Mechano-Sensitive Hypoxia-Inducible Factor-MMP Pathway in Venous Insufficiency

描述(申请人提供)：慢性静脉功能不全(CVI)是一种常见且代价高昂的疾病，其特征是静脉过度扩张和静脉曲张。CVI多发生在下肢，提示有静脉压和静脉壁拉伸的作用。此外，CVI患者血浆和静脉组织中基质金属蛋白酶(MMPs)水平升高，提示MMPs在静脉扩张中起作用。我们发现，延长大鼠静脉的伸展与收缩减少和基质金属蛋白酶-2和-9的表达增加有关。此外，与对照大隐静脉相比，静脉曲张的静脉曲张节段表现为收缩减少。这些新的发现使得研究静脉曲张相关静脉扩张中静脉壁拉伸和MMPs之间的联系，以及确定相关的上游和下游机制变得重要。我们的数据表明，长时间的静脉拉伸与缺氧诱导因子(HIF)的表达增加有关。此外，即使在没有检测到细胞外基质(ECM)降解的情况下，MMP-2和-9也能诱导静脉节段的松弛，这表明抑制静脉平滑肌(VSM)收缩的机制。这一建议的目的是验证中心假说，即静脉压升高和静脉壁延长与机械敏感的HIF-MMP途径上调有关，这反过来会导致VSM收缩机制的下游抑制和ECM蛋白的降解增加，从而导致静脉过度扩张。因此，HIF-MMPs途径的下调应该会改善长时间拉伸的静脉和静脉曲张中的反应性。机制研究将在股静脉压力升高的大鼠模型上进行，并在分离的髂静脉和股静脉上进行。虽然老鼠是四条腿的动物，但老鼠是一个始终如一的品种，避免了人类在年龄、性别和其他混杂因素上的差异。为了加强翻译方面，将在人静脉曲张上进行实验，将其与邻近的近端和远端静脉进行比较，并控制隐静脉。其具体目的是确定：1)静脉压/静脉壁拉伸增加是否与静脉收缩减少和HIF/基质金属蛋白酶途径上调有关。2)HIF/MMP活性升高通过抑制VSM收缩的下游机制(包括[Ca~(2+)]i、蛋白激酶C和Rho-K活性)促进静脉扩张，并增加ECM降解。3)静脉曲张中静脉扩张的增加是由于HIF/MMPs上调所致，因此，使用HIF或MMPI或MMPs抑制剂和siRNA下调HIF或MMPs应可改善静脉曲张节段的收缩，使其接近邻近近端或远端静脉或对照大隐静脉的水平。这些研究将阐明静脉压升高/静脉壁拉伸、HIF/MMP表达/活性、VSM收缩减少的机制和静脉过度扩张之间的关系。这一结果将突出HIF和MMPs的特定抑制剂作为预防静脉曲张进展和复发的新策略的好处。

DESCRIPTION (provided by applicant): Chronic venous insufficiency (CVI) is a common and costly disease characterized by excessive vein dilation and varicose veins. CVI commonly occurs in the lower extremity, suggesting a role of venous pressure and vein wall stretch. Also, the plasma and venous tissue levels of matrix metalloproteinases (MMPs) are elevated in CVI, suggesting a role for MMPs in vein dilation. We have found that prolonged stretch of rat veins is associated with decreased contraction and increased expression of MMP-2 and -9. Also, varix segments of varicose veins demonstrate reduced contraction as compared to control saphenous vein. These novel findings make it important to investigate the link between vein wall stretch and MMPs in the venous dilation associated with varicose veins, and to identify the upstream and downstream mechanisms involved. Our data suggest that prolonged vein stretch is associated with increased expression of hypoxia-inducible factors (HIF). Also, MMP- 2 and -9 induce relaxation of vein segments even in the absence of detectable extracellular matrix (ECM) degradation, suggesting inhibition of venous smooth muscle (VSM) contraction mechanisms. The objective of this proposal is to test the central hypothesis that increased venous pressure and prolonged vein wall stretch are associated with upregulation of a mechano-sensitive HIF-MMP pathway, which in turn causes downstream inhibition of VSM contraction mechanisms and increased degradation of ECM proteins, leading to excessive venous dilation. Consequently, downregulation of the HIF-MMP pathway should improve reactivity in veins subjected to prolonged stretch and in varicose veins. Mechanistic studies will be conducted in a rat model of increased femoral venous pressure, and on isolated iliac and femoral veins. Although the rat is a four-legged animal, the rat is a consistent breed that avoids the variability in age, sex and other confounding factors in humans. To enhance the translational aspects, experiments will be conducted on human varix veins as compared to adjacent proximal and distal veins, and control saphenous veins. The specific aims are to determine whether: 1) Increased venous pressure/vein wall stretch is associated with decreased vein contraction and upregulation of HIF/MMP pathway. 2) Increases in HIF/MMP activity promote venous dilation by downstream inhibition of the mechanisms of VSM contraction including [Ca2+]i, protein kinase C and Rho- kinase activity, and increased ECM degradation. 3) The increased venous dilation in varicose veins occurs as a result of upregulation of HIF/MMPs, and therefore downregulation of HIF or MMPs using HIF or MMP inhibitors and siRNA should improve contraction in varix segments to levels approaching those observed in the adjacent proximal or distal veins, or in control saphenous vein. These studies would elucidate the relation between increased venous pressure/vein wall stretch, HIF/MMP expression/activity, reduced mechanisms of VSM contraction, and excessive venous dilation. The results would highlight the benefits of specific inhibitors of HIF and MMPs as a new strategy to prevent the progression and recurrence of varicose veins.