Mechanisms and Treatment of CNS Edema

描述（由申请人提供）：中风是美国第三大死亡原因和第一大残疾原因，但绝大多数病例没有有效的治疗方法。目前的治疗方案仅仅旨在恢复血液流动，希望挽救处于危险中的组织，但没有提供有针对性的神经保护。由于缺乏对缺血性卒中后继发性损伤至关重要的信号通路的了解，神经保护疗法的发展受到阻碍。十多年来，死亡蛋白酶caspase家族一直与脑缺血和神经变性有关。最近的证据表明，不同的半胱天冬酶途径在缺血时被激活。我们已经确定caspase-9/-6通路与缺血后神经元功能障碍和死亡有关。我们的数据表明，靶向caspase-9活性可在缺血性损伤后提供实质性的神经保护。此外，我们发现caspase-9活性在缺血性发病机制的两个方面是必需的：1)神经元变性和2)脑水肿的发展。水肿是由血管完整性丧失引起的，而不是血管内皮细胞和周细胞（BV）的死亡。这些细胞之间紧密连接的消除使液体从颅内小bv外渗。水肿的形成是严重中风死亡和残疾的主要原因。药物治疗和外科减压手术只能最小限度地改变这种致病过程的自然历史。在我们的研究中，细胞渗透的caspase-9抑制剂Pen1-XBIR3可以降低caspase-9活性，同时消除水肿。这一发现开启了caspase-9活性是否通过损害小脑bv血管完整性而直接导致水肿的问题。我们的初步数据表明，活性caspase-9通过降低基质金属蛋白酶9 （MMP-9）的表达来调节水肿。我们的数据还显示，成熟NGF前体proNGF的表达在中风期间增加。ProNGF是p75NTR的高亲和力配体，我们已经证明通过p75NTR的信号激活caspase-9。p75NTR在脑内的小BVs中发现，p75NTR的表达在中风期间增加。我们的初步数据还显示，活化的caspase-9存在于bv中，caspase-9的抑制可以阻止脑卒中诱导的MMP-9的表达。我们现在提出的假设是，脑卒中水肿的发展是由前eurotrophin （proNT）信号通过p75NTR介导的，该信号激活小bv中的caspase-9来切割对血管完整性至关重要的底物。我们将利用体内和体外模型来检验这一假设，其具体目的如下：目的1：确定诱导proNTs是否会触发小bv中caspase-9的激活。目的2：确定p75NTR信号是否激活caspase-9并导致水肿。目的3：确定caspase-9切割底物如何导致血管完整性丧失。

DESCRIPTION (provided by applicant): Stroke is the 3rd largest cause of death and the largest cause of disability in the U.S., yet there are no effective therapies for the vast majorityof cases. Present therapeutic options merely aim to restore blood flow in the hopes of salvaging at risk tissue, but offer no targeted neuroprotection. Development of neuroprotective therapies has been hindered by lack of knowledge of the signaling pathways critical in secondary injury following ischemic stroke. For more than a decade, the caspase family of death proteases has been implicated in cerebral ischemia and neurodegeneration. Recent evidence shows that distinct caspase pathways are activated during ischemia. We have identified the caspase-9/-6 pathway as responsible for neuronal dysfunction and death after ischemia. Our data show that targeting caspase-9 activity provides substantial neuroprotection following an ischemic insult. Moreover, we find that caspase-9 activity is required for two aspects of ischemic pathogenesis: 1) neuronal degeneration and 2) the development of cerebral edema. Edema is caused by a loss of vascular integrity, rather than death of endothelial cells and pericytes in the blood vessels (BV). The elimination of tight junctions between these cells allows extravasation of fluid from small intracranial BVs. Edema formation is a major contributor to death and disability in severe stroke. Medical therapies and surgical decompressive procedures have only minimally altered the natural history of this pathogenic process. In our studies, a cell permeant caspase-9 inhibitor, Pen1-XBIR3, reduces caspase-9 activity and concomitantly abolishes edema. This finding opens the question of whether caspase-9 activity is a direct cause of edema through the impairment of vascular integrity of small cerebral BVs. Our preliminary data suggest that active caspase-9 regulates edema by decreasing the expression of matrix metalloproteinase 9 (MMP-9). Our data also show that expression of the precursor of mature NGF, proNGF, increases during stroke. ProNGF is a high affinity ligand for p75NTR, and we have shown that signaling through p75NTR activates caspase-9. p75NTR is found in small BVs in the brain, and expression of p75NTR increases during stroke. Our preliminary data also show that activated caspase-9 is present in BVs, and that caspase-9 inhibition prevents the stroke-induced expression of MMP-9. We now propose the hypothesis that the development of edema in stroke is mediated by proneurotrophin (proNT) signaling through p75NTR, which activates caspase-9 in small BVs to cleave substrates vital to the integrity of the vessels. We will utilize in vivo an in vitro models to examine this hypothesis with the following Specific Aims: Aim 1: To determine if induction of proNTs triggers caspase-9 activation in small BVs. Aim 2: To determine if signaling via p75NTR activates caspase-9 and leads to edema. Aim 3: To determine how caspase-9 cleavage of substrates leads to loss of vascular integrity.