Peroxisomal proliferator-activated receptor alpha deficiency diminishes insulin-responsiveness of gluconeogenic/glycolytic/pentose gene expression and substrate cycle flux

Our previous work led to the hypothesis that peroxisomal proliferator-activated receptor alpha (PPAR alpha) modulates insulin action in a compensatory fashion for hepatic glucose balance vs. peripheral glucose disposal. Therefore, we have examined the expression of insulin-dependent gluconeogenic/glycolytic/pentose cycle enzymes and compared these to insulin responsiveness for peripheral vs. hepatic substrate flux and futile cycling in the PPAR alpha knockout mouse. Hepatic gluconeogenic flux, glucose absorption, clearance and recycling, as well as in vivo glucose disposal were evaluated using new mass isotopomer methods. Insulin-dependent gluconeogenic/glycolytic/pentose cycle enzyme expression and glucose futile cycling were diminished; however, glucose disappearance was increased. This supports the hypothesis of hepatic insulin resistance and increased peripheral glucose uptake as compensatory events secondary to the decrease in fatty acid oxidation characteristic of the PPAR alpha knockout. We conclude that 1) the loss of PPAR alpha results in lower expression levels and diminished response to meal regulation for gluconeogenic/glycolytic enzyme expression; and 2) consequently, substrate/futile cycling of glucose is decreased when PPAR alpha is absent despite increased gluconeogenesis. The compensatory changes in liver and peripheral tissue substrate flux and the resultant adaptation for enzyme expression in the liver to have a diminished insulin dependence reflect the loosely linked correlation between phenotype and genotype in hepatic glucose metabolism.     

Activation of liver X receptors promotes lipid accumulation but does not alter insulin action in human skeletal muscle cells

Aims/hypothesis The aim of this study was to investigate the effects of liver X receptor (LXR) activation on lipid metabolism and insulin action in human skeletal muscle cells prepared from control subjects and from patients with type 2 diabetes.Subjects and methods Cultured myotubes were obtained from muscle biopsies of 11 lean, healthy control subjects and ten patients with type 2 diabetes. The mRNA levels of LXR isoforms and lipogenic genes were estimated by RT-quantitative PCR, and the effects of LXR agonists on insulin action were evaluated by assays of protein kinase B serine 473 phosphorylation and glycogen synthesis.Results Both LXRα and LXRβ were expressed in human skeletal muscle and adipose tissue and there was no difference in their mRNA abundance in tissues from patients with type 2 diabetes compared with control subjects. In cultured muscle cells, LXR activation by T0901317 strongly increased expression of the genes encoding lipogenic enzymes, including sterol regulatory element binding protein 1c, fatty acid synthase and stearoyl-CoA desaturase 1, and also promoted triglyceride accumulation in the presence of a high glucose concentration. Importantly, these effects on lipid metabolism did not affect protein kinase B activation by insulin. Furthermore, LXR agonists did not modify insulin action in muscle cells from patients with type 2 diabetes.Conclusions/interpretation These data suggest that LXR agonists may lead to increased utilisation of lipids and glucose in muscle cells without affecting the mechanism of action of insulin. However, the long-term consequences of triglyceride accumulation in muscle should be evaluated before the development of effective LXR-based therapeutic agents.D. Cozzone, C. Debard and N. Dif contributed equally to this work.     

Impaired regulation of glucose transporter 4 gene expression in insulin resistance associated with in utero undernutrition.

