Matrix Metalloproteinases and Neurons in Focal Ischemia

描述(由申请人提供)：缺血性中风是由迅速的深层神经元损伤和微血管完整性丧失引起的。这一建议将检验的假设是，在大脑中动脉闭塞(MCA-O)和再灌注期间，i)神经元的反应部分取决于微血管基膜基质和非血管细胞外基质(ECM)对缺血的反应，ii)阻断导致基膜基质和细胞间ECM变化的特定蛋白酶将挽救微血管系统和神经元免受缺血损伤。根据前人的工作和初步数据，我们认为降解基质硫酸乙酰肝素蛋白多糖(HSPGs)和硫酸乙酰肝素(HSS)的酶是在MCA：O之后产生的，与基质金属蛋白酶Pro-MMP2同时产生，并与微血管和神经元损伤有关。本项目的目标是证明特定的基质降解酶在局灶性脑缺血期间微血管完整性和神经元存活中发挥重要作用。其具体目的是：1)定位缺血后改变微血管基质中含有基质的HSPG和肝素硫酸盐(包括部分组织蛋白酶和肝素酶)的蛋白酶的表达，并将它们的出现与前-MMP-2的表达、基底膜和细胞间ECM的完整性以及神经元存活联系起来；2)证明早期再灌注减少HSPG和HS蛋白的表达，从而恢复微血管内皮细胞和星形胶质细胞整合素、基底板、细胞间ECM和神经元的完整性；3)证明抑制HSPG和HS降解酶的活性可以挽救微血管内皮细胞和星形胶质细胞整合素、基底板、细胞间ECM和神经元的完整性；4)抑制基质金属蛋白酶-2或选择性组织蛋白酶和肝素酶活性，可减少基质热休克蛋白和HS的降解，缩小脑梗塞体积。 HSPG/HS降解酶在微血管完整性丧失和神经元损伤中的作用的研究是对这一提议正在进行的研究的延伸，也是卒中研究的一个新方向。他们预计将大大增加我们对微血管基质和大脑中神经元存活之间的关系的理解。他们还提出了一个新的前提，即在局灶性缺血期间，神经元损伤部分是通过特定敏感基质成分的变化来介导的，这些基质成分维持微血管的完整性并支持神经元的存活。这些研究可能导致新的可测试的治疗方法来保护微血管和神经元功能，这将与其他血管神经元退行性疾病相关。

DESCRIPTION (provided by applicant): Ischemic stroke results from rapid profound neuron injury and loss of microvascular integrity. The hypotheses to be tested by this Proposal state that during middle cerebral artery occlusion (MCA-O) and reperfusion, i) neuron responses are determined, in part, by the responses to ischemia of the microvessel basal lamina matrix and the nonvascular extracellular matrix (ECM), and ii) blocking specific proteases responsible for these changes in the basal lamina matrix and intercellular ECM will salvage the microvasculature and neurons from ischemic injury. Based upon previous work and preliminary data, we propose that enzymes which degrade matrix heparan sulfate proteoglycans (HSPGs) and heparan sulfates (HSs) are generated following MCA:O, simultaneously with the matrix metalloproteinase pro-MMP-2, and are responsible for microvessel and neuron injury. The goal of this Project is to demonstrate that specific matrix-degrading enzymes play significant roles in microvessel integrity and neuron survival during focal cerebral ischemia. The Specific Aims are to: 1) Localize the postischemic expression of proteases that alter matrix-containing HSPGs and heparan sulfates (including select cathepsins and heparanase) in the microvessel matrix, and relate their appearance to pro-MMP-2 expression, basal lamina and intercellular ECM integrity, and to neuron survival; 2) Demonstrate that early reperfusion reduces expression of HSPG and HS proteases, and thereby recovers microvessel endothelial cell and astrocyte integrins, basal lamina, intercellular ECM, and neuron integrity; 3) Demonstrate that inhibiting HSPG and HS-degrading protease activities can salvage microvessel endothelial cell and astrocyte integrins, basal lamina, intercellular ECM, and neuron integrity; and, 4) Demonstrate that inhibition of MMP-2 or selected cathepsin and heparanase activities decreases matrix HSPG and HS degradation, and reduces the volume of cerebral infarction. Studies of the contributions of HSPG/HS-degrading proteases to the loss of microvessel integrity and neuron injury represent an extension of the ongoing studies of this Proposal, and a new direction in stroke research. They are expected to add substantially to our understanding of the relationships between microvessel matrix and neuron survival in the brain. They also suggest the novel premise that during focal ischemia, neuron injury is mediated in part by changes in specific sensitive matrix components which maintain microvessel integrity and support neuron viability. These studies are likely to lead to new testable therapeutic approaches to preserve mlcrovascular and neuron function which will have relevance to other vascular neuron degenerative disorders.