Insulin Resistance In Rats Research Articles

Age-dependent membrane release and degradation of full-length glycosylphosphatidylinositol-anchored proteins in rats

Glycosylphosphatidylinositol (GPI)-anchored proteins (GPI-APs) are associated with the surface of eucaryotic cells only through a covalently coupled carboxy-terminal GPI glycolipid structure which is anchored at the outer leaflet of plasma membranes. This mode of membrane association may be responsible for the recent observations that full-length GPI-APs harbouring the complete GPI anchor are (i) released from isolated rat adipocytes in vitro and (ii) expressed in rat and human serum. The upregulation of the adipocyte release in response to increased cell size and blood glucose/insulin levels of the donor rats and downregulation of the expression in serum of insulin resistant and diabetic rats have been reconciled with enhanced degradation of the full-length GPI-APs released into micelle-like complexes together with (lyso) phospholipids and cholesterol by serum GPI-specific phospholipase D (GPI-PLD).Here by using a sensitive and reliable sensing method for full-length GPI-APs, which relies on surface acoustic waves propagating over microfluidic chips, the upregulation of (i) the release of the full-length GPI-APs CD73, alkaline phosphatase and CD55 from isolated adipocyte plasma membranes monitored in a “lab-on-the-chip” configuration, (ii) their release from isolated rat adipocytes into the incubation medium and (iii) the lipolytic cleavage of their GPI anchors in serum was demonstrated to increase with age (3–16 weeks) and body weight (87−477 g) of (healthy) donor rats. In contrast, the amount of full-length GPI-APs in rat serum, as determined by chip-based sensing, turned out to decline with age/body weight. These correlations suggest that age-/weight-induced alterations (in certain biophysical/biochemical characteristics) of plasma membranes are responsible for the release of full-length GPI-APs which becomes counteracted by elevated GPI-PLD activity in serum. Thus, sensitive and specific measurement of these GPI-AP-relevant parameters may be useful for monitoring of age-related cell surface changes, in general, and diseases, in particular.

Exercise improves high-fat diet- and ovariectomy-induced insulin resistance in rats with altered hepatic fat regulation.

A high-fat diet (HFD) and loss of endogenous estrogens increases the risk for type 2 diabetes (T2D) and insulin resistance. Although exercise is known to prevent and manage insulin resistance, the cellular mechanisms remain largely unknown, especially in the context of a combined HFD and endogenous estrogen loss via ovariectomy (OVX). This study uses female Wistar rats to assess the effect of diet, endogenous estrogens, an exercise on insulin resistance, serum hormones, hepatic AMPK, hepatic regulators of fat metabolism, and expression of signaling molecules of the brain reward pathway. The combination of the HFD/OVX increased the homeostatic model assessment of insulin resistance (HOMA-IR), the glucose-insulin (G-I) index, and the serum adiponectin and leptin values, and exercise decreased these factors. The combination of the HFD/OVX decreased hepatic pAMPK, and exercise restored hepatic pAMPK, an important regulator of fat and glucose metabolism. Furthermore, consumption of the HFD by rats with intact ovaries (and endogenous estrogens) did not result in these drastic changes compared to intact rats fed a standard diet, suggesting that the presence of estrogens provides whole body benefits. Additionally, the HFD decreased the hepatic protein expression of acetyl CoA carboxylase (ACC) and fatty acid synthase (FAS), two proteins involved in de novo lipid synthesis and increased the hepatic protein expression of lipoprotein lipase (LPL), a protein involved in fat storage. Finally, exercise increased mRNA expression of the dopamine D2 receptor and tyrosine hydroxylase in the dopaminergic neuron cell body region of the ventral tegmental area, which is a key component of the brain reward pathway. Overall, this study demonstrates that exercise prevents insulin resistance even when a HFD is combined with OVX, despite hepatic changes in ACC, FAS, and LPL.

Apple pomace and rosemary extract ameliorates hepatic steatosis in fructose-fed rats: Association with enhancing fatty acid oxidation and suppressing inflammation.

Apple pomace and rosemary (AR) have been reported to contain rich bioactive molecules, which have numerous metabolic effects. Our preliminary work revealed that AR ameliorated fructose-induced insulin resistance in rats by modulating sarcolemmal CD36 and glucose transporter-4. The present study aimed to further examine how AR improves metabolic disorders by investigating the effect of AR on hepatic steatosis induced by fructose overconsumption. The results demonstrated that AR (100 mg/kg daily by gavage for 5 weeks) attenuated chronic liquid fructose consumption-induced increases in liver triglyceride content in rats. Mechanistically, reverse transcription-quantitative PCR and western blot analysis results indicated that AR reversed fructose-induced suppression of hepatic peroxisome proliferator-activated receptor α, carnitine palmitoyl-transferase 1α, sirtuin 1 and peroxisome proliferator-activated receptor-γ coactivator 1α, which were associated with the fatty acid oxidative (FAO) pathway. In addition, AR treatment decreased the expression levels of the pro-inflammatory proteins NF-κB and tumor necrosis factor-α. However, AR had no effect on the genes related to lipogenesis and the very low-density lipoprotein-export pathway in rat liver. Thus, the present results suggested that AR treatment diminished long-term fructose overconsumption-induced fatty liver, which was associated with enhanced FAO and suppressed inflammation.

