Role of Aldose Reductase in Diabetic Complications

多元醇途径酶醛糖还原酶（AR）参与了几种多效性并发症， 糖尿病在动物模型中，AR抑制剂（ARI）可预防或延迟多种糖尿病并发症。但 ARI的临床疗效仍然不确定，AR的生理作用也不清楚。我们的结果在当前 资助期间显示AR催化的脂质过氧化物-谷胱甘肽缀合物的还原产物（如GS-DHN， 在高血糖诱导的氧化应激下形成的谷胱甘肽二羟基壬烷）介导NF-κ B和API 激活，增加炎症标志物。我们已经证明，亚硝化激活和谷胱甘肽 禁用AR。我们的中心假设是，通过改变细胞的氧化还原状态，并诱导翻译后 由于这些修饰，长期的糖尿病扰乱了氧化还原平衡和应激信号传导，导致细胞因子的增加。 产生和炎症，其又诱导或加剧继发性糖尿病并发症。为了验证这一 假设，我们将扩大研究，以了解高血糖症和高血糖症之间的机制关系， 炎症和确定AR在细胞因子产生和炎症中的作用。具体来说，我们的目标是继续 我们的研究旨在进一步了解AR介导高血糖诱导的 PKC和TACE引起TNF-α分泌，导致平滑肌细胞增生，血管内皮细胞 细胞凋亡、炎症和胰岛素抵抗。因此，目的是扩大目前的赠款。完成 我们的目标是验证我们的假设，并确定AR介导高血糖的机制- 诱导炎症信号，导致继发性糖尿病并发症，包括胰岛素抵抗。结果也是 对AR进行更多的结构研究将有助于我们开发更特异和靶向的AR抑制剂。因此 未来五年的目标是：（1）研究降脂肽-谷胱甘肽的作用机制 高血糖症中的缀合物（如GS-DHN）激活PKC和NF-κ B并触发炎症;（2）描绘高血糖症中的 AR在高血糖期间调节TNF-α产生中的作用;（3）鉴定由AR-α激活的蛋白激酶。 催化的还原型脂质过氧化物-谷胱甘肽缀合物（GS-DHN）磷酸化PKC;和（4）开发特异性 和靶向醛糖还原酶抑制剂。根据我们最近的AR-NADPH-谷胱甘肽类似物的晶体结构， AR谷胱甘肽结合位点的三元复合物和生化分析，分子模拟，定点 将进行突变以进一步探测谷胱甘肽结合位点和AR与谷胱甘肽之间相互作用的性质。 谷胱甘肽结合物。这将有助于开发基于结构的AR抑制剂，其将阻止 GS-HNE不影响毒性脂质醛如HNE的结合和还原。这种方法 在最大限度地减少AR抑制剂的毒性方面具有潜在的重要性，从而大大提高了治疗应用 AR抑制剂。

The polyol pathway enzyme aldose reductase (AR) has been implicated in several pleiotrophic complicationsof diabetes. In animal models, AR inhibitors (ARI) prevent or delay multiple diabetic complications. However, the clinical efficacy of ARI remains uncertain, and the physiological role of AR is unclear. Our results during the current funding period show that AR-catalyzed reduced products of lipid aldehydes-glutathione conjugates (such as GS-DHN, glutathione dihydroxynonane), formed under hyperglycemia-induced oxidative stress, mediate NF-KB and API activation that increases inflammatory markers. We have shown that nitrosation activates and glutathiolation inactivates AR. Our central hypothesis is that by altering the cellular redox state, and inducing post-translational modifications, prolonged diabetes perturbs the redox poise and stress signaling leading to an increase in cytokine production and inflammation which in turn induces or exacerbates secondary diabetic complications. To test this hypothesis, we will extend the studies to understand the mechanistic relationship between hyperglycemia and inflammation and identify the role of AR in cytokine production and inflammation. Specifically our aim is to continue our investigations to further understand the mechanisms by which AR mediates hyperglycemia-induced activation of PKC and TACE that cause TNF-a secretion leading to smooth muscle cell hyperplasia, vascular endothelial cell apoptosis, inflammation, and insulin resistance. Accordingly,the aims are extension of the current grant. Completion of our aims will verify our hypothesis and identify the mechanisms through which AR could mediate hyperglycemia- induced inflammatory signals that cause secondary diabetic complicationsincluding insulin resistance. Also the results of additional structural studies on AR would help us in developing more specific and targeted inhibitor(s) of AR. Thus the aims of the next five years are to: (1) investigate the mechanisms by which reduced lipid aldehydes-glutathione conjugates (such as GS-DHN) in hyperglycemia activate PKC and NF-KB and trigger inflammation; (2) delineate the role of AR in regulating TNF-a production during hyperglycemia; (3) identify protein kinase(s) activated by AR- catalyzed reduced lipid aldehydes-glutathione conjugates (GS-DHN) that phosphorylate PKC; and (4) develop specific and targeted aldose reductase inhibitors. Based on our recent crystal structure of the AR-NADPH-glutathione analogue ternary complex and biochemical analysis of the glutathione binding site of AR, molecular modeling, site-directed mutations will be performed to further probe glutathione binding site and the nature of the interaction between AR and glutathione conjugates. This will help in developing structure-based AR inhibitors which will prevent the binding of GS-HNE without affecting the binding and reduction of toxic lipid aldehydes such as HNE. This approach is potentially important in minimizing the toxicity of AR inhibitors thereby greatly improvingthe therapeutic application of AR inhibitors.