INTRACELLULAR O GLCNAC AND GLUCOTOXICITY

细胞内 O GLNAC 和葡萄糖毒性

• 批准号: 2906369
• 负责人: Jeffrey E Kudlow
• 金额: $ 22.78万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-30 至 2001-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2906369
• 关键词: N acetylglucosamine; acyltransferase; alloxan; animal tissue; cytotoxicity; gene expression; genetically modified animals; glucosamine; glucose metabolism; glycosylation; histopathology; hyperglycemia; immunocytochemistry; in situ hybridization; laboratory mouse; laboratory rat; noninsulin dependent diabetes mellitus; northern blottings; pancreatic islet function; pancreatic islets; posttranslational modifications

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.

临床研究和对分离胰岛或 Beta 细胞系的研究 表明β细胞长期暴露于超生理状态 葡萄糖水平过高会导致β细胞功能受损，而β细胞功能是重要的 2 型糖尿病发病机制的组成部分。 葡萄糖如何发挥作用 这种对β细胞的毒性仍不清楚。 我们建议 葡萄糖代谢物、葡萄糖胺在 Beta 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我们 β细胞特异性毒性机制的观察 毒素，链脲佐菌素（STZ）。 STZ 的化学性质类似于 N- 乙酰氨基葡萄糖（GlcNAc）。 此外，我们发现β细胞 含有大约 100 倍多的编码酶 o-的 mRNA GlcNAc 转移酶 (OGT)。 该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来形成蛋白质 蛋白质中的残留物。 由此产生的 O-GlcNAc 修饰出现 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 治疗大鼠会导致 O-GlcNAc 的积累 特定于胰腺β细胞的修饰，几小时前 β细胞死亡。 因为 Beta 细胞富含 OGT， 这些细胞可能最容易受到 O-GlcNAc 积累的影响 关于细胞内蛋白质。 我们还在其他细胞类型中表明， 核 O-GlcNAc 对环境葡萄糖浓度敏感。 如果β细胞也是如此，那么高血糖和STZ可能 两者都会增加 Beta 细胞 O-GlcNAc 含量，从而导致共同的 β细胞毒性机制。 本文提出的实验 赠款旨在确定高血糖是否确实导致 增加β细胞O-GlcNAc。 我们还将创建转基因小鼠 从葡萄糖合成葡萄糖胺的模型要么被增强 或减少。 我们将确定这些改变的影响 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。我们建议研究其机制 葡萄糖胺合成受到抑制。 这些研究共同将 确定葡萄糖胺在β细胞葡萄糖毒性中的作用 以及控制葡萄糖代谢为葡萄糖胺的方法。