Establishment of high-fat diet-induced obesity and insulin resistance model in rats

To investigate the appropriate conditions and duration for establishing a high-fat diet-induced obesity and insulin resistance model in rats. Forty-five 6-week-old male Sprague-Dawley (SD) rats were randomly assigned into 2 groups: (1) control group (CON), (2) high-fat diet group (HFD). HFD was fed with a high-fat diet (45% kcal from fat) while CON with chow diet. After four-weeks of high-fat diet feeding, the rats of obesity resistance (OR) were eliminated according to body weight sorting, whereas obese (OB) rats were continued feeding a high-fat diet until 12 weeks. Body weight and food intake were recorded weekly. Glucose tolerance was evaluated by oral glucose tolerance test (OGTT) in 4 weeks, 8 weeks and 12 weeks. At the end of 12 weeks, insulin releasing test and visceral fat mass were measured and HE staining of the liver, adipose tissue and pancreatic tissue were conducted. After 4 weeks of a high-fat diet, the body weight of HFD was 17.8% higher than that of CON (P=0.001), and the rate of obesity was 67.6%-78.4%. Glucose tolerance of OB rats was impaired with a higher blood glucose concentration at 120 min (P<0.001) and a higher area under the curve (AUC, P=0.037) in OGTT compared with CON. The rate of obesity and insulin-resistance rats was 79.3%. After 8 weeks of feeding, the body weight in OB was 30.4% higher than CON (P<0.001). In OGTT, blood glucose levels at 60 min and 120 min were 35.6% and 36.4% higher than those in CON (both P<0.001), and AUC was 21.7% (P<0.001) higher than that of CON. The rate of obesity and insulin-resistance rats was 100.0%. After 12 weeks of feeding, the body weight in OB was 36.9% higher than that in CON (P<0.001). In OGTT, the blood glucose levels at 60 min and 120 min were 24.8% (P=0.001) and 34.6% (P<0.001) higher than those in CON, and AUC was 16.1% (P=0.019) higher than that of CON. The rate of obesity and insulin-resistance rats was 93.3%. The insulin releasing test showed that serum insulin concentration at each time point (0, 30, 60, 120 min) was higher than that in CON, with a 6.3-times higher than that in CON at 120 min (P=0.008). Pathological changes were observed in islets and liver in the OB rats. After 4 weeks of a high-fat diet (45% kcal from fat) feeding in six-weeks SD rats, the rats of OR were eliminated. Impaired glucose tolerance was found in OB rats after 4 weeks of feeding, and the rate was higher after 8-12 weeks of high-fat diet feeding.

Effects of Fish Oil and Grape Seed Extract Combination on Hepatic Endogenous Antioxidants and Bioactive Lipids in Diet-Induced Early Stages of Insulin Resistance in Rats.

Diacylglycerols (DAG) and ceramides have been suggested as early predictors of insulin resistance. This study was aimed to examine the combined effects of fish oil (FO) and grape seed extract (GSE) on hepatic endogenous antioxidants, DAG and ceramides in diet-induced early stages of insulin resistance. Thirty-five rats were fed one of the following diets: (1) a standard diet (STD group), (2) a high-fat high-sucrose diet (HFHS group), (3) an HFHS diet enriched with FO (FO group), (4) an HFHS diet enriched with GSE (GSE group) or (5) an HFHS diet enriched with FO and GSE (FO + GSE group). In the liver, endogenous antioxidants were measured using spectrophotometric and fluorometric techniques, and non-targeted lipidomics was conducted for the assessment of DAG and ceramides. After 24 weeks, the FO + GSE group showed increased glutathione peroxidase activity, as well as monounsaturated fatty acid and polyunsaturated fatty acid-containing DAG, and long-chain fatty acid-containing ceramides abundances compared to the STD group. The FO and GSE combination induced similar activation of the antioxidant system and bioactive lipid accumulation in the liver than the HFHS diet without supplementation. In addition, the FO and GSE combination increased the abundances of polyunsaturated fatty acid-containing DAG in the liver.

Mitochondrial Utilization of Competing Fuels Is Altered in Insulin Resistant Skeletal Muscle of Non-obese Rats (Goto-Kakizaki).

