Pathogenic Mechanisms Underlying Diabetic Retinopathy

描述(申请人提供)：糖尿病中引起微血管病理性视网膜病变和其他血管并发症的致病过程知之甚少。一些新的证据表明，炎症发挥着迄今未被认识到的作用。视网膜细胞、血管系统和其他自身细胞受到一系列固有的膜调节蛋白的保护，免受自体补体蛋白的攻击。这些调节因子是衰变加速因子(DAF或CD55)、膜辅助因子蛋白(MCP或CD46)和膜反应溶解抑制因子(CD59)。我们实验室以前的研究表明，这三种表面蛋白都在视网膜及其相关血管系统中高表达，表达水平与肾小球细胞和全身血管系统相似，而在这些部位，内在调节活性至关重要。在最近的体外工作中，我们已经证明DAF的调节功能是1)&gt；90%被葡萄糖或核糖孵育，2)被乙二醛孵育迅速取消，3)由于一个或多个活性位点赖氨酸和精氨酸残基被修饰而丧失，后者被修饰为精氨酸。在体内工作中，我们已经证明了从糖尿病视网膜分离的内源性DAF蛋白可以被包括精嘧啶在内的几种糖加合物修饰。建议的研究目标是：1)确定高血糖和其他与糖尿病状态有关的代谢异常引起的DAF、MCP和CD59的化学修饰的功能效应；2)从结构上表征调节因子上的修饰的类型和位置；3)分析从糖尿病患者的视网膜和其他组织中分离出的内源性DAF、MCP和CD59蛋白的功能和结构；以及4)检查与视网膜病变和其他糖尿病并发症相关的病理变化是否在Daf1-/-(小鼠DAF同源基因)、Crry-/-(小鼠MCP替代基因)、CD59a-/-(小鼠CD59同源物)和双Daf1-/-Curry-/-或Daf1-/-/CD59a-/-小鼠实验性地用链脲佐菌素造成糖尿病。由于血管病变和视网膜病变是最终影响大多数糖尿病患者的衰弱并发症，充分了解它们的发生机制对于设计有效的治疗干预措施非常重要。

DESCRIPTION (provided by applicant): The pathogenic processes that give rise to microangiopathic retinopathy and other vascular complications in diabetes are poorly understood. Several new lines of evidence have pointed toward inflammation as playing a heretofore unrecognized role. Retinal cells and vasculature and other self cells are protected from attack by autologous complement proteins by a set of intrinsic membrane regulatory proteins. These regulators are the decay accelerating factor (DAF or CD55), the membrane cofactor protein (MCP or CD46), and the membrane inhibitor of reactive lysis (CD59). Previous studies from our lab have shown that all three surface proteins are highly expressed in the retina and its associated vasculature in levels similar to those on glomerular cells and on systemic vasculature, sites where intrinsic regulatory activity is critical. In recent in vitro work, we have shown that DAF's regulatory function is 1) >90% inactivated by incubation with glucose or ribose, 2) rapidly abrogated by methylglyoxal incubation, and 3) lost due to modification of one or more active site lysine and arginine residues, the latter to argprymidine. In in vivo work, we have shown that endogenous DAF protein isolated from diabetic retinas is modified by several sugar adducts including argpyrimidine. The proposed studies are directed toward 1) determining the functional effects of chemical modifications in DAF, MCP, (and CD59) that are induced by hyperglycemia and other metabolic abnormalities that pertain in the diabetic state, 2) structurally characterizing the types and sites of the modifications on the regulators, 3) analyzing the functions and structures of endogenous DAF, MCP, (and CD59) proteins isolated from retinas and other tissues of diabetics, and 4) examining whether the pathological changes that are associated with retinopathy and other diabetic complications develop more rapidly in Daf1-/- (murine DAF homolog), Crry-/- (murine MCP surrogate), CD59a-/- (murine CD59 homolog) and double Daf1-/- / -/- Curry-/- or Daf1-/- / CD59a-/- mice experimentally made diabetic with streptozotocln. Since vasculopathy and retinopathy are debilitating complications that eventually affect most patients with diabetes, fully understanding the mechanisms involved in their development is important in designing effective therapeutic interventions.