BACKGROUND: Adipose and hepatic metabolic dysfunctions are critical comorbidities that also aggravate insulin resistance in obese individuals. Melatonin is a low-cost agent and previous studies suggest that its use may promote metabolic health. However, its effects on some comorbidities associated with obesity are unknown. Herein, we investigated the hypothesis that melatonin supplementation would attenuate adipose-hepatic metabolic dysfunction in high fat diet (HFD)-induced obesity in male Wistar rats. MATERIALS AND METHODS: Twenty-four adult male Wistar rats (n = 6/group) were used: Control group received vehicle (normal saline), obese group received 40% high fat diet, melatonin-treated group received 4 mg/kg of melatonin, and obese plus melatonin group received 40% HFD and melatonin. The treatment lasted for 12 weeks. RESULTS: HFD caused increased food intake, body weight, insulin level, insulin resistance and plasma and liver lipid but decreased adipose lipid. In addition, HFD also increased plasma, adipose and liver malondialdehyde, IL-6, uric acid and decreased Glucose-6-phosphate dehydrogenase, glutathione, nitric oxide and circulating obestatin concentration. However, these deleterious effects except food intake were attenuated when supplemented with melatonin. CONCLUSION: Taken together, the present results indicate that HFD exposure causes adipose-hepatic metabolic disturbance in obese animals, which are accompanied by oxidative stress and inflammation. In addition, the present results suggest that melatonin supplementation attenuates adipose-hepatic metabolic dysfunction, accompanying obesity by suppression of oxidative stress/inflammation-dependent mechanism and increasing circulating obestatin.
BACKGROUND: Adipose and hepatic metabolic dysfunctions are critical comorbidities that also aggravate insulin resistance in obese individuals. Melatonin is a low-cost agent and previous studies suggest that its use may promote metabolic health. However, its effects on some comorbidities associated with obesity are unknown. Herein, we investigated the hypothesis that melatonin supplementation would attenuate adipose-hepatic metabolic dysfunction in high fat diet (HFD)-induced obesity in male Wistar rats. MATERIALS AND METHODS: Twenty-four adult male Wistar rats (n = 6/group) were used: Control group received vehicle (normal saline), obese group received 40% high fat diet, melatonin-treated group received 4 mg/kg of melatonin, and obese plus melatonin group received 40% HFD and melatonin. The treatment lasted for 12 weeks. RESULTS: HFD caused increased food intake, body weight, insulin level, insulin resistance and plasma and liver lipid but decreased adipose lipid. In addition, HFD also increased plasma, adipose and liver malondialdehyde, IL-6, uric acid and decreased Glucose-6-phosphate dehydrogenase, glutathione, nitric oxide and circulating obestatin concentration. However, these deleterious effects except food intake were attenuated when supplemented with melatonin. CONCLUSION: Taken together, the present results indicate that HFD exposure causes adipose-hepatic metabolic disturbance in obese animals, which are accompanied by oxidative stress and inflammation. In addition, the present results suggest that melatonin supplementation attenuates adipose-hepatic metabolic dysfunction, accompanying obesity by suppression of oxidative stress/inflammation-dependent mechanism and increasing circulating obestatin.
Obesity has become a global epidemic in the twenty-first century. Overweight
individuals aged 18 and above accounted for more than 1.9 billion people in 2016.
Over 650 million adults in this group were overweight or obese, 39% were overweight
while over 13% of the group were obese. Thus, obesity prevalence nearly tripled
globally between 1975 and 2016. In 2020, 39 million children under the age of five
were overweight or obese [1].