AimInsulin-resistant skeletal muscle is characterized by metabolic inflexibility with associated alterations in substrate selection, mediated by peroxisome-proliferator activated receptor δ (PPARδ). Although it is established that PPARδ contributes to the alteration of energy metabolism, it is not clear whether it plays a role in mitochondrial fuel competition. While nutrient overload may impair metabolic flexibility by fuel congestion within mitochondria, in absence of obesity defects at a mitochondrial level have not yet been excluded. We sought to determine whether reduced PPARδ content in insulin-resistant rat skeletal muscle of a non-obese rat model of T2DM (Goto-Kakizaki, GK) ameliorate the inhibitory effect of fatty acid (i.e., palmitoylcarnitine) on mitochondrial carbohydrate oxidization (i.e., pyruvate) in muscle fibers.MethodsBioenergetic function was characterized in oxidative soleus (S) and glycolytic white gastrocnemius (WG) muscles with measurement of respiration rates in permeabilized fibers in the presence of complex I, II, IV, and fatty acid substrates. Mitochondrial content was measured by citrate synthase (CS) and succinate dehydrogenase activity (SDH). Western blot was used to determine protein expression of PPARδ, PDK isoform 2 and 4.ResultsCS and SDH activity, key markers of mitochondrial content, were reduced by ∼10–30% in diabetic vs. control, and the effect was evident in both oxidative and glycolytic muscles. PPARδ (p < 0.01), PDK2 (p < 0.01), and PDK4 (p = 0.06) protein content was reduced in GK animals compared to Wistar rats (N = 6 per group). Ex vivo respiration rates in permeabilized muscle fibers determined in the presence of complex I, II, IV, and fatty acid substrates, suggested unaltered mitochondrial bioenergetic function in T2DM muscle. Respiration in the presence of pyruvate was higher compared to palmitoylcarnitine in both animal groups and fiber types. Moreover, respiration rates in the presence of both palmitoylcarnitine and pyruvate were reduced by 25 ± 6% (S), 37 ± 6% (WG) and 63 ± 6% (S), 57 ± 8% (WG) compared to pyruvate for both controls and GK, respectively. The inhibitory effect of palmitoylcarnitine on respiration was significantly greater in GK than controls (p < 10–3).ConclusionWith competing fuels, the presence of fatty acids diminishes mitochondria ability to utilize carbohydrate derived substrates in insulin-resistant muscle despite reduced PPARδ content.

Partial Body Mass Recovery After Caloric Restriction Abolishes Improved Glucose Tolerance in Obese, Insulin Resistant Rats.

Caloric restriction, among other behavioral interventions, has demonstrated benefits on improving glycemic control in obesity-associated diabetic subjects. However, an acute and severe intervention without proper maintenance could reverse the initial benefits, with additional metabolic derangements. To assess the effects of an acute caloric restriction in a metabolic syndrome model, a cohort of 15-week old Long Evans Tokushima Otsuka (LETO) and Otsuka Long Evans Tokushima Fatty (OLETF) rats were calorie restricted (CR: 50% × 10 days) with or without a 10-day body mass (BM) recovery period, along with their respective ad libitum controls. An oral glucose tolerance test (oGTT) was performed after CR and BM recovery. Both strains had higher rates of mass gain during recovery vs. ad lib controls; however, the regain was partial (ca. 50% of ad lib controls) over the measurement period. Retroperitoneal and epididymal adipose masses decreased 30% (8.8 g, P < 0.001) in OLETF; however, this loss only accounted for 11.5% of the total BM loss. CR decreased blood glucose AUC 16% in LETO and 19% in OLETF, without significant decreases in insulin. Following CR, hepatic expression of the gluconeogenic enzyme, PEPCK, was reduced 55% in OLETF compared to LETO, and plasma triglycerides (TG) decreased 86%. Acute CR induced improvements in glucose tolerance and TG suggestive of improvements in metabolism; however, partial recovery of BM following CR abolished the improvement in glucose tolerance. The present study highlights the importance of proper maintenance of BM after CR as only partial recovery of the lost BM reversed benefits of the initial mass loss.

Effect of piridoxin on the skin reparation process under experimental insulin resistance

Topicality. According to WHO experts estimation, the worldwide prevalence of diabetes mellitus is pandemic, with more than 80 % of patients dying from the fatal effects of diabetic angiopathy. One of the most common complications of diabetes of both types is the formation of chronic wound defects, and eventually gangrene of the lower limbs with the transformation into sepsis, which cases to 3.2 % of deaths against the background of diabetes.Aim. To study the effect of combined medicine magnesium and pyridoxine on the course of the reparative skin process in rats with experimental insulin resistance.Materials and methods. The study has been carried out in 60 rats males weighing 180-220 g. Experimental insulin resistance in rats was induced under conditions of the hypercaloric diet and intraperitoneal administration of dexamethasone. Studies of the process of skin repair have been performed on models of full-layer stencil and linear cut wounds, as well as determination of carbohydrate metabolism markers and glycosylation products in serum and hemolysate.Results and discussion. The use of pyridoxine in combination with a reparant (retinol acetate) resulted in a significant reduction in wound area as early as 5 day of treatment and effectively reduced the total time of complete epithelialization; and also increased the load under tensiometry, which corresponded to 72.8 % of the reparative activity of this therapeutic scheme. Against the background of the pyridoxine the content of fructosamine and glycosylated hemoglobin also has been significantly reduced, which confirms the antiglycosylative properties of vitamin.Conclusions. Pyridoxine is able to potentiate the wound healing effect of reparants under the insulin resistance due to its antiglycosylation properties. The results of this study allow us to recommend further research of pyridoxine in combination with standard skin repairs in the treatment of wounds in patients with hyperglycemia and diabetes mellitus.

