online 2 2 2
diet (SLCD), this animal exhibits insulin resistance, which occurs as a result of obesity. Psammomys obe-
KEYWORDS
sus was maintained on SLCD, in order to evaluate the effect of spirulina on obesity development with Spirulina; Psammomys
a particular focus on glucose and lipid metabolism, as well as the mRNA expression of some pro- obesus; lipogenesis; obesity;
inflammatory cytokines. After 12 weeks of treatment with spirulina, there was a significant reduction in insulin-resistance
body weight gain, plasma glucose, insulin and triglyceride levels. There was also a significant reduction
in the mRNA expression of genes involved in lipogenesis and inflammation. Spirulina improved insulin
sensitivity, glucose and lipid metabolism. These findings highlight the positive effect of spirulina on
weight maintenance.

Introduction consumed as food for centuries by Mexicans during Aztec

The obesity epidemic was once limited to high-income coun-
tries, though most middle-income and low-income countries
soon joined the global surge (Swinburn et al. 2011).
According to the Global Burden of Disease Collaborators
(2017), an estimated 107.7 million children and 603.7 million
adults were obese. Obesity is a major factor contributing to
the development of type 2 diabetes, dyslipidaemia, fatty liver
disease and cardiovascular diseases (Reaven 1988).
In this experiment, tests were conducted on Psammomys
obesus, a unique animal model of nutritionally-induced meta-
bolic syndrome (Levy et al. 2006). In its native habitat, P. obe-
sus feeds on low caloric vegetation and remains healthy.
However, when this animal feeds on a standard laboratory
chow diet (SLCD), P. obesus becomes obese and develops
type 2 diabetes (Koceir et al. 2003). This is mainly due to
insulin resistance (the failure of insulin to promote peripheral
glucose uptake). However, in this state of insulin resistance,
the insulin continues to activate lipogenesis in the liver
(Lewandowski et al. 1998, Shafrir et al. 2006). The insulin also
stimulates the production of lipoprotein lipase, accentuating
the uptake of preformed lipids by adipose tissue (Chajek-
Shaul et al. 1988).
In this study, P. obesus was subjected to treatment by
spirulina, a multicellular, filamentous cyanobacteria that
grows in high-salt alkaline water reservoirs in subtropical and
tropical areas (Deng and Chow 2010). Spirulina has been
civilisation and also in Central Africa (Dillon et al. 1995). It
has received much attention as a functional food due to its
widely-demonstrated antioxidant effects (Bermejo-Besco
s
et al. 2008, Gargouri et al. 2016) and anti-inflammatory
effects in several studies (Chen et al. 2012, Abdel-Daim et al.
2015). These therapeutic properties enable the use of spiru-
lina in the treatment of many chronic ailments such as arth-
ritis (Remirez et al. 2002) and cardiovascular disease (Khan
et al. 2006, Riss et al. 2007). Moreover, several studies have
shown that spirulina supplementation can reduce lipid accu-
mulation in the liver (Kato et al. 1984), and regulate serum
cholesterol (Devi and Venkataraman 1983, Kato et al. 1984)
and triglycerides (Rodrıguez-Hern
andez et al. 2001, Serban
et al. 2016).
To our knowledge, little is known about the molecular
mechanisms through which spirulina affects obesity develop-
ment, lipid metabolism, and inflammation. The present study
was designed to investigate the effects of spirulina supple-
mentation on SLCD-induced obesity, insulin resistance,
hyperlipidaemia and low-grade inflammation in P. obesus
with a particular focus on the mRNA expression of genes
involved in glucose and lipid metabolism, as well as some
pro-inflammatory cytokines. The chosen animal model is par-
ticularly suitable to address these issues given the animal’s
potential to develop nutrition-dependent obesity, type-2 dia-
betes, and hepatic steatosis. Psammomys obesus also shares
most of the metabolic parameters of human metabolic

CONTACT Hamza Saidi saidi.hamza.dz@gmail.com Bioenergetics and Intermediary Metabolism team, Laboratory of Biology and Organism Physiology,
University of Sciences and Technology Houari Boumediene, Algiers, Algeria
ß 2020 Informa UK Limited, trading as Taylor & Francis Group
2 H. SAIDI ET AL.

syndrome. In this study, the effects of spirulina on P. obesus
were probed via the transcription level of several genes
involved in lipogenesis, glucose homeostasis and the inflam-
matory process.
