Ezetimibe decreases SREBP-1c expression in liver and reverses hepatic insulin resistance in mice fed a high-fat diet

Ezetimibe inhibits intestinal cholesterol absorption, thereby reducing serum cholesterol. Recent studies suggest that ezetimibe affects liver steatosis and insulin resistance. We investigated the impact of ezetimibe on insulin sensitivity and glucose metabolism in C57BL/6 mice. We analyzed 4 mouse groups fed the following diets: normal chow (4% fat) for 12 weeks, normal chow for 10 weeks followed by normal chow plus ezetimibe for 2 weeks, high-fat chow (32% fat) for 12 weeks, and high-fat chow for 10 weeks followed by high-fat chow plus ezetimibe for 2 weeks. In the normal chow + ezetimibe group, ezetimibe had no impact on body weight, fat mass, lipid metabolism, liver steatosis, glucose tolerance, or insulin sensitivity. In the high-fat chow + ezetimibe group, ezetimibe had no impact on body weight or fat mass but significantly decreased serum low-density lipoprotein cholesterol, triglyceride, and glutamate pyruvate transaminase levels; liver weight; hepatic triglyceride content; and hepatic cholesterol content and increased the hepatic total bile acid content. In association with increases in IRS-2 and Akt phosphorylation, ezetimibe ameliorated hepatic insulin resistance in the high-fat chow + ezetimibe group, but had no effect on insulin sensitivity in primary cultured hepatocytes. A DNA microarray and Taqman polymerase chain reaction revealed that ezetimibe up-regulated hepatic SREBP2 and SHP expression and down-regulated hepatic SREBP-1c expression. SHP silencing mainly in the liver worsened insulin resistance, and ezetimibe protected against insulin resistance induced by down-regulation of SHP. Ezetimibe down-regulated SREBP-1c in the liver and reversed hepatic insulin resistance in mice fed a high-fat diet.     

Mutations of the human glucokinase gene and diabetes mellitus.

The enzyme glucokinase catalyzes the phosphorylation of glucose and plays a key role in the regulation o f insulin secretion by pancreatic beta cells and glucose disposal in hepatocytes. Recent studies have shown that mutations in the gene encoding this key regulatory enzyme of glycolysis are a common cause of an autosomal dominant form of non-insulin-dependent (type 2) diabetes mellitus that has an onset often during childhood. The association of mutations in the glucokinase gene with impaired pancreatic cell function underscores the importance of glycolysis in the regulation of insulin secretion and suggests that mutations in other genes expressed in the beta-cell that also control rate-limiting steps in glucose metabolism may lead to diabetes.     

Sterol regulatory element-binding protein 1 mediates liver X receptor-beta-induced increases in insulin secretion and insulin messenger ribonucleic acid levels

Liver X receptors (LXRalpha and LXRbeta) regulate glucose and lipid metabolism. Pancreatic beta-cells and INS-1E insulinoma cells express only the LXRbeta isoform. Activation of LXRbeta with the synthetic agonist T0901317 increased glucose-induced insulin secretion and insulin content, whereas deletion of the receptor in LXRbeta knockout mice severely blunted insulin secretion. Analysis of gene expression in LXR agonist-treated INS-1E cells and islets from LXRbeta-deficient mice revealed that LXRbeta positively regulated expression of ATP-binding cassette transporter A1 (ABCA1), sterol regulatory element-binding protein 1 (SREBP-1), insulin, PDX-1, glucokinase, and glucose transporter 2 (Glut2). Down-regulation of SREBP-1 expression with the specific small interfering RNA blocked basal and LXRbeta-induced expression of pancreatic duodenal homeobox 1 (PDX-1), insulin, and Glut2 genes. SREBP-1 small interfering RNA also prevented an increase in insulin secretion and insulin content induced by T0901317. Moreover, 5-(tetradecyloxy)-2-furoic acid, an inhibitor of the SREBP-1 target gene acetyl-coenzyme A carboxylase, blocked T0901317-induced stimulation of insulin secretion. In conclusion, activation of LXRbeta in pancreatic beta-cells increases insulin secretion and insulin mRNA expression via SREBP-1-regulated pathway. These data support the role of LXRbeta, SREBP-1, and cataplerosis/anaplerosis pathways in the control of insulin secretion in pancreatic beta-cells.     