The aim of this study was to investigate whether insulin resistance-associated in utero undernutrition was related to changes in insulin action on gene expression of molecules involved in the insulin signaling pathway and peripheral glucose metabolism in muscle and adipose tissue. Thirteen insulin-resistant subjects born with intrauterine growth retardation (IUGR) were matched for age, gender, and body mass index to 13 controls. Gene expression of insulin receptor, insulin receptor substrate-1, p85alpha phosphatidylinositol 3-kinase, glucose transporter-4 (GLUT4), hexokinase II, and glycogen synthase was studied in skeletal muscle at baseline and after a 3-h euglycemic insulin stimulation. Target messenger ribonucleic acid (mRNA) levels were quantified using the RT-competitive PCR method. Insulin-stimulated glucose uptake was significantly lower in IUGR-born subjects than in controls (36.9 +/- 12, 7 vs. 53.9 +/- 12.7 micromol/kg.min; P = 0.007), affecting both the glucose oxidation rate and the nonoxidative glucose disposal rate. At baseline, the expression of the six genes in muscle did not significantly differ between the two groups. The insulin-induced changes over baseline were comparable in both groups for all mRNAs, except GLUT4. In contrast to what observed in the control group (mean increment, 49 +/- 23%; P = 0.0009), GLUT4 expression was not stimulated by insulin in the IUGR group (8 +/- 8%; P = 0.42). Moreover, the magnitude of the defect in GLUT4 mRNA regulation by insulin was correlated to the degree of insulin resistance (r = 0.73; P = 0.01). A similar lack of significant GLUT4 mRNA stimulation by insulin was observed in the adipose tissue of IUGR-born subjects. In conclusion, insulin resistance in IUGR-born subjects is associated with an impaired regulation of GLUT4 expression by insulin in muscle and adipose tissue. Our data provide additional information about the mechanism of insulin resistance associated with in utero undernutrition and strengthen the role of glucose transport in the control of insulin sensitivity.     

Myocardial gene expression of glucose transporter 1 and glucose transporter 4 in response to uteroplacental insufficiency in the rat

Uteroplacental insufficiency causes intrauterine growth retardation (IUGR) and subsequent low birth weight, which predisposes the affected newborn towards adult Syndrome X. Individuals with Syndrome X suffer increased morbidity from adult ischemic heart disease. Myocardial ischemia initiates a defensive increase in cardiac glucose metabolism, and individuals with Syndrome X demonstrate reduced insulin sensitivity and reduced glucose uptake. Glucose transporters GLUT1 and GLUT4 facilitate glucose uptake across cardiac plasma membranes, and hexokinase II (HKII) is the predominant hexokinase isoform in adult cardiac tissue. We therefore hypothesized that GLUT1, GLUT4 and HKII gene expression would be reduced in heart muscle of growth-retarded rats, and that reduced gene expression would result in reduced myocardial glucose uptake. To prove this hypothesis, we measured cardiac GLUT1 and GLUT4 mRNA and protein in control IUGR rat hearts at day 21 and at day 120 of life. HKII mRNA quantification and 2-deoxyglucose-uptake studies were performed in day-120 control and IUGR cardiac muscle. Both GLUT1 and GLUT4 mRNA and protein were significantly reduced at day 21 and at day 120 of life in IUGR hearts. HKII mRNA was also reduced at day 120. Similarly, both basal and insulin-stimulated glucose uptake were significantly reduced in day-120 IUGR cardiac muscle. We conclude that adult rats showing IUGR as a result of uteroplacental insufficiency express significantly less cardiac GLUT1 and GLUT4 mRNA and protein than control animals (which underwent sham operations), and that this decrease in gene expression occurs in parallel with reduced myocardial glucose uptake. We speculate that this reduced GLUT gene expression and glucose uptake contribute towards mortality from ischemic heart disease seen in adults born with IUGR.     

肺癌肿瘤抑制因子1基因过表达对肝细胞糖脂代谢相关基因表达的影响

探讨肺癌肿瘤抑制因子1(TSLC1)基因表达上调对小鼠肝癌细胞Hepal-6糖脂代谢相关基因mRNA表达的影响.结果 显示,TSLC1基因过表达降低脂代谢基因脂肪酸合成酶(FAS)、乙酰辅酶A羧化酶(ACC)的表达(P<0.05或P<0.01),增加脂肪组织甘油三酯水解酶(ATGL)的表达(P<0.05),对激素敏感脂肪酶(HSL)、葡萄糖转运蛋白(GLUT)1和GLUT4的表达无显著影响,提示TSLC1过表达可能抑制肝细胞的脂肪生成,促进脂肪分解.