Unveiling the presence of epigenetic mark by Lactobacillus supplementation in high-fat diet-induced metabolic disorder in Sprague-Dawley rats

Gut dysbiosis, particularly bacteria from Firmicutes and Bacteroidetes phyla, plays a fundamental role in the progression of metabolic disorders. Probiotics have shown to restore the gut microbiota composition in metabolic disorders with subsequent beneficial effects. Recent studies have reported that several species of Lactobacillus as probiotic supplementation improve insulin sensitivity and glucose metabolism. Nonetheless, whether Lactobacillus could influence the epigenetic modifications that underlie insulin-resistant conditions is still unexplored. Therefore, the current study examined the therapeutic effects and underlying epigenetic mechanisms of three different species of Lactobacillus in the high-fat diet (HFD)-induced insulin-resistant rats. Three different species of Lactobacillus; Lactobacillus casei, Lactobacillus gasseri, and Lactobacillus rhamnosus were individually supplemented orally (109 CFU/mL) to insulin-resistant SD rats for 12 weeks. Lactobacillus supplementation led to a significant reduction in the hyperglycemia, hyperinsulinemia, and hyperlipidemia associated with HFD-induced insulin resistance. Histopathological examination also indicated the protective effects of Lactobacillus supplementation against the hepatic and intestinal damage caused by the high-fat diet. Lactobacillus supplementation also down-regulated the expression of FOXO1, a major transcription factor of insulin signaling. In addition, at the epigenetic level, Lactobacillus supplementation predominantly prevented methylation and demethylation of H3K79me2 and H3K27me3, respectively. Chromatin Immunoprecipitation (ChIP) coupled with quantitative PCR (ChIP-qPCR) assay revealed the presence of cross-talk between these two histone modifications at the promoter region of FOXO1. Taken together, this is the first report to observe that the effects of Lactobacillus supplementation involve alteration in FOXO1 expression via cross-talking between H3K79me2 and H3K27me3 histone modifications.

Effect of different fractions of chia (Salvia hispanica L.) on glucose metabolism, in vivo and in vitro

• Chia oil recovered glycolysis enzymes, glucose tolerance and AKT phosphorylation. • Chia oil and flour decreased adiposity and increased AMPK gene expression. • Chia hydrolyzed phenolics did not improve AKT phosphorylation of protein in HepG2 cells. • Chia hydrolyzed phenolics decreased glycolytic and gluconeogenic enzymes in vitro. The influence of chia ( Salvia hispanica L.) flour and oil on glucose metabolism (GM) in insulin resistant (IR) Wistar rats and the effect of chia hydrolyzed phenolics extract (CHPE) on GM in IR HepG2 cells were evaluated. In vivo study: animals were divided into four groups: AIN-93M, high-fat and high-fructose (HFHF), HFHF with chia flour (14.7%) or chia oil (4%). In vitro study: IR HepG2 cells were treated with CHPE (80 ppm). In vivo , chia flour and oil reduced adiposity and increased AMPK mRNA. Chia oil improved glucose tolerance, increased AKT1[pS473] protein level, mRNA of insulin receptor, FOXO1 and glycolysis enzymes. In vitro , CHPE decreased gluconeogenesis enzymes mRNA. Chia flour and oil decreased adiposity, but only chia oil was able to improve glucose tolerance and restore energy fuel system in liver of rats fed HFHF diet. CHPE decreased mRNA levels of gluconeogenesis enzymes.

Serum metabolomics analysis of the intervention effect of whole grain oats on insulin resistance induced by high-fat diet in rats

This study aimed to evaluate the intervention effect of whole grain oats (WGO) on a high-fat (HF) diet induced insulin resistance (IR) rat model by metabolomic approach. Sixty rats were randomly allocated into six groups, including the control group, HF group, and four oat-intervention groups. Rats in the control group and HF group were respectively given the standard diet and HF diet respectively. The four oat-intervention groups were fed with a special HF diet, in which sucrose and starch in the HF diet were replaced by whole grain oats powder in different proportions of 25%, 50%, 75%, and 100% respectively. After 8-week feeding, serum samples of all rats were collected for biochemical detection, and the metabolomics analysis was carried out by ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS). The results showed that the WGO could regulate body weight, blood glucose, blood lipid, and inflammatory response, which indicated that WGO could prevent IR induced by the HF diet, and it was effective enough by 50% WGO replacement. By serum metabolomics analysis, six differential metabolites closely related to WGO intervention on IR were found, and the related metabolic pathways involved energy metabolism, amino acid metabolism, and lipid metabolism.

