Is the effect of high fat diet on lipid and carbohydrate metabolism related to inflammation?
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- Abd El-Hamid Ibrahim, I.A., Abd El-Aziz, M.F.M., Ahmed, A.F. et al. Mediterr J Nutr Metab (2011) 4: 203. doi:10.1007/s12349-011-0056-9
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Obesity is an emerging health problem in world wide countries. High fat diet (HFD), which is the main cause of obesity, induces inflammation and affects both lipid and carbohydrate metabolism. This study investigates the effect of inflammation induced by HFD on lipid and carbohydrate metabolism. This study involves two parts, in vivo and in vitro. In the in vivo part, adult male albino rats were fed a HFD for 8 and 14 weeks. After each feeding period, the following parameters were measured (body weight, blood glucose levels, oral glucose tolerance, serum insulin, insulin resistance, total cholesterol, HDLc, LDLc, triglycerides, free fatty acids, neutrophils percentage and serum CRP levels) in addition to pancreas tissue histopathology. In the in vitro part, insulin release from isolated perfused pancreas preparations was measured. Pancreas preparations were isolated from adult male albino rats fed a HFD for 16 weeks. Feeding rats a HFD for 8 and 14 weeks induced inflammation, impaired lipid profile and carbohydrate metabolism. Feeding rats a HFD for 16 weeks increased in vitro glucose-stimulated insulin release. HFD induces inflammation which plays an important role in the impairment of lipid and carbohydrate metabolism.
Feeding rats a HFD for prolonged periods induces a state of insulin resistance associated with diminished insulin-stimulated glycolysis and glycogen synthesis . This may be attributed to increased lipid availability and oxidation. The increased lipid oxidation causes impaired insulin stimulation of glycogen synthase (GS) activity in skeletal muscle . Also, it may be due to decreased glucose transporter (GLUT)-4 expression  or impaired insulin action on glucose transport in skeletal muscle .
The fatty acid composition of dietary fat may determine its effect on glucose metabolism and insulin action. Saturated and n-6 unsaturated fatty acids in the diet may lead to insulin resistance in experimental animals, whereas n-3 fatty acids, particularly the unsaturated types prevent the development of insulin resistance and ameliorate metabolic dysfunctions, including improvement of insulin sensitivity  and lowering of plasma triacylglycerol levels . This is attributed to increased hepatic fatty acid oxidation .
Also, it has been found that fat may contribute to activation of innate immunity  and that HFD intake increases adipose tissue IL-6 production . This may be attributed to increased visceral adiposity which is characterized by increased release of IL-6, TNF-α and CRP from adipocytes .
The new concept is that obesity or type 2 diabetes is a type of low grade inflammation. However, the debate is about which occurs first, (obesity or inflammation) and (insulin resistance or inflammation). Also, the role of low grade inflammation in the metabolic changes is still under investigation. This is what our work tries to explore.
In all experiments performed, 28 male adult albino rats, weighing 120–140 g, obtained from the national research centre, Egypt, were used. They were kept under constant environmental conditions throughout the period of investigation (13 h dark/11 h light cycles). The animals were housed in cages (7 rats/cage) with wood shaving bedding, and were fed a standard chow diet (SCD) (for 1 week as adaptation period) and tap water ad libitum. All procedures were made according to the ethical guidelines of the Canadian committee.
The SCD consisted of the following components:
Protein (25.6 g%), carbohydrate (50.3 g%), fat (5.1 g%), casein (20 g%), l-cystine (0.3 g%), maltodextrin (2.5 g%), corn starch (31.5 g%), sucrose (35 g%), soybean oil (2.5 g%), lard (2 g%), vitamins and minerals (6.2 g%), calories (3.4 kcal/g) .
Soy bean oil: 1-unsaturated fatty acids: 7% alpha-linolenic acid (C-18:3); 51% linoleic acid (C-18:2); and 23% oleic acid (C-18:1).
2-saturated fatty acids 4% stearic acid and 10% palmitic acid.
Lard: 1-unsaturated fatty acids: oleic acid 44–47%, palmitoleic acid 3%, Linoleic acid 6–10%.
2-saturated fatty acids: palmitic acid 25–28%, stearic acid 12–14%, myristic acid.
Induction of insulin resistance
Insulin resistance was induced according to the method described by Tacikowski et al.  with little modifications by feeding the rats a HFD for 8, 14 and 16 weeks.
The HFD consisted of the following components:
Protein (26 g%), carbohydrate (26 g%), fat (35 g%), casein (20 g%), l-cystine (0.3 g%), maltodextrin (12.5 g%), corn starch (31.5 g%), Sucrose (6.88 g%), soybean oil (2.5 g%), lard (24.5 g%), vitamins and minerals (1.82 g%), calories (5.2 kcal/g) .
Composition of soybean oil and lard is the same of SCD.
In vivo study
After 1-week adaptation period, rats were randomly distributed into 2 groups. The first group is the SCD group at which rats were fed a SCD for 8 and 14 weeks. The second group is the HFD group at which rats were fed a HFD for 8 and 14 weeks.
