十五烷酸对糖代谢的调控及机理探究

发布时间：2018-03-21 12:06

本文选题：十五烷酸　切入点：3T3-L1成熟脂肪细胞　出处：《华东师范大学》2017年硕士论文　论文类型：学位论文

【摘要】：糖代谢紊乱是引起糖尿病的一个关键的危险因素。饱和长链脂肪酸棕榈酸和硬脂酸对糖代谢产生不利影响,增加2型糖尿病的发病危险性,但不同的脂肪酸对糖代谢及糖尿病的发生和发展产生不同影响,与其链长度及饱和度相关。通过前期对降糖植物中活性天然产物的筛选,我们发现十五烷酸对成熟脂肪细胞具有非常显著的促进葡萄糖消耗的作用,提示十五烷酸可能对糖代谢起到一定的调节作用。属于长链脂肪酸的十五烷酸对糖代谢的影响尚不清楚,其作用机制也不明确。为此本研究采用脂肪细胞、骨骼肌细胞以及实验性类似2型糖尿病小鼠,在细胞和整体水平系统研究了十五烷酸对糖代谢的影响,并深入探究了其作用机制,旨在明确十五烷酸对糖代谢的调节作用及其作用靶点,为全面认识饱和长链脂肪酸的功能提供实验依据,并对合理摄入膳食脂肪酸提供了一定的理论依据。在细胞模型方面,不同浓度的十五烷酸能够显著促进基础状态3T3-L1成熟脂肪细胞和C2C12骨骼肌细胞对葡萄糖的消耗。在3T3-L1成熟脂肪细胞或者是C2C12骨骼肌细胞中十五烷酸与胰岛素联合给药,促进葡萄糖消耗的作用均显著强于胰岛素单独作用。进一步的机制探究,发现在基础状态下,十五烷酸对2种细胞中CAP、ERK、PPARγ、PPARβ、PPARα的蛋白表达无明显的影响。但能明显降低P-ERK、P-PPARγ(Ser112)的表达,并且在与胰岛素的联合作用下,十五烷酸同样可降低骨骼肌细胞中P-ERK、P-PPARy(Ser112)的表达,说明十五烷酸可以通过降低ERK的磷酸化来提高PPARγ的活性,这可能为促进细胞糖消耗起到了一定的帮助作用。此外,在2种细胞中十五烷酸都会降低基础状态及胰岛素联合处理下胰岛素信号通路中IRS、PI3K、AKT的磷酸化水平;但同时十五烷酸可增强基础状态及胰岛素联合处理下AMPK的磷酸化及其下游P38的磷酸化水平,提高AKT下游的AS160的磷酸化和膜蛋白Glut4的表达。不同的是,在骨骼肌细胞中十五烷酸能够明显提高P-PKC(ζ/λ)的表达,但是在脂肪细胞中却没有发现这一现象。上述结果提示,十五烷酸促进细胞糖消耗可能是通过提高AMPK的活性,激活P38的磷酸化进而促进下游的AS160的磷酸化,最终导致膜蛋白Glut4的转运。采用AMPK的抑制剂Compound C进行验证,发现Compound C可逆转十五烷酸促进的糖消耗,并能逆转十五烷酸促进的AMPK及AMPK下游的AS16O、aPKC、P38的磷酸化,证明十五烷酸是通过AMPK信号通路发挥促进糖消耗的作用。进一步采用高热量饮食联合STZ诱导的2型糖尿病小鼠模型,观察十五烷酸对2型糖尿病小鼠糖代谢的影响情况。十五烷酸对2型糖尿病小鼠的饮水、饮食、体重无明显影响;对小鼠空腹血糖、糖基化血红蛋白和胰岛素抵抗指数也无明显的影响,但十五烷酸能降低2型糖尿病小鼠异常升高的血清胰岛素水平,改善高胰岛素血症,降低血清炎症因子TNF-α平,通过HE染色观察到,十五烷酸阻止糖尿病对胰岛细胞的破坏作用。对于与糖代谢紊乱密切相关的脂代谢方面,十五烷酸对模型小鼠血清甘油三酯、高、低密度脂蛋白胆固醇、游离脂肪酸水平均无明显的影响,但它能显著性的减少附睾脂肪垫的重量,可降低血清总胆固醇含量。综上所述,十五烷酸主要通过AMPK途径促进脂肪细胞和骨骼肌细胞对葡萄糖的摄取。十五烷酸不影响2型糖尿病小鼠空腹血糖水平,但能改善高胰岛素血症,保护胰岛细胞,并能降低糖尿病小鼠血清胆固醇,减少脂肪的积累,改善糖尿病小鼠的炎症状态。
[Abstract]:Glucose metabolism is a key risk factor for diabetes. Saturated long-chain fatty acid palmitic acid and stearic acid on glucose metabolism adversely affected, increases the risk of type 2 diabetes, but have different effects of different fatty acids on glucose metabolism and the occurrence and development of diabetes, and the chain length and saturation. Through the screening of hypoglycemic activity. Plant natural products early, we found that fifteen alkyl acid has a very significant role in promoting the consumption of glucose on mature adipocytes, suggesting that fifteen alkyl acid may regulate the glucose metabolism. The effects of fifteen belong to acid and long chain fatty acid on glucose metabolism is unclear, the mechanism is not clear. This study uses fat cells, skeletal muscle cells and similar experimental type 2 diabetic mice, in the cell and the overall level of fifteen alkyl acid The influence on glucose metabolism, and explores its mechanism, to clear the fifteen alkyl acid on glucose metabolism regulation and its targets, and provide the experimental basis for the comprehensive understanding of saturated long-chain fatty acids, and certain theoretical basis for the reasonable intake of dietary fatty acids. In the cell model, fifteen acid of different concentrations to consumption based 3T3-L1 adipocytes and C2C12 skeletal muscle cells to glucose significantly enhanced. In 3T3-L1 adipocytes or C2C12 skeletal muscle cells in fifteen acetic acid combined with insulin administration, to promote the consumption of glucose effect was significantly stronger than insulin alone. Further study the mechanism, found