Glucose Transporter mRNA Research Articles

High glucose dialysate enhances peritoneal fibrosis through upregulating glucose transporters GLUT1 and SGLT1

Objective: To investigate the role of glucose transporter 1 (GLUT1) and sodium-glucose cotransporter 1 (SGLT1) in high glucose dialysate-induced peritoneal fibrosis. Methods: Thirty six male SD rats were randomly divided into 6 groups (6 in each):normal control group, sham operation group, peritoneal dialysis group (PD group), PD+phloretin group (PD+T group), PD+phlorizin group (PD+Z group), PD+phloretin+phlorizin group (PD+T+Z group). Rat model of uraemia was established using 5/6 nephrotomy, and 2.5% dextrose peritoneal dialysis solution was used in peritoneal dialysis. Peritoneal equilibration test was performed 24 h after dialysis to evaluate transport function of peritoneum in rats; HE staining was used to observe the morphology of peritoneal tissue; and immunohistochemistry was used to detect the expression of GLUT1, SGLT1, TGF-β1 and connective tissue growth factor (CTGF) in peritoneum. Human peritoneal microvascular endothelial cells (HPECs) were divided into 5 groups:normal control group, peritoneal dialysis group (PD group), PD+phloretin group (PD+T group), PD+phlorezin group (PD+Z group), and PD+phloretin+phlorezin group (PD+T+Z group). Real time PCR and Western blotting were used to detect mRNA and protein expressions of GLUT1, SGLT1, TGF-β1, CTGF in peritoneal membrane and HPECs. Results:In vivo, compared with sham operation group, rats in PD group had thickened peritoneum, higher ultrafiltration volume, and the mRNA and protein expressions of GLUT1, SGLT1, CTGF, TGF-β1 were significantly increased (all P<0.05); compared with PD group, thickened peritoneum was attenuated, and the mRNA and protein expressions of GLUT1, SGLT1, CTGF, TGF-β1 were significantly decreased in PD+T, PD+Z and PD+T+Z groups (all P<0.05). Pearson's correlation analysis showed that the expressions of GLUT1, SGLT1 in peritoneum were positively correlated with the expressions of TGF-β1 and CTGF (all P<0.05). In vitro, the mRNA and protein expressions of GLUT1, SGLT1, TGF-β1, CTGF were significantly increased in HPECs of peritoneal dialysis group (all P<0.05), and those in PD+T, PD+Z, and PD+T+Z groups were decreased (all P<0.05). Pearson's correlation analysis showed that the expressions of GLUT1, SGLT1 in HPECs were positively correlated with the expressions of TGF-β1 and CTGF (all P<0.05). Conclusion: High glucose peritoneal dialysis fluid may promote peritoneal fibrosis by upregulating the expressions of GLUT1 and SGLT1.

Central injection of GALR1 agonist M617 attenuates diabetic rat skeletal muscle insulin resistance through the Akt/AS160/GLUT4 pathway

Insulin resistance of skeletal muscle plays an important role in the pathogenesis of type 2 diabetes. Galanin, a 29/30-amino-acid neuropeptide, plays multiple biological actions, including anti-diabetic effects. Although recent results of our study showed that administration of galanin could mitigate insulin resistance by promoting glucose transporter 4 (GLUT4) expression and translocation in skeletal muscle of rats, there is no literature available regarding to the effect of type 1 of galanin receptors (GALR1) on insulin resistance in skeletal muscle of type 2 diabetic rats. Herein, we intended to survey the central effect of GALR1 agonist M617 on insulin resistance in skeletal muscle and its underlying mechanisms. We found that the intracerebroventricular injection of M617 increased glucose infusion rates in hyperinsulinemic euglycemic clamp tests, but attenuated the plasma insulin and glucose concentrations of diabetic rats. Furthermore, administration of M617 markedly increased GLUT4 mRNA expression and GLUT4 translocation in skeletal muscle of diabetic rats. Last, perfusion of M617 increased phosphorylated Akt and phosphorylated AS160 levels in the skeletal muscle of diabetic rats. In conclusion, central injection of M617 mitigated insulin resistance of skeletal muscle by enhancing GLUT4 translocation from intracellular pools to plasma membranes via the activation of the Akt/AS160/GLUT4 signaling pathway.

Glycolysis gene expression analysis and selective metabolic advantage in the clinical progression of colorectal cancer.