In vitro study
Animals were divided into two groups, one group fed a SCD, the second group fed a HFD for 16 weeks before isolation of their pancreas.
Isolation and perfusion of pancreas
The perfusion medium is a Krebs–Ringer-bicarbonate solution with 6% hydroxyethylstarch (HAES) and 0.25% bovine serum albumin and supplemented with 3 and 16.7 mM glucose. The pH of the final solution was always adjusted at 7.4 and checked using a pH meter. It was continuously gassed with a carbogen mixture of 95% O2 and 5% CO2 and the temperature was maintained constant at 37 °C using a thermostat. It was introduced into the celiac artery at a flow rate of 1 ml/min and the effluents were collected from the portal vein. The venous effluent was collected (after a single passage through the pancreas preparation) at different time intervals into pre-chilled tubes, then frozen and stored at −20 °C until the assay of insulin.
HFD components (Hilab company and local sources), Thiopental (EPICO company, Egypt), HAES (Intramed Division of Pharmacare LTD. South Africa), bovine serum albumin (Sigma Chemical. St Louis, Mo, USA), glucose (Adwic. Company Egypt), carbogen (Egyptian Gases Company, Cairo, Egypt).
Animals were killed and dissected. Pancreas was removed and kept in 10% formalin in saline solution and processed for histopathological examination.
Effect of feeding rats a HFD for 8 and 14 weeks on different parameters
HFD (8 weeks)
HFD (14 weeks)
Body weight (gm)
169.4 ± 6 (8 weeks)
182 ± 7 (14 weeks)
258.7 ± 10*
269.7 ± 7*
Blood glucose (mg/dl)
84.6 ± 4
114.3 ± 4*
130.3 ± 5*
AUC of OGTT (mg min/dl)
13,886 ± 170
16,788 ± 394*
23,411 ± 647*
Serum insulin (μIu/ml)
2.9 ± 0.2
2.8 ± 0.2
6.6 ± 0.7*
Insulin resistance (mmol μIu/ml)
0.6 ± 0.02
0.7 ± 0.07
1.97 ± 0.2*
C-reactive protein (μg/ml)
4.84 ± 0.2
6.6 ± 0.5*
6.77 ± 0.4*
7.2 ± 0.8
17.7 ± 1.6*
41 ± 5*
Total cholesterol (mg/dl)
52.3 ± 1.6
67.5 ± 7.8
80.4 ± 4.8*
49.4 ± 1.4
67.6 ± 3.7*
96.6 ± 7*
36.3 ± 2
27.7 ± 2.3*
26.9 ± 1.7*
6.12 ± 0.68
26.3 ± 4.76*
34.18 ± 1.7*
52.12 ± 3.25
46.8 ± 2.54
51.1 ± 1.1
Data analysis and statistical procedures
The difference between values (Mean ± SEM) was tested for significance by student’s t test for unpaired data  using SPSS program version 10. Results were considered significant at P > 0.05.
Graph Pad prism program version 5 was used to calculate the area under the insulin secretion time curves.
In vivo study
Effect of feeding rats a HFD for 8 and 14 weeks on oral glucose tolerance (OGT)
Blood glucose levels (mg/dl)
AUC of OGTT
84.67 ± 3.60
119.14 ± 2.95
125.28 ± 3.62
120.67 ± 3.56
112.20 ± 2.39
13,886.25 ± 170.19
HFD (8 weeks)
114.3 ± 4.31*
126.00 ± 6.00
139.5 ± 6.26*
155.1 ± 5.82*
146.4 ± 3.96*
16,788.7 ± 394.22*
HFD (14 weeks)
130.4 ± 4.94*
144.9 ± 9.5*
192.7 ± 7.6*
189.3 ± 9.9*
189.5 ± 3.81*
23,411.25 ± 647.21*
In vitro study
The effect of 3 and 16.7 mM of glucose on insulin release from the isolated perfused pancreas preparations obtained from rats fed a HFD
Area under the insulin secretion-time curves (AUC, min μIU/ml)
0 ± 0
213.8 ± 13.8
50.1 ± 4.7
308.3 ± 28.9
0 ± 0
343.8 ± 33.7*
63.1 ± 7.74
471.8 ± 45.8*
Discussion and conclusions
HFD caused a significant increase in body weight. This may be attributed to that HFD is less satiating than SCD leading to passive diet over-consumption and visceral obesity .
Increased body weight and visceral adiposity lead to elevation in inflammatory markers as observed with serum CRP levels and percentage of neutrophils creating a state of low grade inflammation. Increased inflammation associated with visceral adiposity may be due to increased oxidative stress. Production of reactive oxygen species (ROS) increased selectively in adipose tissue of obese mice, accompanied by augmented expression of NADPH oxidase and decreased expression of antioxidative enzymes. In cultured adipocytes, elevated levels of fatty acids increased oxidative stress via NADPH oxidase activation, and oxidative stress caused dysregulated production of adipocytokines (fat-derived hormones), including adiponectin, plasminogen activator inhibitor–1, IL-6, and monocyte chemotactic protein–1 .