SUN-347 Glucagon-like Peptide 1 (GLP-1) Acts Directly On Human Osteoclasts To Increase Differentiation And Bone Resorptive Activity

Glucagon-like peptide 1 (GLP-1) is an intestinal hormone released in response to nutrient intake that promotes glucose-dependent insulin secretion by acting upon the pancreatic GLP-1 receptor (GLP-1R). GLP-1R agonists (GLP-1RAs) are widely used in treatment of type 2 diabetes. Preclinical data indicate that GLP-1RAs could be repurposed to treat low bone mass as GLP-1R-depleted mice have higher bone resorption and thinner cortical bones, while insulinopenic and insulin resistant rats have improved bone formation and reduced bone mass deterioration when treated with GLP-1 or GLP-1RAs. However, the effect of GLP-1 and GLP-1RAs on human bone cells remains undetermined. We aimed to elucidate the effect of GLP-1 on primary human osteoclast (OC) and osteoblast (OB) cultures. OCs were differentiated over 10 days from human blood-derived CD14+ monocytes and OBs over 4–6 weeks from human bone. Cells were seeded on bovine bone slices and studies performed using fetal bovine serum, MCSF and RANKL (OC monocultures) or MCSF only (OB monocultures and OB+OC co-cultures). We first investigated the effect of GLP-1 on bone resorptive activity of mature OCs on bovine bone slices. GLP-1 increased the eroded bone surface percentage compared to vehicle in both OC monocultures (1nM P=0.002; 10nM P=0.023; n=8 donors) and OC+OB co-cultures (1nM P=0.013; 10nM P=0.012; n=8 donors). We then tested the effects of GLP-1 on osteoblast activity in OC+OB co-cultures by measuring alkaline phosphatase (ALP). We found that GLP-1 increased ALP in OC+OB cultures (1nM, P=0.049; 10nM, P=0.019) and these effects were reversed by the GLP-1R antagonist exendin 9–39 (1nM, P=0.93, 10nM, P=0.64). However, in OB monocultures GLP-1 had no effects on ALP (1nM P=0.93, 10nM P=0.64) indicating a GLP-1-driven increase in osteoblast activity through osteoclast-osteoblast coupling. We then assessed the effect of GLP-1 on OC differentiation by assessing TRAcP activity. Although there was a trend towards increased TRAcP activity upon stimulation with GLP-1 on day 10 of osteoclastogenesis, this was not statistically significant (1nM P=0.12; n=8 donors; 10nM P=0.29, n=4 donors). Our studies indicated GLP-1 may have a direct effect on osteoclasts, and we therefore sought to characterise GLP-1-mediated signalling in these cells. We assessed the effect of GLP-1 on cAMP signalling using LANCE assays and assessed phosphorylation of ERK proteins by Western blot analysis in human OC cultures. OCs treated with 10nM GLP-1 for 30 minutes had increased cAMP signaling (P=0.004, n=12 bone slices from 2 donors) when compared to vehicle. Furthermore, 10nM GLP-1 induced rapid increases in phosphorylated ERK (P=0.03 following 2 minutes exposure, n=4 blots). In conclusion, our studies reveal that GLP-1 increases activity in primary mature human OCs, and OBs, via OCs. Our signaling studies in OCs indicate this is mediated by direct action of GLP-1 on human bone cells.

Titin and CK2α are New Intracellular Targets in Acute Insulin Application-Associated Benefits on Electrophysiological Parameters of Left Ventricular Cardiomyocytes From Insulin-Resistant Metabolic Syndrome Rats.

Previous studies have demonstrated that a high-carbohydrate intake could induce metabolic syndrome (MetS) in male rats with marked cardiac functional abnormalities. In addition, studies mentioned some benefits of insulin application on these complications, but there are considerable disagreements among their findings. Therefore, we aimed to extend our knowledge on the in-vitro influence of insulin on left ventricular dysfunction and also in the isolated cardiomyocytes from MetS rats. At the organ function level, an acute insulin application (100-nM) provided an important beneficial effect on the left ventricular developed pressure in MetS rats. Furthermore, to treat the freshly isolated cardiomyocytes from MetS rats with insulin provided marked recoveries in elevated resting intracellular Ca2+-level, as well as significant prevention of prolonged action potential through an augmentation in depressed K+-channel currents. Insulin also normalized the cellular levels of increased ROS and phosphorylation of PKCα, together with normalizations of apoptotic markers in MetS cardiomyocytes through the insulin-mediated regulation of phospho-Akt. Since not only elevated PKCα-activity but also reductions in phospho-Akt are key modulators of titin-based cardiomyocyte stiffening in hyperglycemia, insulin treatment of the cardiomyocytes prevented the activation of titin via the above pathways. Furthermore, CK2α-activation and NOS-phosphorylation could be prevented with insulin treatment. Mechanistically, we found that impaired insulin signaling and elevated PKCα and CK2α activities, as well as depressed Akt phosphorylation, are key modulators of titin-based cardiomyocyte stiffening in MetS rats. We propose that restoring normal kinase activities and also increases in phospho-Akt by insulin can contribute marked recoveries in MetS heart function, indicating a promising approach to modulate titin-associated factors in heart dysfunction associated with type-2 diabetes mellitus. Graphical Abstract.

Deregulation of hepatic lipid metabolism associated with insulin resistance in rats subjected to chronic monocrotophos exposure.

