基于“脂肪-胰岛轴”探讨黄芪散改善胰岛素敏感性的机理研究

Insulin resistance is the key pathophysiologic process of Type Ⅱ diabetes, in which adipose tissue plays an essential role. The recent finding of "adipoinsular axis" elucidates that crosstalk between insulin and leptin maintains the homeostasis of glucolipide metabolism through a feedback loop. Clinical treatments of diabetes mellitus prompt combined medication, to develop new drug with multi-target is very important, and Chinese medicine formula are potential candidate. Till now there is no report of the application of "adipoinsular axis" in the research of anti-diabetes with Chinese Medicine formula. The preliminary study showed that HuangQiSan (HQS), a formula from an ancient medical book < Sheng Ji Zong Lu >, improved insulin sensitivity, accompanied by reducing the concentration of blood insulin and letpin, decreasing adipose mass and body weight, and with no impact on appetite. So we hypothesize that HQS improves insulin sensitivity by keeping the homeostasis of "adipoinsular axis". In this study the mechanism of insulin resistance improvement of HQS through affecting the generation, secretion and function of insulin and leptin is to be evaluated at the levels of general, cell and molecular, with the application of animal models involving leptin absent ob/ob obese mouse, lipodystrophia aP2-SREBF1c mouse and STZ plus high fat diet induced type Ⅱ diabetic obese mouse, and cell lines involving original cell, 3T3-L1 pro-adipose cell and MIN6 β cell, and the techniques of siRNA and co-culture of 3T3-L1 pro-adipose cell and MIN6 β cell. The outcomes of this study will throw lights on the knowledge of multi-target effects of Chinese Medicine formula, and prompt the development of indigenous innovative drugs.

胰岛素抵抗是Ⅱ型糖尿病核心病理变化，脂肪组织在其中发挥重要作用。近年"脂肪-胰岛轴"的发现揭示了胰岛素与脂肪因子瘦素之间存在交互作用机制，共同维持糖脂代谢平衡。糖尿病防治强调联合用药，多靶点药物是研发重点，中药复方潜力巨大。目前未见"脂肪-胰岛轴"应用于中药复方防治糖尿病的报道。前期研究证明古方黄芪散能改善胰岛素抵抗，降低血中胰岛素、瘦素浓度，降脂减肥。本项目提出黄芪散通过平衡"脂肪-胰岛轴"改善胰岛素抵抗假说，采用leptin缺失ob/ob肥胖小鼠、脂肪萎缩aP2-SREBF1c小鼠、STZ+高脂饮食诱导的Ⅱ型糖尿病肥胖小鼠等动物模型，及原代细胞、3T3-L1前脂肪细胞、MIN6β细胞等细胞模型，结合siRNA干扰、脂肪细胞与β细胞混合培养等技术方法，体内外结合，从整体、细胞、分子水平综合评价黄芪散对胰岛素、leptin生成、分泌及功能互动的影响，阐明黄芪散改善胰岛素抵抗的作用机制。

申请人经系统研究，结合“脂肪-胰岛轴”的内涵，提出工作假说：黄芪散改善胰岛素抵抗的关键是平衡“脂肪-胰岛轴”通过调节胰岛素和leptin的生成与分泌，增强胰岛素和leptin受体敏感性，协调二者作用，从而纠正糖脂代谢异常。.通过采用STZ+高脂饮食诱导的Ⅱ型糖尿病肥胖小鼠、leptin缺失ob/ob肥胖小鼠、脂肪萎缩aP2-SREBF1c小鼠等动物模型，及多种细胞模型、siRNA干扰等技术方法进行实验研究，发现了：.1、黄芪散降低STZ+高脂饮食引起的肥胖型Ⅱ型糖尿病小鼠模型的血糖、血TC、insulin、leptin，改善胰岛素敏感性，降低皮下及附睾周围脂肪组织重量，肝中TG和TC含量，改善机体糖脂代谢。.2、黄芪散能下调PEPCK的表达，提示黄芪散对糖异生有明显抑制作用；上调PPARγ和LPL，对SREBP1c、PPAR-α、Cpt1α、IRS-1、ACO的表达有下调趋势，促进脂肪组织的脂质合成，减少循环中脂质含量和脂质异位沉积，改善胰岛素敏感性，为减少“脂毒性”及改善胰岛素抵抗提供依据。.3、aP2-SREBP1c转基因小鼠是因脂肪萎缩引起的胰岛素抵抗模型，黄芪散对该模型小鼠无改善作用。黄芪散联合leptin能下调该模型肝脏PPARγ、 FAS及SCD1的表达，上调PPARa、 CPT1a、HSL的表达，减少脂质合成，促进脂质的氧化分解，降低脂质沉积；下调G6PC的表达，可能抑制糖异生；下调脂肪组织的炎症因子表达。.4、leptin缺失ob/ob肥胖小鼠为常用的Ⅱ型糖尿病模型，黄芪散对该模型小鼠无改善作用，表明缺乏leptin使黄芪散的作用受限。黄芪散联合leptin能上调肝脏中JAK2、STAT3、IRS2、PI3K、Akt2的mRNA表达，改善肝脏的胰岛素敏感性，同时减少外周组织脂质含量；下调皮下脂肪FAS、LPL、MCP1的表达，抑制脂肪合成，促进脂解，降低炎症反应，协同调节机体胰岛素敏感性。.5、细胞实验表明，黄芪散能活化AKT磷酸化，激活AKT通路；上调共培养体系中的SREBP1的表达，提示黄芪散能影响脂质合成，调节糖脂代谢。.本项目已发表中文核心3篇，待发表SCI 2篇。初步阐明黄芪散通过平衡“脂肪-胰岛轴”改善胰岛素抵抗的作用机制。为黄芪散改善糖脂代谢紊乱提供科学依据，并丰富了中药复方改善胰岛素敏感性作用机制的内涵