Materials and methods
Animal and diet
Animal experimentation conformed to the Guide for the
Care and Use of Laboratory Animals issued by the
National Institutes of Health, USA [DHEW Publication No.
(NIH) 85–23, Revised 1996, Office of Science and Health
Reports, DRR/NIH, Bethesda, MD 20205, USA]. Animal
experimentation was also approved by the Algerian
Institutional Animal Care Committee, which belongs to the
National Administration of Algerian Higher Education and
Scientific Research (Ethical approval number: 981–1 law of
22, August 1998).
Adult male P. obesus were trapped in the area of Beni-
Abbes (30
70 N; 2
100 W) in the Algerian West Sahara and
transported to Algiers. The animals were kept under condi-
tions as similar as possible to those of their natural environ-
ment, at 22–26
C, 60 ± 10% relative humidity and 12/12 h
light/dark cycle. In order to satisfy their nutritional require-
ments, P. obesus were always fed on a natural diet (ND)
composed mainly of halophile plants of the Chenopodiaceae
family (especially Salsola foetida) with the following compos-
ition: 80.8% water, 8.4% carbohydrates, 6.9% ash, 3.5% pro-
teins, 0.4% lipids and a caloric value of 0.4 kcal/g (Gouaref
et al. 2017). After 15 days of acclimatisation, P. obesus were
randomly divided into three groups of ten animals each. The
first group was used as a control and continued to be fed
with ND. The second group was fed with SLCD obtained
from ONAB, Algiers with the following composition: 47% car-
bohydrates, 25% proteins, 7.5% fat and a caloric value of
3.85 kcal/g (Gouaref et al. 2017). The third group (SLCD þ Spi)
was fed with SLCD and administered spirulina (Spirulina pla-
tensis), which was cultured in our laboratory as previously
described by Amara-Leffed et al. (2018). Spirulina was admin-
istered at a dose of 2 g/kg, once a day, orally. This dose was
chosen based on previous work by El-Desouki et al. (2015).
Control and experimental groups had free access to food
and water. Food consumption was measured daily and pre-
sented in Figure 1(C), and body weight was determined once
a week.
After 12 weeks of experimentation, the animals were
euthanized with urethane (1 g/kg). Blood was collected in
heparinised tubes and centrifuged (5000g, 10 min), and
plasma was stored at 80
C. Liver tissues were quickly
removed, weighted and immediately conserved in liquid
nitrogen or fixed in a suitable medium.

Figure 1. The effect of spirulina on body weight, food and energy intake, and the relative weight of the liver in P. obesus. The animals were maintained on ND,
SLCD or SLCD þ Spi for 12 weeks. The animals were weighed weekly (A). (B) Represents weight gain at the end of experimentation. Food intake and food energy
(C) were determined daily till the 12th week of the study. (D) Represent the relative weight of the liver. Data is presented as means ± SEM (n ¼ 10). ,  and 
represent p < .05, p < .01 and p < .001 respectively as compared to ND-fed animals. # and ## represent p < .05 and p < .01 respectively as compared to SLCD-fed
animals. p values were obtained using Student’s t-test.
 ARCHIVES OF PHYSIOLOGY AND BIOCHEMISTRY 3

Determination of biochemical parameters Histological examination

Plasma triglyceride, total cholesterol and glucose concentra-
tions were measured using the Roche Hitachi 902 auto ana-
lyser (Roche Molecular Systems, Branchburg, NJ). Plasma
insulin concentration was determined by an enzyme-linked
immunosorbent assay (DRG Instruments GmbH, Marburg,
Germany). To determine plasma fatty acid composition, the
lipid extract was analysed as previously described (Atek-
Mebarki et al. 2015) with minor modification. The desaturase
activity of Stearoyl-CoA desaturase (SCD-act) was calculated
as the ratio between product and precursor according to the
following equation: SCD-act ¼ [16:1 (n-7)/16:0] or [18:1 (n-9)/
18:0] (Warensjo € et al. 2008). Stearoyl-CoA desaturase 1
(SCD1) catalyses the desaturation of palmitic acid (16:0) to
palmitoleic acid (16:1 n-9) and stearic acid (18:0) to oleic acid
(18:1 n-9) (Kawano and Cohen 2013).