In our previous study, we demonstrated the potential of monocrotophos (MCP), an organophosphorus insecticide (OPI), to induce glucose intolerance, insulin resistance (IR), and dyslipidemia with hyperinsulinemia in rats after chronic exposure. As hyperinsulinemia is likely to exert an impact on hepatic lipid metabolism, we carried out this study to establish the effect of chronic MCP exposure (0.9 and 1.8 mg/kg/day for 180 days) on hepatic lipid metabolism in rats. The state of IR induced by MCP in rats was associated with an increase in the liver lipid content (triglyceride and cholesterol) and expression levels of sterol regulatory element-binding proteins, PPARγ, acetyl-CoA carboxylase, and fatty acid synthase in the liver. Similarly, activities of key enzymes (acetyl-COA carboxylase, fatty acid synthase, lipin 1, malic enzyme, glucose-6-phosphate dehydrogenase, and glycerol-3-phosphate dehydrogenase), which regulate lipogenesis, were enhanced in livers of pesticide-treated rats. A strong correlation was observed between insulin levels, hepatic lipid content, and plasma lipid profile in treated rats. Our study suggests that long-term exposure to OPIs not only has a propensity to induce a state of hyperinsulinemic IR, but it is also associated with augmented hepatic lipogenesis, which may explain dyslipidemia induced by chronic exposure to MCP.

An investigation of acute effects at various doses of malathion on glucose homeostasis and insulin resistance in rat liver, pancreas and serum

Objective. This study investigates acute effects of various doses of Malathion on glucose homeostasis and insulin resistance in rat. Methods. Rats were randomly divided into four groups of 6 animals each. Corn oil was given orally to Group 1. Group 2, Group 3, and Group 4 received malathion dissolved in corn oil via oral administration at the doses of 100, 200 and 400 mg/kg, respectively. 24 hours later the rats were sacrificed. Results. Acute administration of Malathion led to a decrease in serum butryl cholinesterase (BChE) levels at all doses tested. It also caused a significant increase in serum advanced glycation end products (AGEs), insulin, and TNF-α levels at all doses. Moreover, Malathion administration raised the liver ALT, AST and LDH, TNF-α, and glycogen levels in a dose dependent manner. It also led to a remarkable increase in pancreatic insulin levels at all doses. Conclusions. Acute administrations of Malathion affect glucose homeostasis in a dose dependent manner through its effects on the liver, serum, and pancreas.

Effects of dapagliflozin and statins attenuate renal injury and liver steatosis in high-fat/high-fructose diet-induced insulin resistant rats

High-fat high-fructose diet (HFF) in obesity can induce dyslipidemia and lipid accumulation both in kidney and liver which related to insulin resistance and lipotoxicity-induced cellular damage. We investigated whether dapagliflozin with or without atorvastatin could improve lipid accumulation-induced kidney and liver injury in HFF-induced insulin resistant rats. Male Wistar rats were fed with HFF for 16 weeks and then received drug treatments for 4 weeks; vehicle, dapagliflozin, atorvastatin and dapagliflozin plus atorvastatin treatment groups. HFF rats demonstrated insulin resistance, dyslipidemia, liver injury and renal dysfunction associated with impaired renal lipid metabolism and lipid accumulation. Dapagliflozin and combination treatment could improve HFF-induced insulin resistance, lipogenesis and lipotoxicity-related renal oxidative stress, inflammation, fibrosis and apoptosis leading to kidney dysfunction recovery. Liver injury-associated inflammation was also improved by these two regimens. Notably, the reduced lipid accumulation in liver and kidney that linked to an improvement of lipid oxidation was prominent in the combination treatment. Therefore, dapagliflozin combined with atorvastatin treatment exert the beneficial effects on lipid metabolism and lipotoxicity in liver and kidney injury via the attenuation of oxidative stress, fibrosis and apoptosis in insulin resistant model.

Body mass recovery after visceral lipectomy improves glucose tolerance without adipose recovery in obese, insulin resistant rats.