Abstract：

The effects of tumor suppressor in lung cancer-1(TSLC1)upregulation on gene expressions involved in glucose and lipid metabolism in Hepa1-6 cells were investigated.The results showed that TSLC1 overexpression decreased fatty acid synthase and acetyl CoA carboxylase expressions(P<0.05 or P<0.01),increased adipose triglyceride lipase expression(P<0.05),and did not change hormone-sensitive lipase,glucose transporter (GLUT)1,and GLUT4 expressions.These results suggest that TSLC1 overexpression may promote lipolysis and inhibit adipogenesis in liver cells.     

Genome-wide expression profiling; a panel of mouse tissues discloses novel biological functions of liver X receptors in adrenals

The liver X receptors alpha and beta (LXRalpha and LXRbeta ) are members of the nuclear receptor superfamily of proteins which are highly expressed in metabolically active tissues. They regulate gene expression of critical genes involved in cholesterol catabolism and transport, lipid and triglyceride biosynthesis and carbohydrate metabolism in response to distinct oxysterols and intermediates in the cholesterol metabolic pathway. The biological roles of the LXRs in tissues other than liver, intestine and adipose tissue are poorly elucidated. In this study we used global gene-expression profiling analysis to detect differences in expression patterns in several tissues from mice fed an LXR agonist or vehicle. Our results show that LXR plays an important role in the kidney, lung, adrenals, brain, testis and heart where several putative LXR target genes were found. The effects of the LXRs were further analysed in adrenals where treatment with an LXR agonist induced expression of adrenocorticotrophic hormone receptor, suppressed expression of uncoupling protein (UCP)-1 and UCP-3 as well as several glycolytic enzymes and led to increased serum corticosterone levels. These results indicate novel biological roles of the LXR including regulation of energy metabolism, glycolysis and steroidogenesis in the adrenals via alteration of expression profiles of putative target genes.     

不同糖代谢状态下大鼠肝X受体的表达及其对肝糖代谢的影响

目的 观察肝X受体(LXR)在不同糖代谢状态下肝内表达的变化情况及其对肝糖代谢酶磷酸烯醇式丙酮酸羧激酶(PEPCK)mRNA和葡萄糖激酶(GCK)mRNA表达的调节,阐明LXR信号通路对糖代谢的影响机制.方法 SD雄鼠36只分为正常对照组、糖尿病组、肥胖组和肥胖伴糖尿病组.测定各组大鼠的体重、血糖、胆固醇和甘油三酯.采用实时PCR检测各组大鼠肝脏LXR mRNA、PEPCK mRNA和GCK mRNA的表达情况,应用Western印迹测定肝脏LXR蛋白质的表达.结果 各实验组大鼠肝LXRmRNA表达均显著高于对照组(P＜0.05).Western印迹结果示各组LXR蛋白表达量和转录水平一致.与对照组和肥胖组相比,肥胖伴糖尿病组和糖尿病组肝PEPCK mRNA表达均明显上调,GCK mRNA均明显下调;与对照组相比,肥胖组PEPCKmRNA表达明显下降,GCKmRNA明显上升(均P＜0.05).结论 在非糖尿病阶段,LXR可能作为糖代谢的保护性受体,通过调节肝糖代谢酶,使血糖保持稳态.进入糖尿病阶段后,LXR的保护作用并不能逆转因胰岛素不足所致的糖代谢相关酶的异常改变,血糖不能恢复正常.

Abstract：

Objective To investigate the mechanism of liver X receptor(LXR)signal pathway in regulating glucose metabolism by observing the variety of LXR expression and its impacts on regulating the mRNA expression of PEPCK and GCK, the key enzyme of hepatic glucose metabolism in various glucose metabolic status in rats. Methods SD rats were chosen and divided into four groups:the CON group, the induced DM group, the OB group, and the induced OB+DM group. For each group of rats, body weight, blood glucose, serum triglycerides, and cholesterol were measured. Then the rats were sacrificed and the livers were collected and studied. Real-time PGR was used to measure the expressions of LXR mRNA, PEPCK mRNA, and GCK mRNA in the livers. Finally, the Western Blot assay was used to measure the liver LXR protein expression. Results The expression of LXR mRNA was significantly higher in DM,OB, and OB+DM groups than in CON group(P＜0.05).The Western blot results showed that the levels of protein were in accordance with the mRNA expression. Comparing to the CON and the OB groups, the PEPCK mRNA expression of the OB+DM and the DM groups was significantly higher, while the GCK mRNA expression of these two groups was significantly lower(P＜0.05). Comparing to the CON group, the PEPCK mRNA expression of the OB group was significantly lower, while the GCK mRNA expression of OB group was significantly higher(P＜0.05).Conclusions During non-diabetic phase, LXR could act as a protective receptor for glucose metabolism and keep glucose homeostasis by regulating the key enzymes of the hepatic glucose metabolism. While in the diabetic phase, the protective receptor LXR failed to reverse the change of the related enzymes caused by insulin deficiency, and finally the plasma glucose level was raised.     