Hepatic lipids were extracted using the Folch method
(Folch et al. 1957) and hepatic triglyceride and cholesterol
levels were measured using a colorimetric kit (Sodioba, USA).
Small pieces of liver were fixed in Bouin’s solution. After
embedding these liver pieces in paraffin, they were sectioned
at 3 mm and stained with masson trichrome. Sections were
examined using light microscopy (Leica, USA). The area and
number of lipid droplets were measured using the particle
sizing function provided by the ImageJ software version 1.52
(Rasband, ImageJ, National Institutes of Health, Bethesda,
MD, USA). To account for discrepancies in sinusoid size
between animals, the number and the surface area of the
lipid droplets are expressed for a standardised surface of liver
section of 10,000 mm2.
Statistical analyses
Results are expressed as mean ± SEM. The statistical differ-
ence between groups was evaluated by Student’s t-test
using GraphPad Prism version 6.01 (GraphPad Software,
USA). All differences with a p values ˂ .05 were considered
statistically significant.

Results
Determination of mRNA expression
The effect of spirulina on energy intake, body weight,
Real-time PCR (RT-PCR) was used to measure mRNA expres-
relative weight of liver and liver lipids
sion of sterol responsive element binding-proteins-1c (SREBP-
1c), carbohydrate-responsive element-binding protein Figure 1(A) shows that Psammomys obesus, which were
(ChREBP), acetyl-CoA carboxylase-1 (ACC1), fatty acid syn- maintained on SLCD, gained weight as a function of time.
thase (FAS), SCD1, interleukin-1b (IL-1b) and tumour necrosis The SLCD promoted a significant increase in body weight
factor-a (TNF-a) in the liver. Total RNA was isolated from liver over the twelve-week period (Figure 1(A)). Differences in
samples using Trizol reagent (Invitrogen, USA), as per the body weight gain among groups became noticeable at the
manufacturer’s instructions. cDNA was generated from the first week of the diet and became significant after the
isolated RNA using the iScript cDNA synthesis kit (Bio-Rad, second week between ND-fed animals and SLCD-fed animals.
USA). Aliquots of cDNA were subjected to RT-PCR using the After four weeks of treatment, significant differences
Step-One Plus Detection System and SYBR Green PCR Master occurred between the SLCD-fed animals and the SLCD þ Spi-
Mix (both from Applied Biosystems, USA). Primers for genes fed animals. At the end of the experiment, the weight gain
of interest were taken from the literature (Jo €rns et al. 2006, was higher in SLCD-fed animals than in the SLCD þ Spi-fed
animals (Figure 1(B)).
Hamlat et al. 2010) (Table 1). The 18S gene was used as an
The mean body weight increased by 17.65 ± 0.6 g in the
internal control. The data are presented as the expression of
ND-fed animals, 40.29 ± 1.5 g in the SLCD-fed animals and
the genes of interest relative to the internal control gene as
25.69 ± 0.8 g in the SLCD þ Spi-fed animals (Figure 1(B)). The
determined by the 2(-DDCT) method (Livak and
average food intake of the SLCD-fed animals was lower than
Schmittgen 2001).
that of the ND-fed animals (12.02 g/d and 44.9 g/day,
respectively), but did not differ between the SLCD and
Table 1. Primers used for mRNA expression.
SLCD þ Spi (10.9 g/d) groups (Figure 1(C)). Conversely, due to
Genes Primers sequences Ref.
the higher caloric value of SLCD (3.85 kcal/g) compared to
18S F: 50 -TGAGGCCATGATTAAGAGGG-30 Hamlat et al. (2010)
R: 50 -AGTCGGCATCGTTTATGGTC-30 ND (0.4 kcal/g), both SLCD and SLCD þ Spi-fed animals had
SREBP-1c F: 50 -CGCTACCGTTCCTCTATCAA-30 Hamlat et al. (2010) higher energy intake (46.28 kcal/day and 41.97 kcal/day,
R: 50 -TTCGCAGGGTCAGGTTCTCC-30 respectively) than ND-fed (17.96 kcal/day) animals
ChREBP F: 50 -CGGGACATGTTTGATGACTATGTC-30 Hamlat et al. (2010)
R: 50 -AATAAAGGTCGGATGAGGATGCT-30 (Figure 1(C)).