Production of lactate even in the presence of sufficient levels of oxygen (aerobic glycolysis) seems the prevalent energy metabolism pathway in cancer cells. The analysis of altered expression of effectors causing redirection of glucose metabolism would help to characterize this phenomenon with possible therapeutic implications. We analyzed mRNA expression of the key enzymes involved in aerobic glycolysis in normal mucosa (NM), primary tumor (PT) and liver metastasis (LM) of colorectal cancer (CRC) patients (pts) who underwent primary tumor surgery and liver metastasectomy. Tissues of 48 CRC pts were analyzed by RT-qPCR for mRNA expression of the following genes: hexokinase-1 (HK-1) and 2 (HK-2), embryonic pyruvate kinase (PKM-2), lactate dehydrogenase-A (LDH-A), glucose transporter-1 (GLUT-1), voltage-dependent anion-selective channel protein-1 (VDAC-1). Differences in the expression of the candidate genes between tissues and associations with clinical/pathologic features were studied. GLUT-1, LDH-A, HK-1, PKM-2 and VDAC-1 mRNA expression levels were significantly higher in PT/LM tissues compared with NM. There was a trend for higher expression of these genes in LM compared with PT tissues, but differences were statistically significant for LDH-A expression only. RAS mutation-positive disease was associated with high GLUT-1 mRNA expression levels only. Right-sided colon tumors showed significantly higher GLUT-1, PKM-2 and LDH-A mRNA expression levels. High glycolytic profile was significantly associated with poor prognosis in 20 metastatic, RAS-mutated pts treated with first-line chemotherapy plus Bevacizumab. Altered expression of effectors associated with upregulated glucose uptake and aerobic glycolysis occurs in CRC tissues. Additional analyses are warranted for addressing the role of these changes in anti-angiogenic resistance and for developing novel therapeutics.

Adiponectin plays a role in energy metabolism for musk secretion in scent glands of muskrats (Ondatra zibethicus).

Adult male muskrats (Ondatra zibethicus) secret musk from their scent glands to attract females for seasonal mating. The goal of the present study was to investigate whether the changes in energy metabolism related to musk secretion during the breeding and non-breeding seasons are mediated by adiponectin. We found that the secretion of musk during the breeding season was markedly greater than that during the non-breeding season. The serum adiponectin concentration measured using an ELISA kit was higher during the breeding season than during the non-breeding season. Glandular cells, interstitial cells, epithelial cells and glandular cavities were detected in scent glands using histological methods. Immunohistochemical methods were used to show that AMP-activated protein kinase-gamma-1 (AMPKG1), and glucose transporter 1 (GLUT1) were more strongly expressed in glandular cells during the breeding season than the non-breeding season, whereas the immunoreactivity for acetyl-CoA carboxylase 1 (ACC1) was stronger during the non-breeding season. Consistent with these qualitative results, RNA-Seq analysis indicated that the expression of AdipoR1 mRNA was not significantly different during the two seasons. However, AMPKG1 and GLUT1 mRNA levels were higher in scent glands during the breeding season than during the non-breeding season, whereas ACC1 mRNA levels notably decreased during the breeding season. These results suggest that greater musk secretion requires additional energy, which may be provided by an adiponectin-mediated increase in β-oxidation and glucose absorption.

Influence of hyperuricemia to the expression of IRS-1, IRS-2, and GLUT-4 mRNA in fat and skeletal muscle of male rats

Basic biochemical index were measured weekly in hyperuricemia rats. Real-time quantitative PCR technology was employed to measure glucose transporter-4(GLUT-4), insulin receptor substrate(IRS)-1, and IRS-2 mRNA expression in the skeletal muscle and adipose tissue. The gene expression of GLUT-4 and IRS-2 are significantly lower in the last weeks of the hyperuricemia rat. Insulin resistance caused by hyperuricemia is probably related to the lowering expression levels of GLUT-4 and IRS-2 mRNA in the target tissues. (Chin J Endocrinol Metab, 2015, 31: 902-905) Key words: Hyperuricemia; Insulin receptor substrates; Glucose transporter-4

Osthole improves glucose and lipid metabolism via modulation of PPARα/γ-mediated target gene expression in liver, adipose tissue, and skeletal muscle in fatty liver rats