Obesity is a multifaceted, diverse disease influenced by hormones, nutritional
consumption, sedentary lifestyles, physical activity, genetics, and environmental
variables [2, 3]. This metabolic disease is
rising with comorbidities, including non-alcoholic fatty liver disease (NAFLD) that
reduce life quality and expectancy, primarily due to cardiometabolic problems [4]. The pathogenesis of
cardiometabolic dysfunctions are low-grade systemic inflammation and insulin
resistance caused by cytokines. These cytokines are released by excess adipose
tissue in the body, especially in the visceral site [5-7].In the quest for effective control of obesity, four hormones were discovered to have
a link; insulin, leptin, ghrelin and obestatin and the growth hormone secretagogue
receptor (GHS-R) [8]. Insulin
is secreted in the pancreas by β-cells islets of Langerhans and has a variety of
biological functions, such as body weight regulation and glucose homeostasis [9, 10]. However, insulin resistance arises as a
result of obesity and obesity-related dysfunctions including type 2 diabetes
mellitus (T2DM) and cardiovascular disorders. Hyperinsulinemia, resulting from
either hypersecretion or reduced insulin clearance, is a symptom of obesity and can
lead to IR sensitivity [11,
12]. Leptin was the
initial cytokine derived from adipose tissue linked with energy balance [13]) and is an anorexigenic
hormone produced mostly by adipose tissue. Leptin synthesis and secretion into
circulation are increased when fat depots expand in conjunction with a favorable
energy balance [14]. It has
been observed that obesity promotes hyperleptinemia and leptin resistance [15]. Ghrelin initially
identified as an endogenous ligand of the growth hormone secretagogue receptor
(GHSR1) is an orexigenic peptide. It is derived primarily from the stomach and a
peripheral signal that promotes food intake [16]. Obese people have lower plasma ghrelin
level and their meal-related ghrelin variations are similarly affected [17]. A study showed that there
was decreased ghrelin sensitivity after the administration of leptin, implying that
the increased leptinemia observed in obesity is responsible for the resistance of
ghrelin [18].Associated with the pathophysiology of obesity-related metabolic dysfunctions are
hyperleptinemia and hyperinsulinemia, and body adiposity in obesity is relative to
insulin and leptin levels in the circulation. Ghrelin dysregulation can also occur
in obesity and play a role in mediating some of the pathological signs and symptoms
[19]. Obestatin is a
23-amino acid anorexic hormone, a peptide that is involved in appetite control and
long-term energy regulation together with ghrelin [20]. Ghrelin and obestatin are both derived
from a single preproghrelin gene and produced by post-translation modification of
preproghrelin but obestatin has a distinct terminus [21]. Hence, it is reported to have opposite
effect on food intake as ghrelin [22]. It is an anorexigenic hormone that suppresses appetite and
gastrointestinal motility and modulates growth hormone and lipid metabolism [23, 24]. However, previous studies have reported
that obestatin acts as antagonist to the actions of ghrelin on appetite, food
intake, gastric emptying and the secretion of growth hormone [25, 26]. Zhao et al., also
reported that obestatin is reduced in obese humans [27].Melatonin is a hormone secreted by the pineal gland in the dark hours via the control
of the suprachiasmatic nucleus of the hypothalamus. It is associated with many
physiological roles in the central nervous system, sleep and wakefulness cycles,
energy metabolism and thermoregulation, immune and endocrine regulation among others
[28]. Melatonin is the
significant mediator molecule in the incorporation of the cyclic environment and the
circadian distribution of physiological and cognitive processes, as well as the
optimization of energy hemostasis and regulation of body weight, which are important
for a healthy metabolism [29]. The islets of Langerhans of the pancreas are important sites of the
action of melatonin where it stimulates the synthesis and secretion of insulin and
glucagon synthesis in reference to the regulation of energy metabolism. The
melatonin receptors MT1 and/or MT2- facilitated the action of melatonin decreasing
the glucose-stimulated insulin secretion (GSIS) in the isolated pancreatic islets
and insulinoma beta cells in rats [30, 31]. Through
the regulation of GLUT4 expression or triggering the insulin signaling pathway,
melatonin functions in potentiating central and peripheral action of insulin. Thus,
it induces, via its G-protein-coupled membrane receptors, the phosphorylation of the
insulin receptor and its intracellular substrates. It has also been considered that
melatonin’s association with all the physiological processes typical of the daily
activity-wakefulness/rest-sleep rhythm may impact body weight and possibly
contribute to energy homeostasis [28, 32]. However,
information on the role of melatonin in obesity-associated adipose-hepatic metabolic
dysregulation is lacking. The present study was therefore designed to investigate
the role of melatonin on adipose-hepatic metabolic perturbations in obese male
Wistar rats. The study in addition determined the probable involvement of
obestatin.