ACC1 F: 50 -ACATCCCTACGCTAAACA-30 Hamlat et al. (2010) For SLCD-fed animals, this resulted in the accumulation of
R: 50 -AGAACATCGCTGACACTA-30 energy in the form of triglycerides stored in adipose tissue
FAS F: 50 -CCCAATGCCTTGTTCCC-30 Hamlat et al. (2010)
R: 50 -CTGTGGTCCCACTTGATGAG-30 as well as ectopic deposition in tissues, such as the liver.
SCD1 F: 50 -TTCTTCTCTCACGTGGGTTG-30 Hamlat et al. (2010 Thus, there was a significant increase in the relative weight
F: 50 -TTGTAGTACCTCCTCTGGAA-30 of the liver in SLCD-fed animals compared with ND-fed ani-
TNFa F: 50 -GACATCGAGCTGGCGGAGGA-30 J€
orns et al. (2006)
R: 50 -CCGCCACGAGCAGGAAAGAG-30 mals. However, 2 g/kg/day spirulina supplementation resulted
IL1b F: 50 -AGCTTCCGGCAGGTGGTATCG-30 J€
orns et al. (2006) in a significant reduction in the relative weight of the liver in
R: 50 -TGGAAGGTCCACGGGCAAGA-30 SLCD þ Spi-fed animals in comparison to SLCD-fed animals
4 H. SAIDI ET AL.
Figure 2. The effect of spirulina on liver lipid parameters in P. obesus. The animals were maintained on ND, SLCD or SLCD þ Spi for 12 weeks. After euthanasia, liver
samples were taken to assess fat mass (A), triglyceride (B) and cholesterol (C) levels as described in Materials and Methods. Data is presented as means ± SEM
(n ¼ 10).  and  represent p < .05 and p < .001 respectively as compared to ND-fed animals. # and ## represent p < .05 and p < .001 respectively as compared
to SLCD-fed animals. p values were obtained using Student’s t-test.
(Figure 1(D)). SLCD þ Spi-fed animals also had lower values
for liver total lipid (Figure 2(A)), triglyceride (Figure 2(B)) and
cholesterol (Figure 2(C)) levels in comparison to SLCD-
fed animals.
Histological findings
Histologic examination of Masson trichrome stained liver sec-
tions demonstrated that in contrast to normal histological
structure observed in ND-fed animals (Figure 3(A)), pro-
nounced fatty degeneration of hepatocytes (Figure 3(B)),
with a large number of lipid droplets (Figure 3(D)), was seen
in the liver of SLCD-fed animals. Both the number and the
size of these droplets were markedly reduced by spirulina
(Figure 3(D,E)).
Spirulina reduced plasma glucose, insulin, triglyceride
and cholesterol levels
As illustrated in Table 2, SLCD-fed animals had a significant
increase in plasma glucose and insulin concentrations in
comparison to ND-fed animals. The treatment of SLCD-fed
animals with spirulina triggered a significant decrease in
plasma glucose and insulin. Accordingly, the homeostatic
model assessment of insulin resistance (HOMA-IR) value was
higher in SLCD-fed animals compared to ND-fed animals.
Animals treated with spirulina showed a significant decrease
in the HOMA-IR value. SLCD-fed animals also exhibited
higher plasma triglyceride and cholesterol concentrations in
comparison to ND-fed animals. Spirulina supplementation
reduced plasma triglyceride and cholesterol concentrations.
