PKC Isoenzymes and Diabetic Microvascular Hyperpermeability

微血管通透性过高是导致血管内皮细胞损伤的重要原因之一。 许多炎症性疾病包括糖尿病并发症。我们研究计划的长期目标是 了解微血管屏障功能调节的细胞和分子机制 在生理和病理条件下。作为该项目的一个组成部分，该项目 旨在阐明负责微血管的信号通路和分子机制， 糖尿病是一种影响大量高渗透性人群的疾病， 以微血管损伤为特征的并发症导致的发病率和死亡率。我们的中央 一种假说认为，糖尿病通过以下途径在多个水平上调微血管内皮细胞中的PKC β， MAPK刺激的基因表达、PDK 1增强的催化活性和DAG介导的激酶活化。 我们进一步提出，上调的PKC β增加了微静脉内皮细胞的细胞旁通透性 通过与细胞间连接偶联的GDI-RhoA-ROCK级联诱导内皮细胞收缩 由β-连环蛋白磷酸化和VE-钙粘蛋白解离引发的组织解体。三个具体目标 提出：1）明确确定PKC β在糖尿病微血管渗漏中的作用; 2） 表征糖尿病猪中PKC β上调的信号传导机制;和3）阐明糖尿病猪中PKC β上调的信号传导机制。 PKC β引起微血管通透性增高的分子基础。这些目标将会实现 通过多方面的分子生理学方法，将分子技术与功能 微血管水平的分析。人类相关的猪糖尿病模型将作为主要模型 用于完整微血管中内皮屏障功能的定量评估。本研究数据 将为糖尿病微血管并发症的发病机制提供新的见解。识别 负责PKC诱导的终点损伤的精确分子机制可能导致一种新的途径， 寻找治疗靶点在此基础上，提出了今后的研究方向 用于诊断和治疗微血管渗漏的分子探针和疗法的发展 与慢性炎症性疾病有关。

Microvascular hyperpermeability represents an important injurious process underlying the development of many inflammatory diseases including diabetic complications. The long-term goal of our research program is to understand the cellular and molecular mechanisms in the regulation of microvascular barrier function under physiological and pathological conditions. As an integral component of the program, this project is designed to elucidate the signaling pathways and molecular mechanisms responsible for microvascular hyperpermeability during development of diabetes, a disease that affects a large population with high morbidity and mortality resulting from complications characterized by microvascular injury. Our central hypothesis states that diabetes upregulates PKCbeta in microvascular endothelium at multiple levels via MAPK-stimulated gene expression, PDK1-potentiated catalytic activity, and DAG-mediated kinase activation. We further propose that upregulated PKCbeta increases the paracellular permeability of venular endothelium by inducing endothelial cell contraction via the GDI-RhoA-ROCK cascade coupled with intercellular junction disorganization triggered by beta-catenin phosphorylation and VE-cadherin dissociation. Three specific aims are proposed: 1) to unequivocally establish the role of PKCbeta in microvascular leakage during diabetes; 2) to characterize the signaling mechanisms of PKCbeta upregulation in diabetic pigs; and 3) to elucidate the molecular basis of PKCbeta-elicited microvascular hyperpermeability. These aims will be accomplished through a multifaceted molecular physiology approach that incorporates molecular techniques with functional analyses at the microvascular level. A human-relevant pig model of diabetes will serve as the primary model for quantitative assessment of endothelial barrier function in intact microvessels. Data derived from this study will provide new insights into the pathogenesis of diabetic microvascular complications. Identification of the precise molecular mechanisms responsible for PKC-induced end-point injury may lead to a new avenue for searching therapeutic targets. Based on this study, a future direction of our research efforts will be directed to the development of molecular probes and therapies for diagnosis and treatment of microvascular leakage associated with chronic inflammatory diseases.