INTRACELLULAR O GLCNAC AND GLUCOTOXICITY

临床研究和对分离的胰岛或β细胞系的研究已经 表明β细胞长期暴露于超生理条件下， 葡萄糖水平导致β细胞功能受损， 2型糖尿病的发病机制。 葡萄糖如何发挥 这种对β细胞的毒性仍不清楚。 我们建议 葡萄糖代谢产物葡萄糖胺在β细胞功能中发挥作用， 这种葡萄糖毒性。 我们得出这个假设是因为 观察β细胞特异性的毒性机制 链脲佐菌素（STZ）。 STZ在化学上类似于N- 乙酰葡糖胺（GlcNAc）。 此外，我们还发现， 含有大约100倍以上的编码o- GlcNAc转移酶（OGT）。 这种酶修饰细胞核和细胞骨架 通过将单糖GlcNAc连接到丝氨酸或苏氨酸上来制备蛋白质 蛋白质中的残基。 由此产生的O-GlcNAc修饰似乎 来改变转录因子的活性。 我们发现STZ 阻断从蛋白质中去除o-GlcNAc的酶的活性。 用STZ处理大鼠导致O-GlcNAc的积累 特别是在胰腺β细胞中的修饰， β细胞死亡。 因为β细胞富含OGT 这些细胞可能对O-GlcNAc的积累最敏感 细胞内蛋白质。 我们还在其他细胞类型中发现， 核O-GlcNAc对周围葡萄糖浓度敏感。 如果β细胞也是如此，那么高血糖症和STZ可能 两者都增加β细胞O-GlcNAc含量，从而导致共同的 β细胞毒性机制。 在此提出的实验 补助金旨在确定高血糖是否确实导致 β细胞O-GlcNAc增加。 我们还将创造转基因小鼠 其中葡萄糖胺合成从葡萄糖增加的模型 或减少。 我们将确定这些变化的影响， 葡萄糖胺代谢对β细胞功能的影响。 我们还发现 cAMP依赖性蛋白激酶抑制负责 葡萄糖胺合成我们建议研究 葡糖胺合成被抑制。 这些研究将 确定葡萄糖胺在葡萄糖对β细胞毒性中的作用 和控制葡萄糖代谢为葡糖胺的手段。

Clinical studies and studies on isolated islets or Beta cell lines have indicated that chronic exposure of the Beta cell to supraphysiological levels of glucose results in impaired Beta cell function, an important component in the pathogenesis of type 2 diabetes. How glucose exerts this toxicity upon the Beta cell remains unclear. We propose that the glucose metabolite, glucosamine plays a role in Beta cell function and this glucose toxicity. We came to this hypothesis as a result of our observations on the mechanism of toxicity of the Beta cell- specific toxin, streptozotocin (STZ). STZ is chemically analogous to N- acetylglucosamine (GlcNAc). Furthermore, we found that the Beta cell contains approximately 100-fold more of the mRNA encoding the enzyme o- GlcNAc transferase (OGT). This enzyme modifies nuclear and cytoskeletal proteins by linking the monosaccharide GlcNAc to serine or threonine residues in the protein. The resulting O-GlcNAc modification appears to modify the activity of transcription factors. We found that STZ blocks the activity of an enzyme that removes o-GlcNAc from proteins. Treatment of rats with STZ results in the accumulation of the O-GlcNAc modification specifically in the pancreatic Beta cells, hours before Beta cell death. Because the Beta cells are so richly endowed with OGT, these cells may be the most susceptible to an accumulation of O-GlcNAc on intracellular proteins. We have also shown in other cell types, that nuclear O-GlcNAc is sensitive to ambient glucose concentrations. If this is also true in the Beta cell, then hyperglycemia and STZ may both increase Beta cell O-GlcNAc content, thereby leading to a common mechanism of Beta cell toxicity. The experiments proposed in this grant are designed to determine if hyperglycemia indeed result in increased Beta cell O-GlcNAc. We will also create transgenic mouse models in which glucosamine synthesis from glucose is either augmented or decreased. We will determine the effect of these alterations in glucosamine metabolism on Beta cell function. We have also found that cAMP-dependent protein kinase inhibits the enzyme responsible for glucosamine synthesis. We propose to study the mechanism by which glucosamine synthesis is inhibited. Together, these studies will establish the role of glucosamine in glucose toxicity on the Beta cell and a means of controlling glucose metabolism to glucosamine.