Effect of spirulina on plasma fatty acid composition and
desaturase activity
Table 3 shows the fatty acid composition of animals under
ND, SLCD or SLCD þ Spi. SLCD-fed animals had higher
amounts of palmitic acid (C16:0), stearic acid (C18:0), palmito-
leic acid (C16:1 n-7), oleic acid (C18:1 n-9) and linoleic acid
(C18:2 n-6) in comparison to ND-fed animals. Spirulina sup-
plementation decreased the contents of these five fatty acids
(C16:0, C18:0, C16:1, C18:1 and C18:2 n-6) in SLCD-fed
animals. Estimated desaturase activity [SCD-act (18:1 n-9/
18:0)] was significantly increased in SLCD-fed animals in com-
parison to ND-fed animals; this was attenuated by the
SLCD þ Spi diet. No differences were observed among the
three animal groups for SCD-act (16:1 n-7/16:0).
Spirulina downregulates lipogenesis-related genes in
SLCD-fed Psammomys obesus
To examine whether the genetic expression of lipogenic
enzymes is affected by spirulina, we measured the mRNA
expression of SCD1, FAS and ACC1 in the liver. We also
measured the transcript levels of SREBP-1c and ChREBP, two
transcription factors involved in the regulation of the expres-
sion of lipogenic genes.
We observed that the mRNA expression of SREBP-1c
(Figure 4(A)) and ChREBP (Figure 4(B)) were significantly
increased in SLCD-fed animals as compared to ND-fed ani-
mals. SLCD-fed animals treated with spirulina showed a sig-
nificant decrease in the mRNA expression of SREBP-1c
and ChREBP.
Also, SLCD led to a significant increase in the mRNA
expression of ACC1 (Figure 4(C)), FAS (Figure 4(D)) and SCD1
(Figure 4(E)) compared to ND. Spirulina treatment induced a
significant decrease in the mRNA expression of ACC1, FAS
and SCD1 in P. obesus under SLCD.
Spirulina modulates the mRNA expression of genes
encoding pro-inflammatory cytokines
To determine the effect of spirulina on pro-inflammatory
cytokines, RT-PCR analysis was conducted on TNF-a and IL-
1b genes in the liver. The mRNA expression of TNF-a (Figure
4(F)) and IL-1b (Figure 4(G)) genes was significantly higher in
SLCD-fed animals in comparison to ND-fed animals. However,
spirulina supplementation significantly dampened the SLCD-
induced expression of theses pro-inflammatory cytokines
(Figure 4(F,G)).
 ARCHIVES OF PHYSIOLOGY AND BIOCHEMISTRY 5

Figure 3. The effect of spirulina on lipid accumulation in liver sections of P. obesus fed ND (A), SLCD (B) or SLCD þ Spi (C) stained with Masson trichrome and
observed with light microscopy. (D) Hepatic lipid droplet number distribution. (E) Mean value of lipid droplet size in each group. For D and E, data is presented as
means ± SEM (n ¼ 10).  represents p < .001 as compared to ND-fed animals. ### represents p < .001 as compared to SLCD-fed animals. p values were obtained
using Student’s t-test. CV: Centrolobular Vein; LD: Lipid Droplet.

Table 2. Biochemical parameters of Psammomys obesus under different diets.

Parameters P. obesus-ND P. obesus-SLCD P. obesus-SLCD þ Spi
Glucose (mM) 3.39 ± 0.06 5.93 ± 0.18 3.95 ± 0.09##
Insulin (pM) 246.24 ± 10.08 714.67 ± 37.94 462.55 ± 17.85#
HOMA-IR 5.38 ± 0.25 28.7 ± 1.89 11.85 ± 0.58##
Triglyceride (mM) 0.74 ± 0.04 2.76 ± 0.21 0.91 ± 0.05#
Total cholesterol (mM) 2.2 ± 0.04 3.49 ± 0.07 2.48 ± 0.06##
HDL-cholesterol (mM) 1.21 ± 0.03 1.48 ± 0.03 0.97 ± 0.03##
LDL-cholesterol (mM) 0.65 ± 0.05 1.95 ± 0.12 1.1 ± 0.07#
Data represent means ± SEM (n ¼ 10).  ,
and  represent p < .05, p < .01 and p < .001 respectively as compared to ND-fed animals. # and ## represent
p < .05 and p < .01 respectively as compared to SLCD-fed animals. p values were obtained using Student’s t-test.