Context: Osthole may be a dual agonist of peroxisome proliferator-activated receptors (PPAR) α/γ and ameliorate the insulin resistance (IR), but its mechanisms are not yet understood completely.Objective: We investigated the effects of osthole on PPARα/γ-mediated target genes involved in glucose and lipid metabolism in liver, adipose tissue, and skeletal muscle in fatty liver and IR rats.Materials and methods: The rat model was established by orally feeding high-fat and high-sucrose emulsion for 9 weeks. The experimental rats were treated with osthole 5–10 mg/kg by gavage after feeding the emulsion for 6 weeks, and were sacrificed 4 weeks after administration.Results: After treatment with osthole 5–10 mg/kg for 4 weeks, the lipid levels in serum and liver were decreased by 37.9–67.2% and 31.4–38.5% for triglyceride, 33.1–47.5% and 28.5–31.2% for free fatty acid, respectively, the fasting blood glucose, fasting serum insulin, and homeostasis model assessment of IR were also decreased by 17.2–22.7%, 25.9–26.7%, and 37.5–42.8%, respectively. Osthole treatment might simultaneously decrease the sterol regulatory element binding protein-1c, diacylglycerol acyltransferase, and fatty acid synthase mRNA expressions in liver and adipose tissue, and increase the carnitine palmitoyltransferase-1A mRNA expression in liver and glucose transporter-4 mRNA expression in skeletal muscle, especially in the osthole 10 mg/kg group (p < 0.01).Discussion and conclusion: Osthole can improve glucose and lipid metabolism in fatty liver and IR rats, and its mechanisms may be associated with synergic modulation of PPARα/γ-mediated target genes involved in glucose and lipid metabolism in liver, adipose tissue, and skeletal muscle.

Delayed access of low body weight-selected chicks to food at hatch is associated with up-regulated pancreatic glucagon and glucose transporter gene expression

Chickens selected for low (LWS) and high (HWS) juvenile body weight (BW) for 55 generations differ in BW by 10-fold at selection age. High (HWR) and low (LWR) body weight-relaxed lines have been random-bred since the 46th generation. Our objective was to evaluate the developmental and nutritional regulation of pancreatic mRNA abundance of pancreatic and duodenal homeobox 1 (PDX1), preproinsulin (PPI), preproglucagon (PPG), and glucose transporter 2 (GLUT2). At day of hatch (DOH) and days 1, 3, 7, and 15 (D1, 3, 7 and 15, respectively), pancreas was collected and real time PCR was performed in Experiment 1. In Experiment 2, HWS and LWS were fed or delayed access to food for 72h post-hatch, and pancreas collected at D15. There was an interaction of line and age for GLUT2 (P=0.001), PPI (P<0.0001), PPG (P=0.034), and PDX1 (P<0.0001). Expression was greater in chicks from LWR and LWS than HWR and HWS. There was an interaction of line and nutrition on PPG (P<0.0001) and GLUT2 (P=0.001) mRNA, where expression was similar among chicks that were fed but greater in LWS than HWS when chicks were delayed access to food. Thus, the first two weeks is important for maturation of pancreatic endocrine function. Long-term selection for BW is associated with differences in pancreas development, and delaying access to food at hatch may have persisting effects on glucose regulatory function.

Inhibiting GLUT-1 expression and PI3K/Akt signaling using apigenin improves the radiosensitivity of laryngeal carcinoma in vivo.

Hypoxia is an important factor in radioresistance of laryngeal carcinoma. Glucose transporter-1 (GLUT-1) is an important hypoxic marker in malignant tumors, including laryngeal carcinoma. Apigenin is a natural phytoestrogen flavonoid that has potential anticancer effects. Various studies have reported that the effects of apigenin on lowering GLUT-1 expression were involved in downregulation of the PI3K/Akt pathway. Thus, apigenin may improve the radiosensitivity of laryngeal carcinoma by suppressing the expression of GLUT-1 via the PI3K/Akt pathway. The effect of GLUT-1 and PI3K/Akt pathway-related factor expressions by apigenin or antisense oligonucleotides (AS-ODNs) on the radiosensitivity of laryngeal carcinoma in vivo was assessed. The xenograft volume, xenograft weight and apoptosis detection were performed to determine radiosensitivity. The results showed that apigenin or apigenin plus GLUT-1 AS-ODNs improved the radiosensitivity of xenografts. Apigenin or apigenin plus GLUT-1 reduced the expression of GLUT-1, Akt, and PI3K mRNA after X-ray radiation. We found similar results at the protein level. The results suggest that the effects of apigenin on inhibiting xenograft growth and enhancing xenograft radiosensitivity may be associated with suppressing the expression of GLUT-1 via the PI3K/Akt pathway. In addition, apigenin may enhance the effects of GLUT-1 AS-ODNs via the same mechanism.

Metabolic fingerprints in testicular biopsies from type 1 diabetic patients.