2. Materials and methods
2.1. Animals
All experimental protocols for this study were conducted in accordance with the
National Institutes of Health Guide for the Care and Use of Laboratory Animals
and was approved by the Institutional Ethical Review Board of Afe Babalola
University, Nigeria (ABUADERC/10/2020), and every effort was made to minimize
both the number of animals used and their suffering. Twenty-four male Wistar
rats weighing 170–200 g were procured from the animal house of the College of
Health Sciences, Afe Babalola University, Nigeria. Rats had unrestricted access
to standard rat chow and tap water. After 2 weeks of acclimatization, the
animals were randomly assigned into four groups (n = 6 per group). Rats were
maintained in a colony under standard environmental conditions of temperature
(22–26°C), relative humidity (50–60%), and 12-hour dark/light cycle.
2.2. Treatment
Control (CTL) received diet and distilled water (vehicle; po),
Melatonin-treated group (MLT-treated) received melatonin (4 mg/kg body weight;
Sigma-Aldrich, St Louis, MI), Obese group (OBS) received 40% high fat diet (HFD)
and Obese with melatonin-treated group (OBS+MLT-treated) received combination of
high fat and melatonin daily. Animals were treated with melatonin between
8:00–10:00 am and obesity was induced by exposing the animals to 40% HFD ad
libitum as previously described [33] The administration lasted for 12 weeks. Initial and final body
weights were determined, and body weight gain was estimated. In addition, daily
food and water consumptions were monitored for week 0 (initial) and week 12
(final) by subtracting the left-over food and water after 24 h from the food and
water that were introduced to the animals. The changes in food and water
consumptions were estimated by subtracting the initial consumption from the
final consumption.
2.3. Sample preparation
After 12 weeks of administration, the animals were fasted overnight for 12 h.
Thereafter, the animals were anesthetized by intraperitoneal injection of 50
mg/kg b.w. of sodium pentobarbital. Cardiac
puncture was used for the collection of blood into the heparinized tube and
blood was centrifuged at room temperature for 5 mins at 3000 rpm. Plasma was
decanted and stored frozen until when it was needed for the biochemical
analysis.
2.4. Preparation of liver and adipose tissue homogenates
After weighing the liver and visceral fat, 100 mg section of each tissue was
carefully removed and homogenized with a glass homogenizer in phosphate buffer
solution, centrifuged at 10000 rpm for 10 min at 4°C.
2.5. Blood glucose and insulin resistance (IR)
Fasting blood glucose was determined with a hand-held glucometer
(ONETOUCH®-LifeScan, Inc., Milpitas, CA, USA). Insulin resistance was estimated
using the Homeostatic model assessment for IR (HOMA-IR = fasting glucose
(mmol/l) * fasting insulin (μU/l)/22.5) [34, 35].
2.6. Biochemical assays
2.6.1. Plasma insulin
The plasma level of insulin was determined with Rat ELISA kits obtained from
Calbiotech Inc. (Cordell Ct., El Cajon, CA 92020, USA) in compliance with
the manufacturer’s procedures and based on the direct sandwich technique in
which two monoclonal antibodies are directed against separate antigenic
determinants on the insulin molecule.
2.6.2. Obestatin
Obestatin concentration was determined in the plasma using Rat ELISA kits
obtained from Calbiotech Inc. (El Cajon, USA) in compliance with the
manufacturer’s assay procedure.
2.6.3. Lipid profile
Concentration of triglycerides (TG) and total cholesterol (TC) were estimated
in the plasma, liver and adipose tissue homogenates by standardized
colorimetric methods using reagents obtained from Fortress Diagnostics Ltd.
(Antrim, UK).
2.6.4. Oxidative stress markers
Malondialdehyde (MDA) was determined from the plasma, liver and adipose
tissue homogenate by standard non-enzymatic spectrophotometric method using
assay kits from Randox Laboratory Ltd. (Co. Antrim, UK). This method was
carried out as previously described [35], whereas Glutathione (GSH) was
determined using non-enzymatic spectrophotometric method with assay kits
obtained from Oxford Biomedical Research Inc. (Oxford, USA). Glutathione was
determined by spectrophotometric method based on the oxidation of GSH in the
sample by the sulfhydryl reagent 5,5′-dithio-bis (2-nitrobenzoic acid)
(DTNB) to form the yellow derivative 5′-thio-2-nitrobenzoic acid (TNB),
measured at 412 nm. While Glucose-6-phosphate dehydrogenase (G6PD) activity
was determined from the plasma, liver and adipose tissue using standard
spectrophotometric method with assay kits obtained from Calbiotech Inc. (El
Cajon, USA).
