Neurovascular Adhesion Receptors and Barrier Integrity

描述(由申请人提供)：缺血性中风会导致微血管完整性的迅速严重丧失。局灶性缺血后早期可检测到脑微血管通透性屏障的破坏，内皮细胞整合素基质迅速丧失。 感受器。在淀粉样血管病中，微血管的改变增加了微出血的风险。这一竞争性更新方案将检验的假设是：i)微血管内皮细胞(和星形胶质细胞)上的基质受体与基底膜的基质成分的相互作用是血脑屏障表型的主要决定因素，ii)局灶性脑缺血破坏受体-基质相互作用，以及iii)受体-基质相互作用的中断导致血脑屏障表型的丧失。我们已经证明，内皮细胞与完整的基底板基质的黏附是屏障完整性的核心。屏障表型(如紧密连接(TJ))和血管基质由内皮细胞和星形胶质细胞共同产生，并由两个细胞室维持。原代培养的小鼠内皮细胞和星形胶质细胞可模拟体内内皮细胞整合素和星形胶质细胞肌营养不良蛋白的表达及其对局灶性缺血的反应。这些反应依赖于矩阵。基于成功完成的工作和初步数据，我们认为1整合素可以决定TJ蛋白的表达，并构成血脑屏障和基质屏障的“垂直”成分。该项目的目标是证明特定的整合素家族成员的作用是微血管屏障完整性所必需的，而这些特定的整合素抑制剂、基因构建和敲除、局灶性缺血和A？1肽破坏了受体-基质的相互作用，导致屏障失效。其具体目的是证明：1)融合内皮上的整合素与基质蛋白的相互作用决定了内皮细胞间的凝聚力，而这些凝聚力是由星形胶质细胞调节的；2)内皮细胞内整合素信号的机制介导了基质黏附受体介导的TJ表达的变化(没有或有星形胶质细胞)；3)实验性缺血通过改变内皮细胞的整合素-基质相互作用来显著改变内皮细胞的TJ表达；以及4)内皮细胞(没有或与星形胶质细胞)暴露。 TO A？-肽通过整合素依赖的机制调节TJ的表达和微血管通透性屏障表型。这些新的研究为局灶性缺血和淀粉样血管病变后微血管完整性的破坏提供了一个可信的解释。内皮细胞1整合素-基质黏附介导的“垂直”成分表明，它的破坏是血脑屏障丧失的前提。了解整合素-基质黏附的机制可能会导致新的可测试的方法来保存或选择性地改变其他神经血管疾病的微血管屏障功能。 与公共卫生相关：缺血性中风是一种破坏性的疾病，它破坏脑部小血管衬里细胞(微血管内皮细胞)之间的重要联系，这些细胞被认为可以防止液体从血管内泄漏，导致血液泄漏和脑肿胀。我们最近发现，将内皮细胞与它们所在的支架(基质)结合在一起的分子(整合素)是必不可少的 以维持正常的渗漏屏障，而缺血性中风迅速破坏整合素，从而导致渗漏发生。这项持续的研究工作将确定内皮细胞整合素-基质结合维持屏障的确切机制，以及它的丢失如何导致缺血性中风和其他脑血管疾病期间的脑损伤：开发新的中风治疗策略所需的基础研究。

DESCRIPTION (provided by applicant): Ischemic stroke produces rapid profound loss of microvascular integrity. Early following focal ischemia detectable disruption in the permeability barrier of cerebral microvessels occurs, with rapid loss of the endothelial cell ¿1 integrin-matrix receptors. In amyloid angiopathy, microvessel alterations increase the risk of microhemorrhages. The hypotheses to be tested by this competing renewal Proposal state that i) the interaction of matrix receptors on microvessel endothelial cells (and astrocytes) with matri components of the basal lamina are a major determinant of the blood-brain barrier phenotype, ii) focal cerebral ischemia disrupts receptor-matrix interactions, and iii) interruptions of receptor-matrix interactions result in loss of the blood brain barrier phenotype. We have shown that adhesion of endothelial cells to the intact basal lamina matrix is central to the integrity of the barrier. The barrier phenotype (e.g. tight junctions (TJs)) and vascular matrix are generated by endothelial cells and astrocytes in concert, and are maintained by both cell compartments. The expression of ¿1 integrins on endothelial cells and ¿¿-dystroglycan on astrocytes in vivo, and their responses to focal ischemia, are mimicked by primary endothelial cells and astrocytes of murine origin in culture. These responses are matrix-dependent. Based upon successful completed work and preliminary data, we propose that ¿1 integrins can determine TJ protein expression, and constitute the "vertical" component of the blood- brain and matrix barriers. The goal of this Project is to demonstrate that the actions of specific ¿1 integrin family members are required for the integrity of the microvessel barrier, and those specific integrin inhibitors, geneic constructs and knockdowns, focal ischemia, and A¿1 peptides disrupt the receptor-matrix interactions, producing barrier failure. The Specific Aims are to demonstrate that: 1) the interactions of ¿1 integrins on confluent endothelium with matrix proteins determine inter-endothelial cell cohesion by the TJ proteins, and these are modulated by astrocytes, 2) mechanisms of ¿1 integrin signaling within endothelial cells mediate the matrix adhesion receptor-mediated changes in TJ expression (without or with astrocytes), 3) experimental ischemia significantly alters endothelial cell TJ expression by altering the endothelial cell ¿1 integrin-matrix interactions, and 4) the exposure of endothelial cells (without or with astrocytes) to A¿-peptides modulates TJ expression and the microvessel permeability barrier phenotype via ¿1 integrin-dependent mechanisms. These novel studies provide a plausible explanation for the disruption of microvessel integrity immediately following focal ischemia and by amyloid angiopathy. The "vertical" component mediated by endothelial ¿1 integrin-matrix adhesion suggests the premise that its disruption is responsible for loss of the blood-brain barrier. Understanding the mechanisms of the ¿1 integrin-matrix adhesion is likely to lead to new testable approaches to preserve or selectively alter microvessel barrier function in other neurovascular disorders. PUBLIC HEALTH RELEVANCE: Ischemic stroke is a devastating disorder that disrupts important bonds between the cells lining the small blood vessels of the brain (microvessel endothelial cells), that are thought to prevent leakage of fluid from inside the vessels, causing blood leakage and brain swelling. We have recently found that molecules (¿1 integrins) that bind the endothelial cells to the scaffolding they sit on (matrix) within each microvessel are essential for maintaining the normal barrier to leakage, and that ischemic stroke quickly disrupts ¿1 integrins, thereby causing leakage to occur. This continuing research work will define the exact mechanisms by which endothelial cell ¿1 integrin-matrix binding maintains the barrier, and how its loss contributes to brain injury during ischemic stroke and other brain vascular disorders: fundamental studies needed for the development of new treatment strategies for stroke.