EARLY BBB DISRUPTION AND HEMORRHAGIC TRANSFORMATION IN TPA THROMBOLYSIS

这个子项目是许多利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 目标和成果 组织纤溶酶原激活剂（tPA）溶栓治疗是FDA批准的唯一治疗急性缺血性卒中的方法，主要由于症状性出血性转化（HT）风险增加（10倍），仅限于5%的缺血性卒中患者。严重的血脑屏障（BBB）缺血性损伤被认为是HT发生的前提。然而，关于BBB损伤和HT之间的因果关系，仍有几个关键问题尚未解决。例如，BBB损伤的程度如何与HT的严重程度相关？ 是否存在一个BBB损伤阈值，特别是在中风早期，这预示着随后的HT？导致早期严重BBB损伤的机制是什么？我们最近开发了磁共振成像（MRI）技术，通过评估钆-二乙烯三胺五乙酸（Gd-DTPA）从血液到大脑的内流率，非侵入性和定量测量大鼠血脑屏障损伤。这一技术进步将使我们能够将早期BBB损伤与tPA治疗后的HT相关联。此外，获得的MRI渗透性图也将确定脑卒中早期严重BBB损伤的特定区域，从而指导我们探索这种严重早期损伤的潜在分子机制。我们以前的工作已经表明，基质金属蛋白酶（MMP），特别是MMP-2/9，在缺血性脑中增加，并有助于通过降解紧密连接蛋白occludin和claudin-5的BBB破坏，和tPA可能通过增加缺血诱导的MMP-9上调诱导HT。事实上，这些研究都集中在MMP-2/9的亚急性变化，几乎没有研究过它们在超急性期（中风发作后的最初几个小时内）的变化和在BBB损伤中的作用，此时从头合成不太可能导致MMP-2/9的增加，因为它们需要时间才能在受影响的脑细胞中转录和翻译。在这个项目中，我们建议研究早期血脑屏障破坏及其与缺血性卒中出血性转化的关系。该项目将利用UNM BRaIN中心现有的最先进技术，使用体内缺血性卒中模型（大脑中动脉闭塞，MCAO）和体外缺血模型（氧葡萄糖剥夺，OGD）研究BBB损伤和潜在的分子机制。 我们的方法包括MRI，双光子显微镜，荧光显微镜和其他细胞和分子技术。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Aims and Results Thrombolytic therapy with tissue plasminogen activator (tPA), the only FDA-approved treatment for acute ischemic stroke, is constrained to 5% ischemic stroke patients largely due to the increased (10-fold) risk of symptomatic hemorrhagic transformation (HT). Severe ischemic damage to the blood brain barrier (BBB) has been considered as the prerequisite for the occurrence of HT. However, several key questions remain outstanding about the causal link between BBB damage and HT. For example, how does the degree of BBB damage correlate to the severity of HT? Is there a threshold of BBB damage, particularly at early-stage of stroke, which portends subsequent HT? What are the mechanisms leading to early severe BBB damage? We have recently developed magnetic resonance imaging (MRI) technique to noninvasively and quantitatively measure BBB damage in rats by assessing the influx rate of gadolinium-diethylene-triamine penta-acetic acid (Gd-DTPA) from blood to the brain. This technical advance would allow us to correlate early BBB damage to subsequent HT following tPA treatment. In addition, the obtained MRI permeability map will also define specific regions with severe BBB damage at early-stage of stroke, and thus guide us to explore the underlying molecular mechanisms for this severe early damage. Our previous work has shown that matrix metalloproteases (MMPs), in particular MMP-2/9, are increased in the ischemic brain and contribute to BBB disruption by degrading tight junction protein occludin and claudin-5, and tPA may induce HT via augmenting ischemia-induced MMP-9 up-regulation. As a fact, these studies have focused on the subacute changes of MMP-2/9, and little has been studied about their changes and roles in BBB damage at hyperacute stage (within the first few hours after stroke onset), when de novo synthesis is not likely responsible for the increase of MMP-2/9 because it takes time for them to be transcribed and translated in affected brain cells. In this project, we propose to study early BBB disruption and its relation to hemorrhagic transformation in ischemic stroke. The project will utilize state of the art technology available in UNM BRaIN center to study BBB damage and the underlying molecular mechanisms using in vivo ischemic stroke model (middle cerebral artery occlusion, MCAO) and in vitro ischemic model (oxygen glucose deprivation, OGD). Our approaches include MRI, two-photon microscopy, fluorescence microscopy, and other cellular and molecular techniques.