Increased visceral fat is associated with metabolic dysregulation, which may be a consequence of chronic exposure of the liver to adipokines and fatty acids derived from visceral adipose, increased hepatic glucose output, and insulin resistance. Therefore, lipectomy (rapid removal of adipose) may ameliorate insulin resistance and other associated maladies. However, abdominal lipectomy did not significantly improve insulin resistance in obese, non‐diabetic females studied one year after their surgery. Moreover, in Sprague‐Dawley rats, fasting glucose and plasma leptin and insulin increased and lipogenesis was stimulated 10 days after lipectomy suggesting that maladaptation may be a consequence of lipectomy. We have previously demonstrated that 50% caloric restriction for 10 days (gradual reduction in adiposity) decreased visceral fat 2.1% associated with an improvement in insulin resistance index and triglyceridemia in OLETF rats. Our goal was to assess the effects of visceral lipectomy (rapid loss) with fat removal comparable to the adipose lost (gradual) in our previous study, with and without mass recovery, in a model of metabolic syndrome.Sixteen lean LETO and 32 obese, insulin resistant OLETF rats were weighted daily (Fig. 1) and fed ad libidum until 18 weeks of age. OLETF rats (n=18) were subjected to removal of retroperitoneal and epididymal fat equivalent to 2 ± 0.1% of their total body mass, while the rest of the rats were subjected to sham surgery. Half of sham LETO and OLETF (Lpx Ctrl), and half of the lipectomized rats (OLETF Lpx) were subjected to oGTT two days after surgery and dissected 3 days later. The rest of the groups (LETO, OLETF Rec Ctrl, and OLETF Rec), were subjected to oGTT 10 days after surgery and dissected 3 days later.Mean glucose AUC (mg*h/dL) was 2‐fold higher in OLETF vs LETO at 18 w.o. (669 ± 39 vs 332 ± 9, P=0.428). Glucose AUC did not change in OLETF Lpx compared to control (726 ± 73 vs 669 ± 39, P=0.428), however it decreased 28% after mass recovery (408 ± 7 vs 570 ± 52, P=0.044) (Fig. 2). Visceral (epididimal + retroperitoneal) relative mass did not increase with age in OLETF (6.8 ± 0.2% vs 6.2 ± 0.2% at 19 and 20 w.o, respectively). Moreover, relative mass decreased in OLETF Rec compared to OLETF Lpx (3.7 ± 0.5 vs 5.0 ± 0.2%, P&lt;0.001) despite an increase in total body mass (Lpx: 508 ± 12 vs Rec: 538 ± 24 g). We conclude that glucose tolerance was improved after body mass recovery without recovery of the retroperitoneal or epididymal depots, reaffirming the role of visceral fat in insulin resistance.Support or Funding InformationThis research was supported in part by the University of California Institute for Mexico and the United States and Mexico’s National Council for Science and Technology doctoral fellowship UC MEXUS‐CONACYT 440553 and by National Institute on Minority Health and Health Disparities grant 9T37‐MD001480Mean ± SE body mass (g). After surgery, mass was not recorded until the day after oGTT, two days before dissection.Figure 1Mean ± SE glucose AUC (mg*h/dl). aP&lt;0.05 vs LETO, bP&lt;0.05 vs Ctrl, cP&lt;0.05 vs LpxFigure 2

Combination of dapagliflozin and statins attenuate renal lipotoxicity in high‐fat high‐fructose diet‐induced insulin resistant rats

High‐fat high‐fructose diet (HFF) can induce dyslipidemia and lipid accumulation in the kidney which are related to insulin resistance and metabolic syndrome. Dyslipidemia and lipid accumulation in obesity can lead to lipotoxicity and cellular damage particularly kidney injury by activating oxidative stress, fibrotic and apoptotic pathways. This study investigated whether dapagliflozin alone or combined with atorvastatin could improve metabolic parameters, lipid metabolism and lipid accumulation induced kidney injury in HFF‐induced insulin resistant rats. Male Wistar rats were fed with high‐fat diet with 10% fructose in drinking water for 16 weeks. After that, these rats were received drug treatments for 4 weeks; vehicle (HFF), SGLT2 inhibitor dapagliflozin (HFFD), atorvastatin (HFFA) and SGLT2 inhibitor combined with atorvastatin (HFFDA) groups. HFF rats demonstrated insulin resistance, dyslipidemia, and renal dysfunction associated with impaired renal lipid metabolism and lipid accumulation. Lipid accumulation in the kidney could promote renal dysfunction in HFF rats via the activation of oxidative stress, fibrosis and apoptosis. Dapagliflozin treatment could improve insulin resistance, lipid synthesis and lipotoxicity‐related renal oxidative stress, fibrosis and apoptosis leading to reversal of kidney dysfunction in HFF model. The combined treatment (HFFDA group) could also improve metabolic parameters which was not different from single drug treatment (HFFD or HFFA group). Notably, the reduced lipid accumulation in kidney which were linked to the improvement in lipid oxidation was better in the combined treatment group when compared to a single drug treatment. In summary, dapagliflozin combined with atorvastatin treatment exert the beneficial effects on metabolic parameters, lipid metabolism, and lipid accumulation induced kidney injury via the attenuation of oxidative stress, fibrosis and apoptosis in high‐fat high‐fructose diet‐induced insulin resistant model.Support or Funding InformationThailand Research Fund (RSA6080015 AL); the Royal Golden Jubilee PhD program (PhD/0063/2560 LT and AL); the Faculty of Medicine Research Fund, Chiang Mai University (AL and AP); the Graduate Research Scholarship Chiang Mai University (LT); and the Functional Food Research Center for Well‐being, Chiang Mai University (AL).

Chronic oral NaHCO3 treatment and high tubular flow promote insulin resistance in rats