Diabetes mellitus (DM) is a metabolic disease that has grown to pandemic proportions. Recent reports have highlighted the effect of DM on male reproductive function. Here, we hypothesize that testicular metabolism is altered in type 1 diabetic (T1D) men seeking fertility treatment. We propose to determine some metabolic fingerprints in testicular biopsies of diabetic patients. For that, testicular tissue from five normal and five type 1 diabetic men was analyzed by high-resolution magic-angle spinning (HR-MAS) nuclear magnetic resonance (NMR) spectroscopy. mRNA and protein expression of glucose transporters and glycolysis-related enzymes were also evaluated. Our results show that testes from diabetic men presented decreased levels of lactate, alanine, citrate and creatine. The mRNA levels of glucose transporter1 (GLUT1) and phosphofructokinase1 (PFK1) were decreased in testes from diabetic men but only GLUT3 presented decreased mRNA and protein levels. Lactate dehydrogenase (LDH) and glutamate pyruvate transaminase (GPT) protein levels were also found to be decreased in testes from diabetic men. Overall, our results show that T1D alters glycolysis-related transporters and enzymes, compromising lactate content in the testes. Moreover, testicular creatine content was severely depressed in T1D men. Since lactate and creatine are essential for germ cells development and support, the data discussed here open new insights into the molecular mechanism by which DM promotes subfertility/infertility in human males.

Ontogenic Changes of Villus Growth, Lactase Activity, and Intestinal Glucose Transporters in Preterm and Term Born Calves with or without Prolonged Colostrum Feeding.

Oral glucose supply is important for neonatal calves to stabilize postnatal plasma glucose concentration. The objective of this study was to investigate ontogenic development of small intestinal growth, lactase activity, and glucose transporter in calves (n = 7 per group) that were born either preterm (PT; delivered by section 9 d before term) or at term (T; spontaneous vaginal delivery) or spontaneously born and fed colostrum for 4 days (TC). Tissue samples from duodenum and proximal, mid, and distal jejunum were taken to measure villus size and crypt depth, protein concentration of mucosa and brush border membrane vesicles (BBMV), total DNA and RNA concentration of mucosa, mRNA expression and activity of lactase, and mRNA expression of sodium-dependent glucose co-transporter-1 (SGLT1) and facilitative glucose transporter 2 (GLUT2) in mucosal tissue. Additionally, protein expression of SGLT1 in BBMV and GLUT2 in crude mucosal membranes and immunochemical localization of GLUT2 in the enterocytes were determined. Villus height in distal jejunum was lower in TC than in T. Crypt depth in all segments was largest and the villus height/crypt depth ratio in jejunum was smallest in TC calves. Concentration of RNA was highest in duodenal mucosa of TC calves, but neither lactase mRNA and activity nor SGLT1 and GLUT2 mRNA and protein expression differed among groups. Localization of GLUT2 in the apical membrane was greater, whereas in the basolateral membrane was lower in TC than in T and PT calves. Our study indicates maturation processes after birth for mucosal growth and trafficking of GLUT2 from the basolateral to the apical membrane. Minor differences of mucosal growth, lactase activity, and intestinal glucose transporters were seen between PT and T calves, pointing at the importance of postnatal maturation and feeding for mucosal growth and GLUT2 trafficking.

Insulin Control of Blood Glucose and GLUT4 Expression in the Skeletal Muscle of Septic Rats.

Insulin resistance is common in septic patients. The level at which the serum glucose should be maintained using insulin infusions for optimal utilization by skeletal muscles is not yet established. The objective of the present study was to compare glucose transporter 4 (GLUT4) mRNA and GLUT4 expression and glucose utilization at the recommended glucose levels of 6-8 mmol/L (110-140 mg/dL) and 8-10 mmol/L (140-180 mg/dL) in septic rats. This was a prospective randomized study using 44 Sprague-Dawley rats (260-330 g). Rats were anaesthetized with gaseous diethyl ether. Catheters were implanted into the jugular vein and artery. Following a laparotomy, rats in the experimental group (n = 36) were rendered septic by standard caecal ligation and puncture (CLP) and intraperitoneal lipopolysaccharide (LPS) infusion (O111:[B4], 1 mg/kg). Control animals (n = 8) underwent laparotomy, but no caecal ligation or puncture and no LPS injection. Four experimental groups were studied: sham-operated control, sepsis treated with fluid maintenance only, sepsis treated with fluid and insulin infusion controlling blood glucose concentration at 6-8 mmol/L and sepsis treated with fluid and insulin infusion controlling blood glucose concentration at 8-10 mmol/L. Hyperinsulinaemic-euglycaemic clamp experiment was done before fluid maintenance and insulin treatment to calculate average glucose infusion rate. All septic rats were markedly hyperglycaemic compared with sham-operated controls two hours after operation. Glucose infusion rate during hyperinsulinaemic-euglycaemic clamp experiment was slower in septic rats, suggesting that they were insulin resistant. At the 12th and 24th hour, skeletal muscle was taken to observe pathological change and analyse the GLUT4 mRNA and GLUT4 levels. There were more inflammatory cells, less GLUT4 mRNA and GLUT4 expression in the skeletal muscles of septic rats. Insulin increased the expression of GLUT4 mRNA and GLUT4 in the skeletal muscle of septic rats. Among all septic rats, the expression of GLUT4 mRNA and GLUT4 was more in the 8-10 mmol/L group. Blood glucose concentration of 8-10 mmol/L results in more glucose utilization than 6-8 mmol/L in the skeletal muscle of septic rats during insulin therapy.