Expression of genes involved in lipid metabolism correlate with peroxisome proliferator-activated receptor gamma expression in human skeletal muscle

Peroxisome proliferator-activated receptor gamma (PPAR-gamma) activation in adipose tissue is known to regulate genes involved in adipocyte differentiation and lipid metabolism. However, the role of PPAR-gamma in muscle remains unclear. To examine the potential regulation of genes by PPAR-gamma in human skeletal muscle, we used semiquantitative RT-PCR to determine the expression of PPAR-gamma, lipoprotein lipase (LPL), muscle carnitine palmitoyl transferase-1 (mCPT1), fatty acid-binding protein (FABP), carnitine acylcarnitine transferase (CACT), and glucose transporter-4 (GLUT4) in freeze-dried muscle samples from 14 male subjects. These samples were dissected free of adipose and other tissue contamination, as confirmed by minimal or absent adipsin expression. Between individuals, the messenger ribonucleic acid concentration of PPAR-gamma varied up to 3-fold, whereas LPL varied up to 6.5-fold, mCPT1 13-fold, FABP 4-fold, CACT 4-fold, and GLUT4 up to 3-fold. The expression of LPL (r2 = 0.54; P = 0.003), mCPT1 (r2 = 0.42; P = 0.012), and FABP (r2 = 0.324; P = 0.034) all correlated significantly with PPAR-gamma expression in the same samples. No significant correlation was observed between the expression of CACT and PPAR-gamma or between GLUT4 and PPAR-gamma. These findings demonstrate a relationship between PPAR-gamma expression and the expression of other genes of lipid metabolism in muscle and support the hypothesis that PPAR-gamma activators such as the antidiabetic thiazolidinediones may regulate fatty acid metabolism in skeletal muscle as well as in adipose tissue.     

Thiazolidinediones improve insulin sensitivity in adipose tissue and reduce the hyperlipidaemia without affecting the hyperglycaemia in a transgenic model of type 2 diabetes

Aim/hypothesis The aim of this study was to examine the effects of thiazolidinediones on the MKR mouse model of type 2 diabetes.Methods Six-week-old wild-type (WT) and MKR mice were fed with or without rosiglitazone or pioglitazone for 3 weeks. Blood was collected from the tail vein for serum biochemistry analysis. Hyperinsulinaemic–euglycaemic clamp analysis was performed to study effects of thiazolidinediones on insulin sensitivity of tissues in MKR mice. Northern blot analysis was performed to measure levels of target genes of PPAR agonists in white adipose tissue and hepatic gluconeogenic genes.Results Thiazolidinedione treatment of MKR mice significantly lowered serum lipid levels and increased serum adiponectin levels but did not affect levels of blood glucose and serum insulin. Hyperinsulinaemic–euglycaemic clamp showed that whole-body insulin sensitivity and glucose homeostasis failed to improve in MKR mice after rosiglitazone treatment. Insulin suppression of hepatic endogenous glucose production failed to improve in MKR mice following rosiglitazone treatment. This lack of change in hepatic insulin insensitivity was associated with no change in the ratio of HMW : total adiponectin, hepatic triglyceride content, and sustained hepatic expression of PPAR and stearoyl-CoA desaturase 1 mRNA. Interestingly, rosiglitazone markedly enhanced glucose uptake by white adipose tissue with a parallel increase in CD36, aP2 and GLUT4 gene expression.Conclusions/interpretation These data suggest that potentiation of insulin action on tissues other than adipose tissue is required to mediate the antidiabetic effects of thiazolidinediones in our MKR diabetic mice.