Diabetes is the leading cause of chronic kidney disease (CKD), and worsening insulin resistance likely contributes to CKD progression. Several small clinical trials have indicated that sodium bicarbonate (NaHCO3) slows the decline in GFR in CKD. Chronic ingestion of alkali like NaHCO3 can lead to potassium (K+) wasting by the kidney. While K+ wasting has been linked to the dysregulation of insulin metabolism, no studies have examined the metabolic impacts of chronic NaHCO3. We hypothesized that chronic NaHCO3 would promote K+ wasting and worsen insulin resistance in rats. In order to test our hypothesis, 16 male, 12 wk old Sprague Dawley (SD) rats underwent four weeks of high‐fructose (60%) diet (HFruD) prior to a baseline insulin tolerance test (ITT). Treatment with 0.1M NaHCO3 (Bicarb; n=8) or 0.1M NaCl (Vehicle; n=8) was initiated in drinking water after the baseline ITT and continued along with fructose feeding for another 3 weeks. ITTs were performed again following 1 week and 3 weeks of either bicarb or vehicle drinking. A similar protocol was performed using 18 male, 12 wk old SD rats, with the exception that these animals were maintained on normal rat chow, and following a baseline ITT, were placed on either Bicarb (n=6), Vehicle (n=6), or Tap H2O (n=6). At the conclusion of the protocol, animals were sacrificed, and visceral and subcutaneous fat pads and the spleen and kidneys harvested. In HFruD rats, there were no significant differences in the magnitude of the fall in blood glucose (area under the curve (AUC)) in response to insulin (0.75U/kg) at baseline (Veh 4229 ± 1625 vs Bicarb 3624 ± 1093; unpaired t‐test, p=0.76) or following 1 week of treatment (Veh 7187 ± 1613 vs Bicarb 5151 ± 1121; unpaired t‐test, p=0.32). However, following 3 weeks of treatment, animals that received bicarb demonstrated a significant decrease in the response to insulin, when compared to vehicle treated animals (Veh 5170 ± 690.0 vs Bicarb 1902 ± 846.0; unpaired t‐test, p=0.001). In naïve animals, there were no significant differences in the response to insulin at baseline (Veh 2337 ± 396.2 vs Bicarb 3651 ± 690.7 vs Tap H2O 3349 ± 544.2; One‐way ANOVA, p=0.25) or following three weeks of treatment (Veh 1773 ± 559.5 vs Bicarb 1745 ± 338.3 vs Tap H2O 1320 ± 215.7; One‐way ANOVA, p=0.67). In conclusion, our data show that prolonged NaHCO3 ingestion can significantly impair the response to insulin. As this effect was only present in HFruD fed rats, we speculate that high tubular flow may act synergistically with NaHCO3 to increase K+ wasting or that underlying metabolic dysfunction may be required to observe impairments in insulin sensitivity. As both underlying metabolic dysfunction and high single tubular flow due to reduced nephron mass are present in many CKD patients, these findings could have significant implications for CKD patients who are currently supplemented with NaHCO3. Such patients may require more vigilant blood glucose monitoring.Support or Funding InformationNIH PO1HL134604 and DK099548

Lack of Uncoupling Protein 3 Protects from High‐Fat Diet‐Induced Obesity, Systemic Inflammation and Insulin Resistance in Rats

Introduction. Uncoupling protein 3 (UCP3), a mitochondrial anion carrier with antioxidant properties, is downregulated in animal models with obesity, insulin resistance and type 2 diabetes. Although mutations in UCP3 have been linked to obesity in humans, their contribution to increased adiposity and insulin resistance is still unclear. The goal of this study was to investigate the impact of UCP3 loss on the development of high‐fat diet‐induced obesity in rats.Methods. Male UCP3 knockout (KO) rats and their wild type (WT) littermates were maintained on a high‐fat diet (45% kcal from fat) for up to 21 weeks starting at 8 weeks of age. Body composition was analyzed by EchoMRI. Whole‐body insulin sensitivity and tissue‐specific glucose uptake were determined during hyperinsulinemic‐euglycemic clamps studies. White and brown adipose tissue morphologies were assessed by histology and electron microscopy. Potential molecular alterations in brown adipose tissue (BAT) physiology were further investigated by RNA sequencing, quantification of total protein S‐sulfenylation, and determination of tissue glutathione levels.Results. Loss of UCP3 unexpectedly decreased body weight gain on a high‐fat diet in spite of increased food intake. The decrease in body weight gain was driven by a decrease in body fat mass leading to smaller white adipocytes and a decrease in circulating tumor necrosis factor alpha levels. Consequently, the KO rats exhibited preserved insulin sensitivity and glucose tolerance which correlated with increased glucose uptake in BAT. While high‐fat feeding induced lipid accumulation and whitening of BAT in WT rats, this phenomenon was markedly reduced in KO rats. Surprisingly, protein S‐sulfenylation was decreased in BAT from KO rats, suggesting decreased mitochondrial reactive oxygen species generation in spite of UCP3 loss. The RNA sequencing analysis of BAT transcriptome revealed increased expression of multiple enzymes linked to glutathione synthesis. Direct quantification of glutathione confirmed a 44% increase of the antioxidant molecule levels in BAT of KO rats when compared to high‐fat fed WT rats.Conclusions. Lack of UCP3 protects from high‐fat diet‐induced obesity and insulin resistance in rats. We propose that this protective effect is driven by a compensatory increase in synthesis of the antioxidant glutathione in BAT, thereby protecting mitochondrial function and substrate utilization in this tissue. The findings suggest that loss of UCP3 function in humans may unexpectedly decrease whole‐body fat accumulation, inflammation and the development of insulin resistance.Support or Funding InformationThis work was supported by grants R01HL136438, P20GM104357 and P01HL051971 from the National Institutes of Health.