6 H. SAIDI ET AL.

Table 3. The fatty acid composition (mmol/l) of plasma and estimated desaturase activity of Psammomys obesus under different diets.
Fatty acids P. obesus-ND (a) P. obesus-SLCD (b) P. obesus-SLCD þ Spi (c) p values
14:0 0.07 ± 0.01 0.27 ± 0.03 0.05 ± 0.01 a:b (˂.05) ; b:c (˂.05)
16:0 0.88 ± 0.03 2.43 ± 0.15 1.16 ± 0.07 a:b (˂.01) ; b:c (˂.05)
18:0 0.36 ± 0.01 0.61 ± 0.02 0.43 ± 0.01 a:b (˂.001) ; b:c (˂.001)
16:1 n-7 0.12 ± 0.01 0.25 ± 0.02 0.1 ± 0.01 a:b (˂.05) ; b:c (˂.05)
18:1 n-9 0.72 ± 0.02 2.84 ± 0.2 0.9 ± 0.03 a:b (˂.01) ; b:c (˂.05)
18:2 n-6 0.82 ± 0.02 3.08 ± 0.13 1.94 ± 0.1 a:b (˂.001) ; b:c (˂.05)
18:3 N-3 0.49 ± 0.02 0.15 ± 0.01 0.37 ± 0.03 a:b (˂.001) ; b:c (˂.05)
SCD-act 16 0.14 ± 0.01 0.15 ± 0.02 0.11 ± 0.01 NS
SCD-act 18 2.13 ± 0.08 5.1 ± 0.45 2.1 ± 0.07 a:b (˂.05) ; b:c (˂.05)
Values are means ± SEM. SCD-act 16 ¼ C16:1 n - 7/C16:0; SCD-act 18 ¼ C18:1 n - 9/C18:0. Different groups were statistically compared
by Student’s t-test in the following manner: (a) vs (b), (b) vs (c).

Figure 4. The hepatic mRNA expression of genes involved in lipid metabolism and inflammation in animals fed ND, SLCD or SLCD þ Spi for 12 weeks. After euthan-
asia, liver samples were taken to assess the mRNA expression of SREBP-1c (A), ChREBP (B), ACC1 (C), FAS (D), SCD1 (E), TNF-a (F) and IL-1b (G) genes as per the
technique explained in “Materials and methods” section. Data is presented as means ± SEM (n ¼ 10).  and  represent p < .05 and p < .01 respectively as com-
pared to ND-fed animals. # and ## represent p < .05 and p < .01 respectively as compared to SLCD-fed animals. p values were obtained using Student’s t-test.

Discussion
The present study was designed to determine the effect of
3-month oral treatment with spirulina on metabolic disorders
related to obesity in P. obesus. This animal model of obesity
and metabolic syndrome is characterised by high hepatic
lipogenic activity and innate insulin resistance (Shafrir
et al. 2006).
In P. obesus, SLCD has been reported to induce hepatic
lipid accumulation, hypertriglyceridaemia, hyperglycaemia
and hyperinsulinemia, thus promoting obesity (Koceir et al.
2003). In this study, spirulina supplementation for 12 weeks
resulted in a 36.23% decrease in body weight gain in
SLCD þ Spi-fed animals in comparison to SLCD-fed animals;
this occurred despite the fact that there was no significant
difference in average energy intake. The anti-obesity effect
of spirulina has also been demonstrated in human clinical tri-
als by Szulinska et al. (2017), who found that spirulina (at a
dose of 2 g/day during 12 weeks) decreased body weight by
4.29%. Zeinalian et al. (2017) also found that spirulina (at a
dose of 1 g/day for 12 weeks) decreased body weight
by 1.79%.
Based on the HOMA-IR values presented in this study,
SLCD-fed Psammomys obesus were more insulin-resistant
than ND-fed animals, suggesting that the high degree of
obesity in these SLCD-fed animals might be caused, in part,
by the significantly higher fasting blood glucose and insulin
levels (Khan et al. 2017). The treatment of P. obesus with
spirulina improved the SLCD-induced alteration of the
HOMA-IR value. These findings highlight the ability of spiru-
lina to reduce insulin resistance and weight gain in P. obesus
under SLCD.