This state of low grade inflammation leads to increase in fasting serum glucose, insulin levels, insulin resistance and impaired OGT.
Inflammation activates IKK causing serine phosphorylation of IRS-1, this results in decreased insulin activation of IRS-1 tyrosin phosphorylation, and decreased activation of IRS-1-associated PI3K. This is followed by a decrease in insulin-stimulated glucose uptake and glycogen synthesis causing insulin resistance, hyperglycemia and reduced OGT .
Down-regulation of PPARγ receptors during inflammation may play a role in insulin resistance development as this is associated with an increase in the levels of cytokines, reduced GLUT4 activity, elevated levels of resistin which is an insulin desensitizer and reduced PI3K activity in skeletal muscles .
The insulin resistance induced by inflammation may contribute to hyperinsulinemia away from the compensatory increased insulin release from beta cells, by causing insulin resistance in liver, leading to decreased insulin clearance and degradation and finally hyperinsulinemia.
The hyperinsulinemia itself initiates another cycle of insulin resistance by increasing delta desaturase enzyme levels, which increases conversion of omega 6 fatty acids to arachidonic acid, which in turn increases prostaglandin 2 synthesis causing increased production of cytokines and inflammation throughout the entire body .
Also, low grade inflammation leads to elevation in serum TG levels. Inflammation increases de novo fatty acid synthesis in liver , decreases fatty acid oxidation  and increases adipose tissue lipolysis which in turn increases fatty acid levels . All the preceding changes direct the fatty acids towards the liver to be involved in TG synthesis. The final result is a significant increase in serum TG levels as shown in the present study.
The increase in serum TC levels observed in the present study may also be due to inflammation. Inflammation increases hepatic cholesterol synthesis by increasing synthesis and activity of hydroxymethylglutaryl COA (HMG-COA) reductase enzyme , decreases hepatic cholesterol catabolism and excretion  and reduces reverse cholesterol transport (RCT)  resulting finally in a significant increase in serum TC levels as shown in our results.
Furthermore inflammation can increase the LDLc/HDLc ratio by decreasing LDL clearance from the circulation  as shown in our results. From the histopathological study, it is clear that, HFD reduced beta cells viability and activity and this may be attributed to inflammation.
The observation that, serum levels of CRP and percentage of neutrophils increased significantly at a time preceding the significant increase in serum insulin and FFA levels, indicates that inflammation initiates the incidence of insulin resistance not the reverse.
In the in vitro study, the elevation in glucose-stimulated insulin secretion (GSIS) may be attributed to elevated pancreatic level of urocortin 3 (Ucn3) mRNA. Urocortin is a corticotropin-releasing factor receptor 2 (CRFR2) agonist. Its level increases after feeding rats a HFD for prolonged periods. It stimulates insulin secretion particularly in presence of excess nutrients and high blood glucose levels .
Another possible mechanism by which HFD increases GSIS, is its ability to reduce the sex-determining region Y-box 6 (SOX6) mRNA level. Its level decreases by both a long term high fat feeding protocol in normal mice and in genetically obese ob/ob mice with a normal chow diet. This protein (SOX6) is expressed in adult pancreatic insulin-producing cells, over-expression of SOX6 decreased glucose-stimulated insulin secretion, which was accompanied by decreased ATP/ADP ratio, Ca2+ mobilization, proinsulin content, and insulin gene expression .
A recent hypothesis said that fatty acids activate the G-protein coupled receptor (GPCR) GPR40 also referred to as the free fatty-acid 1 receptor (FFA1R). This receptor is highly expressed in pancreatic beta cells and insulin-secreting cell lines, binding of fatty acids to GPR40 stimulates phospholipase C-mediated hydrolysis of phosphatidylinositol 4,5-biphosphate into diacylglycerol and inositol trisphosphate. This leads to the respective activation of protein kinase C and calcium mobilization from the endoplasmic reticulum which induces insulin release .
So that the ability of HFD to enhance glucose stimulated insulin secretion may be attributed to its modulatory effects on Ucn3, SOX6 or GPR40.
The results obtained from the in vitro study may appear contradictory to the histopathological results. Especially, when we know that cytokines (produced during inflammation) act directly on beta cells causing reduction in insulin secretion . Our interpretation for this contradiction is that HFD has a direct and indirect effect on pancreatic beta cells. The direct effect is by increasing GSIS. The indirect effect is by inducing a state of low grade inflammation that decreases insulin secretion. It appears that at this stage the direct effect is more potent than the indirect effect. However, after a certain period of time, this condition is reversed leading to type 2 diabetes.
Great thanks for all members of the pharmacology department, faculty of pharmacy, Zagazig university for scientific assistance and support. This research received no specific grant from any funding agency in the public, commercial or not-for-profit sectors.
Conflict of interest
There are no conflicts of interest.