18-Carbon polyunsaturated fatty acids ameliorate palmitate-induced inflammation and insulin resistance in mouse C2C12 myotubes

Skeletal muscle is a major site of insulin action. Intramuscular lipid accumulation results in inflammation, which has a strong correlation with skeletal muscle insulin resistance (IR). The aim of this study was to explore the effects of linoleic acid, alpha-linolenic acid, and gamma-linolenic acid (GLA), 18-carbon polyunsaturated fatty acids (PUFAs), on palmitic acid (PA)-induced inflammatory responses and IR in C2C12 myotubes. Our data demonstrated that these three test 18-carbon PUFAs can inhibit PA-induced interleukin-6 and tumor necrosis factor-α messenger RNA (mRNA) expression and IR as evidenced by increases in phosphorylated AKT and the 160-kD AKT substrate, mRNA and plasma membrane protein expression of glucose transporter 4, and glucose uptake. Moreover, the 18-carbon PUFAs blocked the effects of PA on activation of mitogen-activated protein kinases (MAPKs), protein kinase C-θ (PKC-θ), AMP-activated protein kinase (AMPK) and nuclear factor-κB (NF-κB). Of note, supplementation with GLA-rich borage oil decreased proinflammatory cytokine production and hindered the activation of MAPKs, PKC-θ and NF-κB in the skeletal muscles of diabetic mice. The 18-carbon PUFAs did not reverse PA-induced inflammation or IR in C2C12 myotubes transfected with a constitutively active mutant IκB kinase-β plasmid, which suggests the importance of the inhibition of NF-κB activation by the 18-carbon PUFAs. Moreover, blockade of AMPK activation by short hairpin RNA annulled the inhibitory effects of the 18-carbon PUFAs on PA-induced IR but not inflammation. Our findings suggest that the 18-carbon PUFAs may be useful in the management of PA-induced inflammation and IR in myotubes.

White tea consumption improves cardiac glycolytic and oxidative profile of prediabetic rats

Prediabetes is associated with cardiovascular diseases. We hypothesized that white tea (WTEA) consumption by prediabetic rats improved cardiac glycolytic and oxidative profile. Prenatal male Wistar rats were divided in control group and prediabetes (PrDM) group which received a streptozotocin injection. After one month, PrDM rats were divided: half consumed water and half WTEA during two months. WTEA phytochemical profile was determined. Rats were then subjected to glucose tolerance and insulin resistance tests. Cardiac glycolytic and oxidative profiles were determined. Prediabetes decreased glucose transporters (GLUT1 and GLUT3) mRNA expression, lactate and acetate content and lactate dehydrogenase activity in cardiac tissue. Prediabetes also decreased heart antioxidant capacity, increasing lipid peroxidation and protein oxidation. WTEA consumption improved glucose tolerance and insulin sensitivity in prediabetic rats and increased cardiac acetate and alanine contents and protein oxidation levels. WTEA consumption ameliorated overall metabolic status of prediabetic rats and prevented most of heart-related deleterious effects evaluated.