2.6.5. Interleukin-6 (IL-6), nitric oxide and uric acid
concentration
Plasma, liver and adipose tissue concentration of IL-6 was determined by the
quantitative standard sandwich ELISA technique using monoclonal antibody
specific for these parameters with rat kits obtained from Elabscience
Biotechnology Inc. (Wuhan, Hubei, P.R.C., China). Nitric oxide was assayed
spectrophotometrically by measuring the accumulation of its stable
degradation products, nitrate and nitrite using kits from Oxford Biomedical
Research Inc., (Oxford, UK). This kit employs the NADH-dependent enzyme
nitrate reductase for conversion of nitrate to nitrite prior to the
quantification of nitrite using Griess reagent—thus providing for accurate
determination of total NO production. Furthermore, uric acid uric
concentration was estimated by non-enzymatic colorimetric method using assay
kits from Randox Laboratory Ltd. (Co. Antrim, UK) and in compliance with the
manufacturer’s assay procedures.
2.7. Statistical analysis
Shapiro-Wilk test was used to confirm the data distribution, and the data were
normally distributed. All data were expressed as means ± SD. Statistical group
analysis was performed using the Graphpad prism 5. One-way ANOVA was used to
compare the mean values of variables among the groups. Bonferroni’s test was
used for post hoc analysis. Statistically significant
differences were accepted at p less than 0.05.
3. Results
3.1. Effects of melatonin on food intake, water intake and body weight in
HFD-induced obese rats
There was a significant increase (p<0.05) in food intake in obese and
OBS+MLT-treated rats compared to the control group. Supplementation with
melatonin did not significantly decrease the food intake as shown in
OBS+MLT-treated rats compared with obese rats. In addition, body weight was
increased in obese rats when compared to the control group. However, melatonin
decreased the body weight. There was no alteration in water intake in all the
experimental groups compared to the control group (Table 1).
Table 1
Melatonin attenuates excess body weight but not food intake in
HFD-induced obese animals.
GROUPS
CTL
MLT
OBS
OBS+MLT
Food intake (g/day)
Initial
25.22 ± 0.81
33.15 ± 2.30
30.59 ± 4.24
31.85 ± 2.27
Change
8.01 ± 2.71
5.33 ± 1.79
19.21 ± 3.77*
14.42 ± 0.35*
Water intake (mL/day)
Initial
32.62 ± 1.47
27.79 ± 3.16
26.63 ± 3.43
35.63 ± 3.43
Change
7.34 ± 2.52
5.44 ± 10.19
6.18 ± 5.44
5.86 ± 3.88
Body weight (g)
Initial
172.71 ± 6.41
174.93 ± 8.12
171.00 ± 6.65
171.43 ± 5.70
Gain
44.40 ± 6.70
36.67 ± 9.30
75.87 ± 4.72*
26.69 ± 3.62#
Data are expressed as mean ± SD. n = 6 and analyzed by one-way ANOVA
followed by Bonferroni post hoc test.
(*p<0.05 vs. CTL;
#p<0.05 vs. OBS). Control (CTL); Melatonin
(MLT); Obesity (OBS).
Data are expressed as mean ± SD. n = 6 and analyzed by one-way ANOVA
followed by Bonferroni post hoc test.
(*p<0.05 vs. CTL;
#p<0.05 vs. OBS). Control (CTL); Melatonin
(MLT); Obesity (OBS).
3.2. Effects of melatonin on glucose homeostasis in HFD-induced obese
rats
There was a significant increase (p<0.05) in fasting plasma
insulin but no alteration in blood glucose in obese group compared to the
control group. However, supplementation with melatonin decreased the fasting
plasma insulin in OBS+MLT group compared to the untreated obese group.
Similarly, insulin resistance was observed in the obese animals compared with
control animals. Administration of melatonin significantly reduced insulin
resistance in OBS+MLT group compared to the untreated obese group (Fig 1).
Fig 1
Effects of melatonin on blood glucose (A), insulin (B) and insulin
resistance (C) HFD-induced obese animals. Data are expressed as mean ±
SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 vs.
CTL; #p<0.05 vs. OBS). Control (CTL); Melatonin
(MLT); Obesity (OBS).
Effects of melatonin on blood glucose (A), insulin (B) and insulin
resistance (C) HFD-induced obese animals. Data are expressed as mean ±
SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 vs.
CTL; #p<0.05 vs. OBS). Control (CTL); Melatonin
(MLT); Obesity (OBS).
