Role of Aldose Reductase in Diabetic Complications

描述（由申请人提供）：多元醇途径酶醛糖还原酶（AR）与糖尿病的几种多营养性并发症有关。在动物模型中，AR抑制剂（ARI）可预防或延缓多种糖尿病并发症。然而，ARI的临床疗效仍不确定，AR的生理作用也不清楚。我们在过去资助期间的结果表明，疏水性醛，如脂质过氧化产生的醛，被AR还原的效率高于葡萄糖；提高了AR通常参与脂质衍生醛解毒和减少氧化应激的可能性。我们的中心假设是，通过改变细胞氧化还原状态，并诱导跨国修饰，长期糖尿病扰乱AR的抗氧化作用，因此它有助于而不是对抗高糖的氧化作用。为了验证这一假设，我们将研究人类晶状体上皮细胞（HLEC）、血管平滑肌细胞（VSMC）和血管内皮细胞（VEC）在高血糖和正常血糖条件下AR的翻译后变化。基于我们的研究结果表明AR可以在体外被亚硝化或谷胱甘肽化，我们将确定一氧化氮（NO）合成、亚硝基硫醇和NO供体的增加如何影响细胞AR活性（目的1）。通过免疫沉淀物的电喷雾质谱，我们将确定暴露于高血糖或糖尿病的细胞或组织中AR的共价变化（目的2）。AR在调节内源性氧化应激中的作用将在肿瘤坏死因子- α (tnf - α)诱导的信号传导的背景下进行研究，该信号在糖尿病中增加并由活性氧介导。我们将测试AR的抑制是否会阻止tnf - α对NF-kappaB的激活，从而导致tnf - α诱导的VSMC生长和HLEC或VEC凋亡的减少（Aim 3）。我们将研究ARI是否会阻止高糖对NF-KappaB的基础激活，以及它们是否会阻止高糖对tnf - α的有丝分裂和凋亡作用（Aim 4）。这些研究将有助于确定ARI预防糖尿病并发症的新机制，并将更好地了解AR在正常血糖条件下的调节和作用。我们的研究也将有助于评估抗ar治疗的长期风险或益处。

DESCRIPTION (provided by applicant): The polyol pathway enzyme aldose reductase (AR) has been implicated in several pleiotrophic complications of 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 past funding period show that hydrophobic aldehydes, such as those derived from lipid peroxidation are reduced more efficiently by AR than is glucose; raising the possibility that AR normally participates in the detoxification of lipid-derived aldehydes, and in diminishing oxidative stress. Our central hypothesis is that by altering the cellular redox state, and inducing post-transnational modifications, prolonged diabetes perturbs the antioxidant role of AR, so that it contributes to, rather than combats, the oxidative effects of high glucose. To test this hypothesis, we will investigate the post-translational changes in AR under hyperglycemic vs. normoglycemic conditions in human lens epithelial cells (HLEC), vascular smooth muscle cells (VSMC) and vascular endothelial cells (VEC). Based on our results showing that AR could be nitrosated or glutathiolated in vitro, we will determine how an increase in nitric oxide (NO) synthesis, nitrosothiols, and NO donors affects cellular AR activity (Aim 1). By electrospray mass spectroscopy of immunoprecipitates, we will identify covalent changes in AR in cells or tissues exposed to hyperglycemia or diabetes (Aim 2). The role of AR in regulating endogenous oxidative stress will be examined within the context of tumor necrosis factor-alpha (TNF-alpha)-induced signaling, which is increased in diabetes and is mediated by reactive oxygen species. We will test whether inhibition of AR prevents the activation of NF-kappaB by TNF-alpha and so leads to a decrease in TNFalpha-induced VSMC growth and HLEC or VEC apoptosis (Aim 3). We will examine whether the basal activation of NF-KappaB by high glucose is prevented by ARI, and whether they prevent the mitogenic and apoptotic effects of TNF-alpha in high glucose (Aim 4). These studies should lead to the identification of a novel mechanism by which ARI prevents diabetic complications, and will provide a better understanding to the regulation and the role of AR in normoglycemic conditions. Our investigations will also help in evaluating the long-term risks or benefits of anti-AR therapy.