【作者】 张浩卫；

【导师】 杨公社；

【作者基本信息】 西北农林科技大学 ， 动物学， 2007， 硕士

【摘要】 猪与人在生理生化特点和遗传上的相似性,使得猪成为研究人类相关疾病的良好动物模型,特别是肥胖及其相关的2型糖尿病。Insulin抵抗和leptin抵抗伴随肥胖和2型糖尿病的发生,这两种抵抗也是机体能量代谢紊乱的诱因。脂肪作为能量代谢的重要组织,在能量代新中发挥重要的作用,同时也作为一个内分泌器官分泌诸如leptin、TNFα、IL6等因子,参与能量代谢的调节。SOCS-3作为细胞信号抑制因子家族的主要成员,参与了脂肪细胞中多种细胞因子的信号转导,例如leptin、IGF-1和insulin信号通路,也参与了insulin和leptin抵抗。在胰岛素信号通路中PPARγ和GLUT4基因作为其下游基因在胰岛素抵抗中有着显著的变化。2型糖尿病以胰岛素抵抗为特征,机体常出现高胰岛素血症和高糖血症。因此本实验以培养的猪原代脂肪细胞为研究对象,通过研究在胰岛素诱导、高胰岛素联合高糖诱导的条件下和地塞米松诱导的胰岛素抵抗状态下SOCS-3及其胰岛素信号通路相关基因和OB基因表达规律,以及胰岛素抵抗状态下leptin和insulin对SOCS-3基因表达的影响,初步探讨SOCS-3引起胰岛素抵抗的作用及其机理。主要研究结果如下:1、100 nmol/L的胰岛素作用脂肪细胞2h可以诱导PPARγ、GLUT4和SOCS-3基因的大量表达,而不能诱导OB基因的大量表达。胰岛素诱导GLUT4基因的表达,可能是通过诱导PPARγ的表达,然后通过PPARγ促进了GLUT4的表达。SOCS-3基因在胰岛素诱导高表达的情况下,并没有引起胰岛素抵抗。2、100 nmol/L胰岛素和20 mmol/L葡萄糖可以诱导OB基因表达量的增加,其对PPARγ、GLUT4和SOCS-3基因表达量的影响与胰岛素单一作用一致。推测在肥胖和糖尿病患者中的高胰岛素血症和高糖血症伴随的高leptin血症可能是由于胰岛素和高糖诱导了Leptin的表达。3、300 nmol/L的地塞米松诱导了脂肪细胞中SOCS-3基因的高效表达,随后也抑制了PPARγ和GLUT4的表达,而OB基因的表达量上升。因此,推测地塞米松诱导脂肪细胞胰岛素抵抗主要是通过SOCS-3持续大量的表达引起。而胰岛素刺激SOCS-3基因的表达由于其表达量不足只能抑制胰岛素信号而不能引起胰岛素抵抗。同时OB基因表达量上升,提示OB基因可能也参与了上调SOCS-3基因的表达量。4、300 noml/L的地塞米松处理24h,然后除去地塞米松24h,再用100 nmol/L的insulin和leptin分别处理2小时,而SOCS-3基因的表达量没有上升。推测地塞米松诱导SOCS-3的表达,虽然除去地塞米松24h后SOCS-3基因的表达量下降了,可能SOCS-3蛋白的表达量并没有下降。因此当用胰岛素和leptin处理脂肪细胞时,它们的作用信号被SOCS-3蛋白抑制,而不能象其单独作用一样诱导SOCS-3的表达。进一步表明SOCS-3的持续大量表达是引起胰岛素和leptin抵抗的原因。更多还原

【Abstract】 Porcine is an ideal model of human disease, because porcine is remarkably similar to human biological, metabolic and genetic levels. Insulin resistance and leptin resistance are the character of type 2 diabetes. They induce metabolic disorganized. The adipocyte plays a critical role in energy balance. Adipocyte also has endocrinic function which can secrete many kinds of adipokines. Such as: Leptin, TNFα,IL6 and so on, which can regulate energy metabolism. SOCS-3 is one of the main members of the SOCS family. Many cytokines’signaling such as Leptin, Insulin and IGF-Ⅰwas regulated by SOCS-3 in adipocyte. It can induce insulin resistance and leptin resistance. In this study, we investigated the expression pattern of SOCS-3 mRNA in primary culture adipocyte, which treated with insulin, insulin and glucose and dexamethasone. We also investigated the expression pattern of OB, PPARγand GLUT4 gene and tried to explore the regulating mechanism of SOCS-3 inducing insulin resistance.The main results were summarized as following:1. 100 nmol/L insulin treated adipocyte 2h can increased the expression of PARγ、GLUT4 and SOCS-3 mRNA. But it can be no significantly effect on the expression of OB gene. The increase of GLUT4 induced by insulin, maybe first increased PPARγ, then PPARγcaused increase of GLUT4 mRNA. The expression of SOCS-3 increased, but did not cause insulin resistance.2. 100 nmol/L insulin and 20 mmol/L glucose treated the primary culture adipocyte. It can induce OB gene expression. The effect on PPARγ、GLUT4 and SOCS-3 mRNA were similar with 100noml/L insulin treated. The high insulin and high glucose can induce OB mRNA expression, that is similar with type 2 diabetes, they are hyperinsulinemia, hyperglycemia and hyperleptinemia.3. The expression of PPARγand GLUT4 decreased, SOCS-3 and OB increased in porcine adipocyte which was treated by 300 nmol/L Dex. Maybe the mass increased of SOCS-3 induced insulin resistance. Although insulin induced expression of SOCS-3, SOCS-3 was not enough to cause insulin resistance. We also found the increase of OB gene expression. Maybe OB had partial effect on Dex increased SOCS-3 expression.4. After the primary porcine adipocyte was treated with 300nmol/L Dex for 24h, Dex was taken out for 24h. Then,the cell was treated with 100 nmol/L insulin and leptin, respectively. The arrested effect on SOCS-3 expression can not be alleviated by newly additive insulin or leptin. Maybe Dex increased SOCS-3. After took out Dex, maybe the mRNA expression of SOCS-3 gene was decreased, but the protein of SOCS-3 was also high. So when the adipocyte was treated by insulin and leptin, the SOCS-3 gene expression can not be induced. The effect of insulin and leptin was suppressed by SOCS-3. Maybe the continuance and mass expression of SOCS-3 induced insulin resistance and leptin resistance.更多还原