Fibrotic effects and regulation of MMP proteins in thrombus resolution

描述(由申请人提供)： 深静脉血栓形成(DVT)每年影响200多万美国人。静脉炎后综合征可影响25%-75%的DVT患者，包括腿部肿胀、疼痛、皮肤变化和皮肤溃疡。这只发生在DVT患者的一小部分中，这表明血栓溶解对于确定慢性静脉阻塞和纤维化是否会发展并导致静脉炎后综合征至关重要。基质金属蛋白酶基因是细胞迁移和组织重塑的关键基因，在血栓溶解过程中被表达和激活，提示在这一过程中起着关键作用。它们在静脉炎后综合征的发病机制中的作用有三个特定的目的：1)确定与血栓诱导的MMP-2表达有关的MMP-2基因及其同源转录因子的区域；2)确定基质金属蛋白酶-2、9和14(MT-基质金属蛋白酶-1)在深静脉血栓消退和血栓诱导的静脉壁纤维化中的作用；3)确定基质金属蛋白酶蛋白(基质金属蛋白酶-2、基质金属蛋白酶-9和基质金属蛋白酶-14)的过表达是否加速深静脉血栓的溶解并改变血栓诱导的静脉壁纤维化。一系列独特的转基因基质金属蛋白酶-2报告小鼠将被用来确定基质金属蛋白酶-2基因的哪些区域是血栓诱导的基质金属蛋白酶-2转录所必需的，染色质免疫沉淀将被用来鉴定同源转录因子。各种基质金属蛋白酶基因定向缺失的小鼠将被用来确定这些酶在血栓再通、静脉壁纤维化以及静脉壁顺应性和弹性丧失中的作用，方法是采用新的小鼠静脉生物力学分析方法。在血栓溶解过程中，组织特异性转基因过表达的基质金属蛋白酶-2、-14和-9以及编码这些异构体的腺病毒载体将被用来过表达这些蛋白，以确定加速DVT的溶解以及对纤维化和静脉壁生物力学的影响。这些研究将确定基质金属蛋白酶蛋白在血栓溶解的有利和不利方面的作用，并确定预防静脉炎后综合征的潜在分子治疗方法。相关性：这项提案将确定基质金属蛋白酶蛋白如何在血栓形成后导致静脉结疤和增厚，这可能会导致后来的腿部疼痛、肿胀和溃疡。这些研究将增加对血栓如何损害静脉的了解，并测试防止这种损害的新治疗方法。

Description (provided by Applicant): Deep venous thrombosis (DVT) affects more than 2 million Americans per year. Post-phlebitic syndrome can affect 25-75% of patients following DVT and includes leg swelling, pain, skin changes and ulceration of the skin. This develops only in a subset of DVT patients, suggesting that thrombus resolution is critical in determining whether chronic venous obstruction and fibrosis will develop and lead to post-phlebitic syndrome. Matrix metalloproteinase (MMP) genes, critical to cell migration and tissue remodeling, are expressed and activated during thrombus resolution suggesting a critical role in this process. Their role in the pathogenesis of post-phlebitic syndrome will be defined with three Specific Aims: 1) To define the regions of the MMP-2 gene and cognate transcription factors critical to thrombus-induced MMP-2 expression, 2) To determine the role of MMP-2, MMP-9 and MMP-14 (MT-MMP-1) in DVT resolution and thrombus-induced vein wall fibrosis, 3) To determine if overexpression of MMP proteins (MMP-2, MMP-9 and MMP-14) accelerates DVT resolution and alters thrombus-induced vein wall fibrosis. A unique series of transgenic MMP-2 reporter mice will be used to determine which regions of the MMP-2 gene are essential for thrombus-induced MMP-2 transcription and chromatin immunoprecipitation will be used to identity the cognate transcription factors. Mice with targeted deletion of various MMP genes will be used to determine the role of these enzymes in thrombus recanalization, vein wall fibrosis and loss of vein wall compliance and elasticity using novel assays of mouse vein biomechanics. Tissue-specific transgenic overexpression of MMP-2, -14 and -9 as well as adenoviral vectors encoding these isoforms will be utilized to overexpress these proteins during thrombus resolution to determine resolution of DVT is accelerated and the effects on fibrosis and vein wall biomechanics. These studies will define the role of MMP proteins in the beneficial and detrimental aspects of thrombus resolution, and characterize potential molecular therapy to prevent post- phlebitic syndrome. RELEVANCE: This proposal will determine how matrix metalloproteinase proteins cause scarring and thickening of veins after blood clots, which can cause later leg pain, swelling and ulcers. These studies will increase knowledge of how veins are damaged by clots and test new treatments to prevent this damage.