Ⅱ型糖尿病大鼠海马Alzheimer病样Tau蛋白过度磷酸化修饰及机制探讨

Alzheimer-like Hyperphosphorylation of Tau in Hippocapus of Rat with Type Ⅱ Diabetes

Tau蛋白过度磷酸化是Alzheimer病(Alzheimer′sdisease,AD)的一个重要特征.本研究检测了Ⅱ型糖尿病大鼠海马tau蛋白磷酸化水平,对其形成机制进行探讨.以同龄正常Wistar大鼠作为对照,高脂高蛋白高糖饮食加小剂量链脲佐菌素(streptozotocin,STZ)注射诱导造Ⅱ型糖尿病模型(T2DM组).放免法检测血浆胰岛素;葡萄糖氧化酶法检测血浆葡萄糖;蛋白质印迹技术检测各组大鼠海马内总tau蛋白、tau蛋白上部分位点磷酸化、神经细胞膜上胰岛素受体及葡萄糖转运子3(glucosetransport3,GLUT3)水平;表面等离子共振技术(surfaceplasmonresonance,SPR)检测细胞膜上胰岛素受体与血浆胰岛素结合力;γ-32P标记的ATP和特异性底物肽检测海马内胰岛素信号传导系统中的关键酶糖原合酶激酶-3β(glycogensynthasekinase-3β,GSK-3β)活性.结果显示,T2DM组血浆血糖、血浆胰岛素及运用HOMA-IR公式计算的胰岛素抵抗指数显著高于对照组.蛋白质印迹结果显示两组大鼠海马回总tau蛋白水平无差异;T2DM组中tau蛋白在Ser199、Thr212、Ser214、Thr217、Ser396及Ser422位点上的磷酸化水平均显著高于对照组;T2DM组海马神经细胞膜上胰岛素受体水平及与胰岛素结合的功能均显著低于对照组;GSK-3β活性检测结果显示,T2DM组大鼠模型海马回中GSK-3β活性明显增高.研究结果表明,Ⅱ型糖尿病中由于胰岛素抵抗导致GSK-3β激活从而出现AD样tau蛋白的过度磷酸化,葡萄糖代谢紊乱也可能在tau蛋白的过度磷酸化起一定作用.

Abnormally hyperphosphorylation of tau plays a critical role in the pathogenesis of Alzheimer disease （AD） and type Ⅱ diabetes is a known risk factor of AD. We therefore studied the phosphorylation of tau in type Ⅱ diabetic rats by Western blotting, and found that tau protein was hyperphosphorylated at several AD-related phosphorylation sites. The activity of glycogen synthase kinase 3β （GSK-3β）, a key component of insulin transduction pathway and a known tan kinase, was found to be increased in the brains of diabetic rats. Decreased β-subunit of the membrane insulin receptor and binding activity between insulin and insulin receptor induced down regulation of insulin signal pathway though plasma insulin was dramatically increased. In additional, the glucose transport 3 （GLUT3） on neuronal membrane was found to be reduced in type Ⅱ diabetic rats. These findings suggest that type Ⅱ diabetes increase the probability of AD by decreasing insulin receptor and binding activity to insulin which leads to increased insulin resistance and consequent upregulation of GSK-3β and hyperphosphorylation of tau protein. The decreased GLUT3 which indicated of impaired glucose metabolism may also contribute to tau hyperphosphorylation in type Ⅱ diabetes.