Vaspin promotes 3T3-L1 preadipocyte differentiation

Vaspin, a novel adipocyte factor secreted from visceral adipose tissues, is associated with obesity and insulin resistance and can regulate glucose and lipid metabolism, increase insulin sensitivity, and suppress inflammation; however, the underlying mechanisms remain unknown. Proliferation and maladaptive differentiation are important pathological mechanisms underlying obesity. This study aimed to evaluate the effects of vaspin on the proliferation and differentiation of preadipocyte 3T3-L1 cells and to explore the likely mechanisms responsible for 3T3-L1 differentiation. Vaspin was added to cultured 3T3-L1 cells, and the differentiation of adipocytes was evaluated using Oil Red O staining. The AKT signaling pathway and specific differentiation factors related to the differentiation of preadipocyte 3T3-L1 cells, peroxisome proliferator-activated γ and the CCAAT/enhancer-binding protein (C/EBP) family, were evaluated using reverse transcription polymerase chain reaction (RT-PCR) and western blot analyses during the early phase of differentiation. Additionally, adiponectin mRNA, interleukin-6 mRNA (IL-6 mRNA), and glucose transporter-4 (GLUT4) protein levels were measured in the differentiated adipocytes. The results indicated that vaspin promotes the intracellular accumulation of lipids and increases differentiation-related factors, including peroxisome proliferator-activated receptor γ, C/EBPα, and free fatty acid-binding protein 4 (FABP4), in a dose-dependent manner. Additionally, vaspin (200 ng/mL) increased the mRNA and protein levels of C/EBPβ, peroxisome proliferator-activated γ, C/EBPα, and FABP4. Moreover, compared with the control, significantly smaller eight-day differentiated adipocytes were observed, and these cells exhibited decreased IL-6 mRNA and increased GLUT4 mRNA levels; these results also indicated the potential of vaspin to promote the insulin-mediated AKT signaling pathway during the early phase of differentiation. In conclusion, vaspin is able to promote the differentiation of 3T3-L1 preadipocytes and may increase their sensitivity to insulin and suppress obesity.

Effect of Intermittent Hypoxia and Rimonabant on Glucose Metabolism in Rats: Involvement of Expression of GLUT4 in Skeletal Muscle.

BackgroundObstructive sleep apnea (OSA) and its main feature, chronic intermittent hypoxia (IH) during sleep, is closely associated with insulin resistance (IR) and diabetes. Rimonabant can regulate glucose metabolism and improve IR. The present study aimed to assess the effect of IH and rimonabant on glucose metabolism and insulin sensitivity, and to explore the possible mechanisms.Material/MethodsThirty-two rats were randomly assigned into 4 groups: Control group, subjected to intermittent air only; IH group, subjected to IH only; IH+NS group, subjected to IH and treated with normal saline; and IH+Rim group, subjected to IH and treated with 10 mg/kg/day of rimonabant. All rats were killed after 28 days of exposure. Then, the blood and skeletal muscle were collected. We measured fasting blood glucose levels, fasting blood insulin levels, and the expression of glucose transporter 4 (GLUT4) in both mRNA and protein levels in skeletal muscle.ResultsIH can slow weight gain, increase serum insulin level, and reduce insulin sensitivity in rats. The expressions of GLUT4 mRNA, total GLUT4, and plasma membrane protein of GLUT4 (PM GLUT4) in skeletal muscle were decreased. Rimonabant treatment was demonstrated to improve weight gain and insulin sensitivity of the rats induced by IH. Rimonabant significantly upregulated the expression of GLUT4 mRNA, PM GLUT4, and total GLUT4 in skeletal muscle.ConclusionsThe present study demonstrates that IH can cause IR and reduced expression of GLUT4 in both mRNA and protein levels in skeletal muscle of rats. Rimonabant treatment can improve IH – induced IR, and the upregulation of GLUT4 expression may be involved in this process.

Effect of Solanum surattense on mitochondrial enzymes in diabetic rats and in vitro glucose uptake activity in L6 myotubes.