3.3. Effects of melatonin on plasma, adipose and liver triglyceride and total
cholesterol in HFD-induced obese rats
There was a significant increase (p<0.05) in plasma and liver
TG and TC but a decrease in adipose triglyceride and total cholesterol in obese
group compared to the control group. However, supplementation with melatonin
decreased the plasma and liver TG and TC and as well increased the TG and TC
concentrations in the adipose tissue of OBS+MLT group compared to the untreated
obese group (Fig 2).
Fig 2
Effects of melatonin on plasma, adipose and liver triglyceride (A-C) and
total cholesterol (D-F) in HFD-induced obese rats. Data are expressed as
mean ± SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 vs.
CTL; #p<0.05 vs. OBS). Control (CTL); Melatonin
(MEL); Obesity (OBS); Total cholesterol (TC).
Effects of melatonin on plasma, adipose and liver triglyceride (A-C) and
total cholesterol (D-F) in HFD-induced obese rats. Data are expressed as
mean ± SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 vs.
CTL; #p<0.05 vs. OBS). Control (CTL); Melatonin
(MEL); Obesity (OBS); Total cholesterol (TC).
3.4. Effect of melatonin on malondialdehyde in HFD-induced obese rats
There was a significant increase (p<0.05) in plasma, adipose
and liver MDA in obese group compared to the control group. However,
supplementation with melatonin decreased the plasma, adipose and liver MDA in
OBS+MLT group compared to the untreated obese group (Fig 3).
Fig 3
Effect of melatonin on plasma, adipose and liver malondialdehyde (A-C) in
HFD-induced obese rats. Data are expressed as mean ± SD. n = 6 and
analyzed by one-way ANOVA followed by Bonferroni post hoc
test. (*p<0.05 vs. CTL;
#p<0.05 vs. OBS). Control (CTL); Melatonin
(MLT); Obesity (OBS).
Effect of melatonin on plasma, adipose and liver malondialdehyde (A-C) in
HFD-induced obese rats. Data are expressed as mean ± SD. n = 6 and
analyzed by one-way ANOVA followed by Bonferroni post hoc
test. (*p<0.05 vs. CTL;
#p<0.05 vs. OBS). Control (CTL); Melatonin
(MLT); Obesity (OBS).
3.5. Effect of melatonin on G6PD and GSH in HFD-induced obese rats
There was a significant decrease (p<0.05) in plasma, adipose
and liver G6PD activity and glutathione concentration in obese group compared to
the control group. Nonetheless, supplementation with melatonin increase the
plasma, adipose and liver G6PD and glutathione concentration in OBS+MLT group
compared to the untreated obese group (Fig 4).
Fig 4
Effects of melatonin on plasma, adipose and liver Glucose-6-phosphate
dehydrogenase (A-C) and glutathione (D-F) in HFD-induced obese rats.
Data are expressed as mean ± SD. n = 6 and analyzed by one-way ANOVA
followed by Bonferroni post hoc test.
(*p<0.05 VS. CTL;
#p<0.05 VS. OBS). Control (CTL);
Melatonin (MLT); Obesity (OBS); Glucose 6 phosphate dehydrogenase
(G6PD); Glutathione (GSH).
Effects of melatonin on plasma, adipose and liver Glucose-6-phosphate
dehydrogenase (A-C) and glutathione (D-F) in HFD-induced obese rats.
Data are expressed as mean ± SD. n = 6 and analyzed by one-way ANOVA
followed by Bonferroni post hoc test.
(*p<0.05 VS. CTL;
#p<0.05 VS. OBS). Control (CTL);
Melatonin (MLT); Obesity (OBS); Glucose 6 phosphate dehydrogenase
(G6PD); Glutathione (GSH).
3.6. Effects of melatonin on IL-6 and uric acid concentration in HFD-induced
obese rats
There was a significant increase (p<0.05) in plasma, adipose
and liver IL-6 and uric acid concentration in obese group compared to the
control group. However, supplementation with melatonin decrease the plasma and
liver but not adipose uric acid concentration in OBS+MLT group compared to the
untreated obese group (Fig
5)
Fig 5
Effects of melatonin on plasma, adipose and liver interleukin-6 (A-C) and
uric acid concentration (D-F) HFD-induced obese rats. Data are expressed
as mean ± SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 VS.