In a state of insulin resistance (as with the SLCD-fed ani-
mals), it has been shown that lipogenesis is increased mainly
due to the relatively preserved sensitivity of the lipogenesis
pathway to insulin (Chen et al. 2004). This increase in hepatic
lipogenesis can result in an increase in adipose tissue and
obesity (Strable and Ntambi 2010) as well as the ectopic
deposition of lipids in body organs, especially in the liver
(Liao et al. 2013).
Accordingly, the SLCD-fed animals in this study depicted
an increase in the relative weight of the liver in comparison
to ND-fed animals. The administration of spirulina decreased
the relative weight of the liver, suggesting a decrease in lipo-
genesis and ectopic deposition. Histological sections also
showed that spirulina markedly reduced hepatic lipid drop-
lets. Furthermore, serum total cholesterol and triglycerides
levels were significantly reduced by spirulina. These results
suggest that spirulina attenuates lipid accumulation in the
liver and suppresses hyperlipidaemia in P. obesus
under SLCD.
To confirm the effects of spirulina on lipogenesis, the
expression of lipogenesis-related genes in the liver of SLCD-
fed P. obesus was probed. Various nuclear transcription fac-
tors regulate the rate of lipogenesis such as ChREBP and
SREBP-1c (Zhang et al. 2015). ChREBP is a transcription factor
which mediates the expression of lipogenic genes in
response to glucose (Yamashita et al. 2001) and SREBP-1c is
a transcription factor which mediates the expression of
ARCHIVES OF PHYSIOLOGY AND BIOCHEMISTRY 7
lipogenic genes in response to insulin (Foufelle et al. 1992).
It is well known that high glucose concentration stimulates
the mRNA expression of the ChREBP gene, independently of
insulin. Whereas, high insulin level associated with consump-
tion of high carbohydrate diet increased the mRNA expres-
sion of the SREBP-1c gene (Dentin et al. 2004, Iizuka &
Horikawa, 2008). Thus, as expected, the SLCD-fed animals,
which were on a high carbohydrate diet and which had high
glucose and insulin levels, had significantly higher ChREBP
and SREBP-1c mRNA expression than the ND-fed animals.
Together, ChREBP and SREBP-1c appeared to be responsible
for the increase in lipogenesis resulting from high carbohy-
drate intake (Zhang et al., 2015).
In comparison to ND-fed animals, SLCD-fed animals also
had higher mRNA expression of the downstream target
genes which mediate lipogenesis: ACC1, FAS and SCD1.
These downstream genes are related to fatty acid synthesis
and are transcriptionally regulated by both ChREBP and
SREBP-1c (Bennett et al. 1995; Lopez et al. 1996; Sato et al.
2000). The above-mentioned downstream target genes were
also downregulated by spirulina in SLCD þ Spi-fed animals.
This transcriptional effect of spirulina may be due to an
improvement in plasma glucose and insulin concentrations
in close agreement with previous studies demonstrating the
hypoglycaemic and the hypo-insulinemic effects of spirulina
(Parikh et al. 2001; Strable and Ntambi 2010).
In addition to the regulation of lipogenic gene expression,
excess amounts of glucose via the glycolytic pathway can
provide a carbon source, such as acetyl-CoA, to the lipogen-
esis pathway (Ameer et al. 2014). Acetyl-CoA is then con-
verted to malonyl-CoA by ACC. FAS catalyses the formation
of palmitic acid from malonyl-CoA and acetyl-CoA. Palmitic
acid is then elongated and desaturated by long-chain fatty
acid elongase 6 (ELOVL6) and SCD1 to generate monoun-
saturated fatty acids such as palmitoleic and oleic acids,
which are the major fatty acid constituents of triglycerides
(Kawano and Cohen 2013). In this study, SLCD-fed animals
had high plasma concentrations of palmitic (C16:0), palmito-
leic (C16:1), stearic (C18:0) and oleic acids (C18:1). This can
be attributed to the high mRNA expression of ACC1, FAS
and SCD1 in the liver. Also, SLCD-fed P. obesus receiving a
diet supplemented with spirulina had lower concentrations
of the above-mentioned fatty acids. Therefore, it is likely that
spirulina inhibits fatty acid synthesis in the liver through a
decrease in the mRNA expression of the nuclear factors
ChREBP and SREBP-1c. This can ultimately result in a reduc-
tion in the mRNA expression of ACC1, FAS and SCD1, as well
as a decrease in the plasma concentration of fatty acids.