Dietary olive oil and menhaden oil mitigate induction of lipogenesis in hyperinsulinemic corpulent JCR:LA-cp rats: microarray analysis of lipid-related gene expression

In the corpulent James C. Russell corpulent (JCR:LA-cp) rat, hyperinsulinemia leads to induction of lipogenic enzymes via enhanced expression of sterol-regulatory-binding protein (SREBP)-1c. This results in increased hepatic lipid production and hypertriglyceridemia. Information regarding down-regulation of SREBP-1c and lipogenic enzymes by dietary fatty acids in this model is limited. We therefore assessed de novo hepatic lipogenesis and hepatic and plasma lipids in corpulent JCR rats fed diets enriched in olive oil or menhaden oil. Using microarray and Northern analysis, we determined the effect of these diets on expression of mRNA for lipogenic enzymes and other proteins related to lipid metabolism. In corpulent JCR:LA-cp rats, both the olive oil and menhaden oil diets reduced expression of SREBP-1c, with concomitant reductions in hepatic triglyceride content, lipogenesis, and expression of enzymes related to lipid synthesis. Unexpectedly, expression of many peroxisomal proliferator-activated receptor-dependent enzymes mediating fatty acid oxidation was increased in livers of corpulent JCR rats. The menhaden oil diet further increased expression of these enzymes. Induction of SREBP-1c by insulin is dependent on liver x receptor (LXR)alpha. Although hepatic expression of mRNA for LXR itself was not increased in corpulent rats, expression of Cyp7a1, an LXR-responsive gene, was increased, suggesting increased LXR activity. Expression of mRNA encoding fatty acid translocase and ATP-binding cassette subfamily DALD member 3 was also increased in livers of corpulent JCR rats, indicating a potential role for these fatty acid transporters in the pathogenesis of disordered lipid metabolism in obesity. This study clearly demonstrates that substitution of dietary polyunsaturated fatty acid for carbohydrate in the corpulent JCR:LA-cp rat reduces de novo lipogenesis, at least in part, by reducing hepatic expression of SREBP-1c and that strategies directed toward reducing SREBP-1c expression in the liver may mitigate the adverse effects of hyperinsulinemia on hepatic lipid production.     

Effect of long‐term treatment with a small‐molecule glucokinase activator on glucose metabolism,lipid profiles and hepatic function

We investigated the long‐term effect of a glucokinase (GK) activator (GKA) on the changes in hepatic gene expression, glucose metabolism, lipid profiles and hepatic function in wild‐type mice and the haploinsufficiency of β‐cell‐specific GK mice on a high‐fat (HF) diet. Twenty weeks of GKA treatment had no effect on hepatic GK activity or expression of genes related to glucose or lipid metabolism, suggesting that chronic GK activation by GKA showed a sustained reduction of ambient blood glucose levels without causing significant impact on hepatic lipid and glucose metabolisms. Furthermore, GKA exerted glucose‐lowering efficacy lasted for up to 40 weeks without increasing bodyweight or exerting adverse effects on lipid metabolism or hepatic function in either genotype on the HF diet. The present results show that GKA is capable of chronically improving glucose metabolism in mice on the HF diet without exerting a harmful influence on their lipid profile or hepatic function. (J Diabetes Invest,doi: 10.1111/j.2040‐1124.2011.00103.x, 2011)     

Glucose-induced lipogenesis in pancreatic beta-cells is dependent on SREBP-1

High concentrations of glucose induce de novo fatty acid synthesis in pancreatic β-cells and chronic exposure of elevated glucose and fatty acids synergize to induce accumulation of triglycerides, a phenomenon termed glucolipotoxicity.

Here we investigate the role of sterol-regulatory element binding proteins in glucose-induced lipogenesis in the pancreatic β-cell line INS-1E. We show that glucose induces SREBP-1c expression and SREBP-1 activity independent of insulin secretion and signaling. Using adenoviral expression of SREBP-1c and a SREBP-mutant we show that lipogenic gene expression, de novo fatty acid synthesis and lipid accumulation are induced primarily through sterol-regulatory elements (SREs) and not E-Boxes. Adenoviral expression of a dominant negative SREBP compromises glucose induction of some lipogenic genes and significantly reduces glucose-induction of de novo fatty acid synthesis. Thus, we demonstrate for the first time that SREBP activity is necessary for full glucose induction of de novo fatty acid synthesis in pancreatic β-cells.     

二甲双胍对人肝细胞Huh7脂质蓄积的影响及作用机制

目的研究二甲双胍对肝细胞脂质蓄积的影响及作用机制。方法体外培养人源Huh7肝细胞,细胞随机分为空白对照组及5 mmol/L二甲双胍、10 mmol/L二甲双胍、15 mmol/L二甲双胍处理组,确定最佳浓度后,使用LDL荷脂,然后分为空白对照组、LDL处理组和LDL+15 mmol/L二甲双胍处理组。实验终点,采用细胞增殖及毒性检测试剂盒检测细胞存活率,油红O染色和BODIPY脂质探针检测细胞内脂质含量,Dil-LDL检测细胞脂质摄取能力,Western blot分析固醇调节元件结合蛋白2(SREBP-2)、SREBP-1c、低密度脂蛋白受体(LDLR)和前蛋白转化酶枯草溶菌素9(PCSK9)蛋白的表达情况。结果二甲双胍可抑制LDL诱导的肝细胞脂质蓄积,降低SREBP-2、SREBP-1c、LDLR和PCSK9的蛋白表达。结论二甲双胍可能通过减少转录因子SREBP-2和SREBP-1c,下调LDLR表达,从而抑制细胞脂质蓄积。