CTL; #p<0.05 VS. OBS). Control (CTL),
Melatonin (MLT), Obesity (OBS).
Effects of melatonin on plasma, adipose and liver interleukin-6 (A-C) and
uric acid concentration (D-F) HFD-induced obese rats. Data are expressed
as mean ± SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 VS.
CTL; #p<0.05 VS. OBS). Control (CTL),
Melatonin (MLT), Obesity (OBS).
3.7. Effects of melatonin on nitric oxide concentration in HFD-induced obese
rats
There was a significant reduction (p<0.05) in plasma, adipose
and liver nitric oxide concentration in obese group compared to the control
group. However, supplementation with melatonin increased the plasma, adipose and
liver nitric oxide concentration in OBS+MLT group compared to the untreated
obese group (Fig 6).
Fig 6
Effects of melatonin on plasma, adipose and liver nitric oxide
concentration (A-C) in HFD-induced obese rats. Data are expressed as
mean ± SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 VS.
CTL; #p<0.05 VS. OBS). Control (CTL);
Melatonin (MLT); Obesity (OBS).
Effects of melatonin on plasma, adipose and liver nitric oxide
concentration (A-C) in HFD-induced obese rats. Data are expressed as
mean ± SD. n = 6 and analyzed by one-way ANOVA followed by Bonferroni
post hoc test. (*p<0.05 VS.
CTL; #p<0.05 VS. OBS). Control (CTL);
Melatonin (MLT); Obesity (OBS).
3.8. Effects of melatonin on obestatin level in HFD-induced obese
rats
There was a significant decrease (p<0.05) in the level of plasma obestatin
concentration in obese animal when compared to the control animal. However,
supplementation with melatonin significantly increased the obestatin level in
animal with obesity (Fig
7).
Fig 7
Effects of melatonin on circulating obestatin concentration in
HFD-induced obese rats.
Data are expressed as mean ± SD. n = 6 and analyzed by one-way ANOVA
followed by Bonferroni post hoc test.
(*p<0.05 VS. CTL;
#p<0.05 VS. OBS). Control (CTL),
Melatonin (MLT), Obesity (OBS).
Effects of melatonin on circulating obestatin concentration in
HFD-induced obese rats.
Data are expressed as mean ± SD. n = 6 and analyzed by one-way ANOVA
followed by Bonferroni post hoc test.
(*p<0.05 VS. CTL;
#p<0.05 VS. OBS). Control (CTL),
Melatonin (MLT), Obesity (OBS).
4. Discussion
The data from the present study demonstrated that melatonin reversed the
adipose-hepatic metabolic comorbidities associated with obesity in male Wistar rats
by suppression of oxidative stress, inflammation and increasing circulating
obestatin. Earlier studies have demonstrated a significant decrease in the level of
obestatin in obese children [36], and the present observation that revealed a significant decrease in
the circulating levels of obestatin in obese animals compared to control group is
consistent with previous studies. In addition, Ren et al., reported
that the levels of obestatin were significantly lower in obese subjects and
correlated negatively with body mass index (BMI), insulin, glucose and insulin
resistance [37]. However, as
revealed in the results of the present studies, in addition to decreased obestatin
level, obesity is also characterized with insulin resistance, hyperinsulinemia and
excess body weight, which are consistent with previous observations [23, 37]. Besides, the present results also showed
an increase in food intake in obese animals compared to the control group, which
might contribute to excess body weight possibly due to reduced energy utilization
resulting from insulin resistance. As already demonstrated, HFD causes insulin
resistance in experimental rodents [38, 39].Furthermore, compensatory hyperinsulinemia observed in obese rats might contribute to
normal blood glucose. However, evidence exists that hyperinsulinemia signals
oxidative stress, especially in combination with insulin resistance causing adipose
tissue inflammation that characterized obesity [40-42]. In addition, obesity is considered a
syndrome of excessive visceral adiposity and is linked with metabolic dysfunctions.