Furthermore, the high liver weight and fat content in SLCD-
fed P. obesus was associated to high hepatic and blood tri-
glyceride concentrations. The high amount of liver fatty acids
(as generated by ACC1, FAS and SCD1) could have contrib-
uted to the high liver and blood triglyceride concentration in
SLCD-fed animals. In contrast, P. obesus supplemented with
spirulina showed low hepatic and plasmatic triglyceride con-
centrations, in close agreement with previous studies dem-
onstrating the hypolipidemic effect of spirulina (Kato et al.
1984; Torres-Dur
an et al. 1999; Rodrıguez-Hern
andez et al.
8 H. SAIDI ET AL.
2001). This hypolipidemic effect in SLCD þ Spi animals was
associated with a downregulation in the mRNA expression of
ACC1, FAS and SCD1. In addition, the ratio SCD-act (18:1 n-9/
18:0), reflecting the enzymatic conversion of C18:0 to C18:1,
was twofold increased in SLCD-fed compared to ND-fed ani-
mals; this normalised with spirulina supplementation, in
accordance with the mRNA expression of SCD1 in the liver.
Obesity is also characterised by a state of chronic inflam-
mation (Moschen et al. 2011), which may contribute to the
development of insulin resistance and type 2 diabetes (Xu
et al. 2003; McArdle et al. 2013). In addition, ectopic fat
deposition in the liver enhances the inflammatory response
and the release of pro-inflammatory cytokines by Kupffer
cells (Asrih and Jornayvaz 2013). In this study, SLCD-fed ani-
mals had high mRNA expression of the pro-inflammatory
cytokines IL-1b and TNF-a in the liver. High hepatic levels of
pro-inflammatory cytokines mRNA expression such as TNF-a
and/or IL-1b can promote insulin resistance (Cai et al. 2005).
The reduction in these inflammatory markers has been asso-
ciated with improved insulin sensitivity and glucose metabol-
ism during insulin resistance (Hotamisligil et al. 1993;
Weisberg et al. 2003). In this study, spirulina supplementa-
tion decreased the mRNA expression of pro-inflammatory
cytokines TNF-a and IL-1b in the liver. These results are con-
sistent with those of Ku et al. (2013), who have reported the
anti-inflammatory effect of spirulina. This effect may result
from the inhibition of the nuclear translocation of NF-jB, a
transcription factor which mediates the production of pro-
inflammatory cytokines. Ultimately, the anti-inflammatory
effect of spirulina may be involved in the prevention of insu-
lin resistance (Cai et al. 2005; Tilg 2010).
Conclusion
In this study, supplementation of SLCD-fed P. obesus with
spirulina resulted in a reduction in body weight gain and
plasma lipids. This was associated with the downregulation
of the mRNA expression of genes involved in lipogenesis,
including transcription factors (ChREBP and SREBP-1c) and
their downstream targets (ACC1, FAS and SCD1) involved in
fatty acid synthesis. The reduction in the expression of lipo-
genic genes by spirulina can be attributed to its ability to
lower plasma insulin and glucose concentrations. It is also
possible that spirulina enhanced insulin sensitivity in SLCD-
fed P. obesus by downregulating the mRNA expression of the
pro-inflammatory cytokines TNF-a and IL-1b. The findings of
this study add to current knowledge about the effects of
spirulina supplementation on diet-induced obesity. This may
ultimately provide clinically-relevant insight into the use of
spirulina as a supplement to prevent obesity and its associ-
ated metabolic disorders.
Disclosure statement
No potential conflict of interest was reported by the author(s).
Funding
This work was supported by the Algerian Ministry of Higher Education &
Scientific Research programme under grant [F 0022014 0100].
ORCID
Hamza Saidi http://orcid.org/0000-0001-8340-995X
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