in the baseline state, fifteen alkyl acid of 2 kinds of cells in CAP, ERK, PPAR gamma, PPAR beta, PPAR alpha protein expression had no significant effect. But it can significantly reduce P-ERK, P-PPAR gamma (Ser112). And in the expression, and the joint action of insulin, fifteen acid also reduced P-ERK in skeletal muscle cells, the expression of P-PPARy (Ser112), fifteen acid can improve PPAR gamma activity by inhibiting ERK phosphorylation, which may promote cell sugar consumption played a helpful role in addition. In fifteen, the 2 kinds of cells are under the acid combined treatment of insulin signaling and insulin IRS, reducing PI3K, AKT phosphorylation; but at the same time, fifteen alkyl acid combined treatment of AMPK and its downstream phosphorylation of P38 enhanced the basic condition and the phosphorylation level of insulin, increase the expression of AKT downstream the phosphorylation of AS160 and membrane protein Glut4. The difference is, fifteen alkyl acid can significantly improve P-PKC in skeletal muscle cells (zeta / lambda) expression, but in the fat cells but did not find this phenomenon. These results suggest that fifteen Acid sugar consumption may promote cell by increasing the activity of AMPK, activation of P38 phosphorylation and promote downstream phosphorylation of AS160, resulting in membrane protein translocation of Glut4. Using the AMPK inhibitor Compound C verifies that Compound C can reverse the fifteen alkyl acid promoted sugar consumption, and can reverse the fifteen alkyl acid to promote the downstream AMPK and AMPK AS16O, aPKC, P38 phosphorylation, fifteen alkyl acid is proved to play a role in promoting the consumption of glucose through the AMPK signaling pathway. The high calorie diet in type 2 diabetic mice model induced with STZ, observe fifteen alkyl acid on glucose metabolism in mice with type 2 diabetes diet for fifteen. Acid on mice with type 2 diabetes mellitus, drinking water, no significant effect on body weight in mice; fasting blood glucose, glycosylated hemoglobin and insulin resistance index have no obvious effect, but the fifteen alkyl acid can reduce type 2 diabetes. Serum insulin levels in mice increased abnormally, hyperinsulinemia, decreased serum inflammatory factor TNF- alpha level was observed by HE staining, fifteen damage acid to prevent diabetes on islet cells. The glucose metabolism and lipid metabolism disorders are closely related, fifteen alkyl acid on mice serum triglyceride, high and low the LDL had no significant effect of free fatty acid levels, but it can reduce the weight of epididymal fat pad significantly, can reduce serum total cholesterol content. To sum up, fifteen alkyl acid mainly through the AMPK pathway to promote glucose uptake in adipocytes and skeletal muscle cells. Fifteen acid does not affect the blood glucose level of 2 diabetic mice fasting, but it can improve the hyperinsulinemia, protect the islet cells, and can reduce the serum cholesterol in diabetic mice, reduce fat accumulation, improve diabetic mice A state of inflammation.

【学位授予单位】：华东师范大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R587.1

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