Cortical spreading depression, proteases and ischemia

皮质扩散性抑制(CSD)在实验性卒中期间以及在正常和 受创伤的人脑。近年来，基质金属蛋白酶通过降解基质蛋白、增强血脑屏障通透性和脑水肿、促进tPA治疗后出血等作用，参与了卒中的发病机制和神经血管单位的完整性。虽然脑损伤或血管机制提供了触发因素，但我们最近发现，在单个CSD过程中，强烈的神经元和神经胶质细胞去极化至少会激活MMP9 72小时，并促进血管周围血浆蛋白的泄漏。我们提出的4个目的是为了探索这一新的假设，即CSD期间MMPs的激活部分是由神经元和神经胶质细胞的激活直接引起的，并参与了正常脑缺血时的水肿和血脑屏障的破坏。目标1将确认和扩展初步数据建立 CSD作为基质金属蛋白酶激活的触发物，并将确定相关的异构体和起源细胞。目的2验证CSD伴有基质金属蛋白酶(MMPs)活化引起的血浆蛋白渗漏的假说，并将扩展的初步数据显示：(1)8kD葡聚糖和伊文思蓝在同侧血管内通透性增加，(2)内皮细胞Hard抗原和层粘连蛋白的抗原性在CSD后通过依赖于MMP的机制降低。AIM 3将研究在慢性阻塞性肺疾病期间触发基质金属蛋白酶激活的上游机制。我们还将 检测NFkappaB通路在CSD诱导的基质金属蛋白酶转录调控中的重要性，以及TIMP-1作为CSD后基质金属蛋白酶-9活性调节因子的表达。目的4研究CSD诱导的基质金属蛋白酶在缺血时激活的后果，并探讨基质金属蛋白酶在缺血损伤和远隔脑区血管通透性改变中的作用。利用缺乏表达在白细胞上的基质金属蛋白酶-8的突变小鼠，我们建议确定基质金属蛋白酶-8导致CSD诱导的缺血和非缺血组织中血脑屏障破坏的程度，并通过在白细胞中依赖于基质金属蛋白酶-8的机制来检测CSD诱导的基质金属蛋白酶-9激活的可能性。通过这样做，我们打算探索如何 脑缺血时神经元的活动调节细胞外基质和神经血管单位，从而导致组织损伤。

Cortical spreading depression (CSD) develops during experimental stroke and within normal and traumatically-injured human brain. Recently, matrix metalloproteinases (MMP) have been implicated in stroke pathogenesis and in the integrity of the neurovascular unit by degrading matrix proteins, enhancing blood brain barrier (BBB) permeability and brain edema, and promoting hemorrhage after tPA administration. Although brain injury or vascular mechanisms provide triggers, we recently found that intense neuronal and glial depolarization during a single CSD activates MMP-9 for at least 72 hr and promotes perivascular plasma protein leakage. We propose 4 aims to explore the novel hypothesis that MMP activation during CSD is caused in part by neuronal and glial activation directly and contributes to the development of edema in ischemia and BBB disruption in normal brain. Aim 1 will confirm and extend preliminary data establishing CSD as a trigger for MMP activation and will identify relevant isoforms and cells of origin. Aim 2 will test the hypothesis that CSD is accompanied by leakage of plasma protein caused by MMP activation and will extend preliminary data showing that, (1) permeability of 8kD dextran and Evans blue increases within ipsilateral vessels and that, (2) antigenicity of endothelial harder antigen and laminin is decreased by MMP-dependent mechanisms after CSD. Aim 3 will examine upstream mechanisms triggering MMP activation during CSD. Because nitric oxide (NO) S-nitrosylates the MMP active site to promote activation, we will examine whether CSD-induced NO generation triggers S-nitrosylated MMP activation and if so, examine the relevant NOS isoform promoting MMP activation by using selective NOS knockout mice. We will also examine the importance of NFkappaB pathway in the transcriptional regulation of CSD-induced MMP and the expression of TIMP-1, as a regulator of MMP-9 activity after CSD. Aim 4 will examine the consequences of CSD-induced MMP activation during ischemia and explore the contributions of MMPs to vascular permeability changes both within the ischemic lesion and in remote brain areas. Using mutant mice lacking MMP-8 expressed on leukocytes, we propose to determine the extent to which MMP-8 causes CSD-induced BBB disruption in ischemic and non-ischemic tissue, and examine the potential for CSD-induced MMP-9 activation by MMP-8 dependent mechanisms in leukocytes. By so doing, we intend to explore ways in which neuronal activity during cerebral ischemia modulates the extracellular matrix and neurovascular unit and contribute to tissue injury.