Background:S. surattense is widely used in Siddha medicine for various ailments.Objective:The aim was to evaluate the impact of alcoholic leaf-extract of S. surattense on mitochondrial enzymes in streptozotocin (STZ) induced diabetic rats and to study the in vitro muscle glucose uptake activity on L6 myotubes.Materials and Methods:The male albino Wistar rats were randomly divided into five groups of six animals each. Diabetes was induced by intraperitoneal injection of STZ (40 mg/kg body weight). After being confirmed the diabetic rats were treated with alcoholic leaf-extract of S. surattense (100 mg/kg body weight) for 45 days. The biochemical estimations (liver mitochondrial enzymes, antioxidants, thiobarbituric acid reactive substances [TBARS]) and histopathological studies were performed. Further, the in vitro muscle glucose uptake activity in L6 myotubes and messenger RNA (mRNA) expression of glucose transporter-4 (GLUT-4) was performed.Results:In diabetic rats, the activities of liver mitochondrial enzymes were found to be significantly lowered. The mitochondrial TBARS level increased, whereas the activities/level of enzymatic and non-enzymatic antioxidants decreased in diabetic rats. Administration of S. surattense to diabetic rats significantly reversed the above parameters toward normalcy. Furthermore in diabetic rats, the histopathological studies showed growth of adipose tissue and shrinkage of islets in the pancreas, liver showed fatty change with mild inflammation of portal triad, and kidney showed messangial capillary proliferation of glomeruli and fatty infiltration of tubules. Treatment with S. surattense brought back these changes to near normalcy. The extract was analyzed for in vitro muscle glucose uptake activity in L6 myotubes and mRNA expression of GLUT-4 by semi-quantitative reverse transcriptase-polymerase chain reaction. One nano gram per millilitre of S. surattense leaf-extract gave 115% glucose uptake on L6 myotubes. It also showed elevated levels of GLUT-4 mRNA transcripts, when compared with control cells.Conclusion:These studies strongly support the anti-diabetic nature of S. surattense.

Dysregulation of Glucose Homeostasis Following Chronic Exogenous Administration of Leptin in Healthy Sprague-Dawley Rats.

Impaired glucose utilization is seen in chronic hyperleptinaemia associated conditions such as obesity and type 2 diabetes mellitus. It is unclear if this impaired glucose utilization is due to the effect of persistent hyperleptinaemia on insulin secretion from the beta cells of pancreas. To examine the effects of chronic leptin administration on plasma glucose regulation in rats. Glucose challenge curves were plotted for male Sprague-Dawley rats treated with either normal saline (Control; n=8) or subcutaneous leptin injection for 42 days (60 μg/kg body weight/day; n=8). Plasma glucose and plasma insulin levels were measured at 0, 5, 10, 15, 20 and 25 minutes after glucose challenege. Skeletal muscle tissue was collected at the end of a glucose challenge for glucose transporter-4 protein content, insulin receptor and glucose transporter-4 mRNA expression. Data were analysed using repeated measures and one-way ANOVA with post-hoc analysis. Chronic leptin treatment caused significantly higher fasting insulin level. Post glucose challenge, there was a significant increase in blood glucose levels and insulin level in the leptin treated rats. There was no significant difference in the skeletal muscle glucose transporter-4 content. However, leptin treated rats showed decreased mRNA expression of Insulin Receptor and glucose transporter-4 in the skeletal muscle. Leptin administration for 42 days caused hyperinsulinaemia and decreased the expression of insulin receptors in insulin sensitive tissues leading to the development of an insulin resistance-like state in the rats.

Angiotensin II receptor blocker ameliorates stress-induced adipose tissue inflammation and insulin resistance.

A strong causal link exists between psychological stress and insulin resistance as well with hypertension. Meanwhile, stress-related responses play critical roles in glucose metabolism in hypertensive patients. As clinical trials suggest that angiotensin-receptor blocker delays the onset of diabetes in hypertensive patients, we investigated the effects of irbesartan on stress-induced adipose tissue inflammation and insulin resistance. C57BL/6J mice were subjected to 2-week intermittent restraint stress and orally treated with vehicle, 3 and 10 mg/kg/day irbesartan. The plasma concentrations of lipid and proinflammatory cytokines [Monocyte Chemoattractant Protein-1 (MCP-1), tumor necrosis factor-α, and interleukin-6] were assessed with enzyme-linked immunosorbent assay. Monocyte/macrophage accumulation in inguinal white adipose tissue (WAT) was observed with CD11b-positive cell counts and mRNA expressions of CD68 and F4/80 using immunohistochemistry and RT-PCR methods respectively. The mRNA levels of angiotensinogen, proinflammatory cytokines shown above, and adiponectin in WAT were also assessed with RT-PCR method. Glucose metabolism was assessed by glucose tolerance tests (GTTs) and insulin tolerance tests, and mRNA expression of insulin receptor substrate-1 (IRS-1) and glucose transporter 4 (GLUT4) in WAT. Restraint stress increased monocyte accumulation, plasma free fatty acids, expression of angiotensinogen and proinflammatory cytokines including MCP-1, and reduced adiponectin. Irbesartan reduced stress-induced monocyte accumulation in WAT in a dose dependent manner. Irbesartan treatment also suppressed induction of adipose angiotensinogen and proinflammatory cytokines in WAT and blood, and reversed changes in adiponectin expression. Notably, irbesartan suppressed stress-induced reduction in adipose tissue weight and free fatty acid release, and improved insulin tolerance with restoration of IRS-1 and GLUT4 mRNA expressions in WAT. The results indicate that irbesartan improves stress-induced adipose tissue inflammation and insulin resistance. Our results suggests that irbesartan treatment exerts additive benefits for glucose metabolism in hypertensive patients with mental stress.

