酵母菌葡萄糖耐量因子对3T3-L1脂肪细胞葡萄糖代谢的影响及机理

酵母菌葡萄糖耐量因子（GTF）作为铬的最有效活性形式之一，对维持机体血糖的正常水平具有重要作用。受分离纯化技术以及菌体含量低所限，胰岛素通路下GTF对葡萄糖代谢作用及其机理的研究没有完全展开。 本研究通过已建立的膜过滤、制备型色谱的方法从高GTF酵母菌里批量制备GTF，研究GTF和不同铬制剂对胰岛素抵抗3T3-L1脂肪细胞葡萄糖代谢作用的影响，活性中心铬与葡萄糖代谢作用之间的关系；在此基础上，利用实时荧光定量PCR和蛋白印迹技术分析GTF和不同铬制剂对抵抗型脂肪胰岛素受体、受体底物-1及葡萄糖转运蛋白的mRNA及蛋白的表达和活性的影响、通过免疫组化技术观察细胞内胰岛素受体分布；通过酶连免疫反应等研究代谢过程中酪氨酸激酶等的活性变化规律。从糖代谢调控作用、胰岛素通路中相关的蛋白及基因表达影响以及受体磷酸化过程中参与酶的活性变化规律三个层面分析和互相印证，研究GTF对葡萄糖代谢的作用机理。

As the most bioactive form of chromium，glucose tolerance factor (GTF) play an important role in maintaining normal blood sugar level. The mechanism of GTF on glucose metabolism is not clear. In this study, the pure sample of GTF are prepared from the established method. The glucose metabolism of GTF, high-molecular-weight chromium-binding substance and chromium picolinate on insulin resistent 3T3-L1 adipocyte cell are determinded by GOD-GLU and MTT. The relationship between Chromium and its activity are analyzed. Furthermore, the mRNA expressions of marker genes(IR, IR-1 and GLUT-4) involved in the glucose metabolism are determined by qRT-PCR and their protein expression are determined by Western blot. At last, the regulation of Phosphotyrosine phosphatase(PTP) and Tyrosine Protein Kinase (TPK) during Tyr phosphorylation are intensively studited. The molecular mechanisms of GTF and its pathway in vitro cell are demonstrated from the above three levels.