Metabolic dysfunctions are characterized by cardiovascular and diabetes risk factors
such as abdominal adiposity, hypertension, reduction in high‐density lipoprotein
(HDL), increased triglycerides and glucose intolerance [43]. In this study, there was a significant
increase in plasma and liver TG and TC with corresponding decrease in adipose TG and
TC in obese group compared to the control group, which might lead to hepatic
lipotoxicity that triggered oxidative stress in obese animals as shown by elevated
hepatic lipid peroxidation (MDA) with a decrease in G6PD/GSH-dependent antioxidant
capacity. Previous studies have documented that a decrease in obestatin could also
contribute to increase in TC with consequent oxidative stress [44-46]. Therefore, in this study obesity-induced
hepatic oxidative stress is associated with a decrease in circulating level of
obestatin and excessive lipolysis that led to reduction in adipose TG and TC.In addition, the present study showed a significant increase in plasma, adipose and
liver IL-6 and uric acid concentration and a significant reduction in plasma and
adipose nitric oxide concentration in obese group compared to the control group.
These observations are consistent with earlier studies, including a recent study
from our laboratory animals which demonstrates that metabolic related syndrome such
as obesity causes inflammation in the metabolic tissues, particularly the adipose
and hepatic tissues [47,
48] and these are well
documented pathological features of non-alcoholic fatty liver disease [47, 49]. This therefore suggests obesity as a
predictor of fatty liver disease, which may become one of the common reasons for
liver transplantations by 2030 especially in developed countries [49, 50]. Other studies have also reported that
obestatin could be protective against oxidative stress and inflammation [51, 52]. Therefore, decrease level of obestatin
might in part contribute to adipose/hepatic inflammation that characterized obese
animals compared to the control group.Interestingly, this study also showed that melatonin supplementation reduced the body
weight of obese rats while elevating their levels of obestatin, though without a
significant decrease in food intake compared to the untreated obese group. The
treatment with melatonin also decreased the plasma and liver triglyceride and total
cholesterol in OBS+MLT group compared to the untreated obese group. In addition, the
elevated fasting plasma insulin and insulin resistance were reversed by melatonin
supplementation, which might be due to increase in insulin sensitivity as earlier
reported by McHugh and Cheng that administration of melatonin improves insulin
sensitivity and insulin level [53]. This possibly improved glucose/lipid metabolism and thus prevents
excess energy storage/visceral adiposity that constitutes excess body weight gain.
This observation seems similar to a number of studies that demonstrated improved
body composition following administration of melatonin [32, 39, 54]. Besides, melatonin has also been reported
to modulate cyclic adenosine monophosphate (cAMP) and cyclic gaunosine monophosphate
(cGMP levels), which regulate glucose and energy homeostasis [55] corroborating that melatonin promotes body
maintenance. Other studies have also shown that administration of melatonin
prevented high glucose or lipid levels in pinealectomized rats [56]. However, melatonin in
addition to improving body weight, insulin sensitivity also demonstrated antioxidant
effect against hepatic and adipose oxidative stress with corresponding decrease in
lipid peroxidation and enhancement of G6PD/GSH-dependent antioxidant barrier in
obese animals compared to the untreated obese group. Likewise, the administration of
melatonin increased the plasma, liver and adipose nitric oxide concentration and
decreased the plasma and liver uric acid and IL-6 concentration with corresponding
decrease in adipose IL-6 in obese rats compared to the untreated obese group,
suggesting that treatment with melatonin mitigates inflammatory signals induced by
insulin resistance/hyperinsulinemia with consequent decrease in adipose/hepatic
inflammation. In consonance with previous study, melatonin acts as a free radical
scavenger that eliminates reactive oxygen and promotes the action and expression of
endogenous antioxidants [26,
57]. Our results are also
consistent with a number of studies who have demonstrated anti-inflammatory,
anti-proliferative and apoptotic properties of melatonin in experimental animals
[58-60]. Nevertheless, the present results are not
without limitations in such that the molecular mechanisms underlying the regulatory
role of melatonin in obese animals, and the link between obestatin and other
biochemical parameters were not investigated. However, the present data provide a
justification for further study of molecular mechanisms, and the data perhaps,
provide clinical insight into the diagnosis and management of obesity-associated
adipose-hepatic metabolic comorbidities.
5. Conclusion
Taken together, the present results indicate that HFD exposure causes adipose-hepatic
metabolic disturbance in obese animals, which are accompanied by oxidative stress
and inflammation. In addition, the present results suggest that melatonin
supplementation attenuates adipose-hepatic metabolic dysfunction, accompanying
obesity by suppression of oxidative stress/inflammation-dependent mechanism and
increasing circulating obestatin.(XLSX)Click here for additional data file.
Fig 1
Fig 2
Fig 3
Fig 4
Fig 5
Fig 6
Fig 7