Glycation of Protein in Diabetes

描述（由申请人提供）：本研究计划阐述了非酶糖化和美拉德或布朗宁反应对蛋白质和其他生物分子的损伤有助于糖尿病长期并发症发病机制的假设。提出的机制涉及形成不可逆的晚期糖基化和脂氧化终产物（AGE/ALE），其影响蛋白质结构和功能，并改变并发症发生的组织中的代谢。我们已经通过评估AGE/ALE抑制剂对蛋白质的化学修饰和糖尿病动物模型（包括链脲佐菌素糖尿病大鼠和Zucker肥胖大鼠）中并发症的发展的影响来测试Maillard假说。这些研究提出了关于AGE/ALE抑制剂对蛋白质的酶交联的影响的问题，导致鉴定了一类新的来源于半胱氨酸的AGE/ALE，并对AGE/ALE抑制剂吡哆胺（PM）的羰基捕获活性和作用机制有了深入的了解。在继续资助期间，我们的具体目标是：1）评估糖尿病、高脂血症和AGE/ALE抑制剂对胶原蛋白中酶促和非酶促化学修饰和交联形成的影响，包括AGE/ALE抑制剂体内螯合活性的研究; 2）研究Cys-AGE/ALE在糖尿病和高脂血症中的形成及其生物学意义，包括蛋白质巯基与乙二醛、丙酮醛和延胡索酸反应的基础研究，比较细胞内和细胞外蛋白质中Cys-AGE/ALE的水平，AGE/ALE抑制剂处理的效果，鉴定Cys-AGE/ALE修饰的主要细胞内蛋白质，以及评估Cys-AGE/ALE形成的调节意义; 3）继续对PM进行研究，以在体外和体内鉴定由该化合物捕获的活性中间体，评价PM对血浆中二羰基中间体水平的影响，并评价PM对非高血糖动物模型（糖尿病倾向BB/Wor大鼠）中AGE/ALE形成和病理发展的影响。通过这项工作，我们希望能够更好地了解AGE/ALE在糖尿病并发症发病机制中的作用以及AGE/ALE抑制剂的作用机制，最终开发出更有效的治疗糖尿病及其并发症的疗法。

DESCRIPTION (provided by applicant): This research program addresses the hypothesis that damage to proteins and other biomolecules by nonenzymatic glycation and Maillard or browning reactions contributes to the pathogenesis of the long-term complications of diabetes. The proposed mechanism involves formation of irreversible advanced glycation and lipoxidation end-products (AGE/ALEs) that affect protein structure and function and alter metabolism in tissues in which complication develop. We have tested the Maillard hypothesis by evaluating the effects of AGE/ALE inhibitors on the chemical modification of proteins and development of complications in animal models of diabetes, including the streptozotocin-diabetic and Zucker obese rat These studies have raised questions about the effects of AGE/ALE inhibitors on enzymatic crosslinking of proteins, have led to the identification of a new class of AGE/ALEs derived from cysteine, and have yielded insight into the carbonyl trapping activity and mechanism of action of the AGE/ALE inhibitor pyridoxamine (PM). During the continuation period of this grant, our Specific Aims are: 1) to evaluate the effects of diabetes, hyperlipidemia and AGE/ALE inhibitors on the formation of enzymatic, as well as nonenzymatic, chemical modifications and crosslinks in collagen, including studies on the chelating activity of AGE/ALE inhibitors in vivo; 2) to study the formation and biological significance of Cys-AGE/ALEs in diabetes and hyperlipidemia, including basic research on the reaction of protein sulfhydryl groups with glyoxal, methylglyoxal and fumarate, comparison of levels of Cys-AGE/ALEs in intracellular and extracellular proteins, effects of treatment with AGE/ALE-inhibitors, identification of major intracellular proteins modified by Cys-AGE/ALEs, and evaluation of the regulatory significance of Cys-AGE/ALE formation; 3) to continue studies on PM in order to identify reactive intermediates trapped by this compound in vitro and in vivo, to evaluate effects of PM on levels of dicarbonyl intermediates in plasma, and to evaluate the effects of PM on AGE/ALE formation and development of pathology in a non-hyperlipidemic animal model, the diabetes prone BB/Wor rat. Through this work, we hope to gain a better understanding of the role of AGE/ALEs in the pathogenesis of diabetic complication and the mechanism of action of AGE/ALE inhibitors, leading eventually to the development of more effective therapies for treatment of diabetes and its complications.