177 DOWN-REGULATION OF P450 AROMATASE AND GLUCOSE TRANSPORTER 4 mRNAs EXPRESSION IN SHEEP OVARIAN FOLLICLES AFTER ULTRASHORT NUTRITIONAL FLUSHING

Nutritional supplementation before breeding (flushing) has become a common practice and is a reliable method to increase lambing and twining rates in sheep. We have shown that ultrashort flushing (USF: 1 day long) acutely increases ovulation rate without affecting the body condition or animal weight. A short nutritional stimulus would necessarily be acting either directly at the ovarian level or through hormones and factors other than gonadotropins. The possible mechanisms of action of USF on ovulation rate is the deregulation of the feedback loop between the ovaries and gonadotropin secretion, where nutrition causes a direct inhibition of follicular oestradiol production leading to compensatory secretion of FSH that stimulates folliculogenesis, or direct stimulation of follicular development through hormones and factors other than gonadotropins. We hypothesised that USF enhances the number of follicles selected for ovulation due to either a decrease in the expression of P450 aromatase or alternatively by advancing the maturation of the ovarian follicles, which would be reflected in an increase in LH receptor (LHr) and 3β-hydroxysteroid dehydrogenase (3β-HSD) mRNA expression, an increase in glucose transporter 4 (GLUT4) mRNA expression, or both. To test these hypotheses, the oestrus cycle of 30 ewes was synchronized with progestin intravaginal sponges and prostaglandins. Animals were given the USF on the day of progestin withdrawal. The ovaries were removed surgically before treatment (time 0; n = 6), or at 12, 24, and 48 h after being treated with either glycerol or water (n = 4 per treatment by time category). The ovarian follicles were dissected out and counted. Follicles larger than 3 mm in diameter were pooled and the mRNA extracted for specific P450aromatase, 3β-HSD, LHr, and glucose transporter 4 determinations. USF increased the number of follicles larger than 3 mm at 48 h after treatment (P &lt; 0.05). No effect was observed in the number of small follicles. In the two largest follicles, aromatase and GLUT4 mRNA abundance decreased (P &lt; 0.01) 12 h after treatment. There was no effect of treatment on mRNA for LH receptor or 3β-HSD. These results demonstrate a reduction in aromatase expression in potentially ovulatory follicles 12 h after flushing. The decrease in aromatase may favour a transient increase in FSH concentrations 12 h after flushing that would allow the stimulation and selection of supplementary ovulatory follicles.

Heme oxygenase-1 plays a crucial role in chemoresistance in acute myeloid leukemia

ObjectivesThe heme oxygenase-1 (HO-1) gene may contribute to the development of acquired chemoresistance in solid tumor cells, but its function in acute myeloid leukemia (AML) remains unclear. Therefore, we investigated whether the expressions of HO-1 mRNA and protein were associated with AML chemoresistance.MethodsBone marrow or peripheral blood was obtained from newly diagnosed (n = 26), relapsed (n = 10), and completely remitted (n = 18) patients with AML (M3 exclusion) and healthy donors (n = 10). Small interfering RNA was used to stably silence HO-1 gene expression in AML cell lines. The expressions of HO-1, hypoxia inducible factor-1ɑ (HIF-1ɑ), glucose transporter-1 (GLUT1) mRNA and proteins were measured by quantitative real-time PCR and Western blot. Cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis induction was analyzed by flow cytometry.ResultsThe drug-resistant AML cell line HL-60R was significantly less sensitive to cytarabine and daunorubicin than HL-60 cells. HO-1 mRNA and proteins were highly expressed in HL-60R cells. However, down-regulating HO-1 significantly enhanced the sensitivity of HL-60R to chemotherapy, and the expressions of HIF-1ɑ and GLUT1 mRNA and proteins decreased. Meanwhile, the expressions of caspase-3 and caspase-8 proteins increased, while that of bcl-2 decreased. Overexpressions of HO-1, HIF-1ɑ, and GLUT1 were associated with poor response of AML to chemotherapy.ConclusionsOverexpressions of HO-1, HIF-1ɑ, and GLUT1 might be involved in the chemoresistance of AML. HO-1 is a potential target to overcome the drug resistance of AML.