Mechanisms of Microvascular Remodeling Progression

 描述(由申请人提供)：血管重塑是一种适应血管直径长期改变的机制。在高血压中，向内重构，即阻力血管管腔直径的结构性缩小，与心肌梗死和中风的风险增加有关。然而，尽管它的流行和临床重要性，控制内向重塑过程的机制仍然很大程度上是未知的。我们的目标是确定阻力血管向内重塑过程中的机制，可以通过新的策略进行干预，以防止、停止或逆转重塑过程，从而减少与之相关的危及生命的心血管事件。目前的文献和我们自己的初步数据表明，血管平滑肌细胞(VSMC)内的组织型谷氨酰胺转氨酶(TG2)、LIM激酶(LIMK)和基质金属蛋白酶-2(MMP2)参与了血管重塑过程。因此，随着我们和其他人确定内向重塑的阻力血管具有肌动蛋白细胞骨架结构，从而减少其被动直径和细胞外基质(ECM)的特征，特征是内弹力板(IEL)中的窗孔数量和大小减少：我们的假设是，在阻力血管内向重塑过程的早期阶段，长期的血管收缩导致通过TG2和LIM激酶的细胞内活性形成永久性的VSMC细胞骨架结构，这反过来又刺激MMP2的产生和ECM，尤其是IEL的修饰。我们将在VSMC、分离的阻力动脉和高血压的整个动物模型中检验我们的假设。细胞和组织将来自动物，以及正常血压和高血压患者。在我们的假说中测试的重塑成分的表达和活性将通过药理学和分子手段进行调节。实验结果将使用蛋白质和酶活性分析中的传统和前沿技术，以及原子力、多光子和长期活体显微镜来衡量。我们的特定目标将检验以下假设：1)细胞内TG2激活RhoA、Rho激酶和LIMK使cofilin磷酸化和失活，有利于肌动蛋白网络和应力纤维的形成，TG2进一步交联肌动蛋白结构，使其更持久；2)LIMK激活MMP14，导致VSMC表达/分泌MMP2。然后，MMP2通过其弹性溶解作用产生弹性蛋白肽，激活VSMC产生更多弹性蛋白。这种新的弹性蛋白被合并到IEL中，并减少了IEL中窗孔的大小和数量。我们期望这项研究将为高血压时阻力动脉的细胞骨架和IEL结构的改变提供新的见解。这些知识应该对预防和治疗高血压的策略以及与血管重塑相关的疾病的管理产生积极的影响。

 DESCRIPTION (provided by applicant): Vascular remodeling is an adaptive mechanism for long-term modification of vascular diameter. In hypertension, inward remodeling, that is, the structural reduction of the lumen diameter in resistance vessels, is associated with an increased risk for myocardial infarction and stroke. However, despite its prevalence and clinical importance, the mechanisms that control the inward remodeling process remain largely unknown. Our goal here is to identify mechanisms in the inward remodeling process of the resistance vasculature that may be intervened with novel strategies to prevent, stop or reverse the remodeling process, and consequently diminish the life-threatening cardiovascular events associated with it. Current publications and our own preliminary data indicate that tissue-type transglutaminase (TG2), LIM kinase (LIMK), and matrix metalloproteinase-2 (MMP2) within vascular smooth muscle cells (VSMC) are involved in the remodeling process. Therefore, as we and others have determined that inwardly remodeled resistance vessels have actin cytoskeletal structures that reduce their passive diameters and extracellular matrix (ECM) features characterized by a reduction in the number and size of fenestrae in the internal elastic lamina (IEL): Our hypothesis is that during the early stages of the inward remodeling process in resistance vessels, prolonged vasoconstriction leads to formation of permanent VSMC cytoskeletal structures via the intracellular activity of TG2 and LIM kinase, which in turn stimulate the production of MMP2 and the modification of the ECM, in particular the IEL. We will test our hypothesis in VSMC, isolated resistance arteries and a whole animal model of hypertension. Cells and tissues will come from animals, as well as from normotensive and hypertensive individuals. The expression and activity of the remodeling components tested in our hypotheses will be modulated using pharmacological and molecular means. Experimental outcomes will be measured using traditional and leading-edge techniques in protein and enzymatic activity analyses, as well as, atomic force, multiphoton, and long-term intravital microscopy. Our specific aims will test the hypotheses that: 1) Intracellular TG2 activates RhoA, Rho kinase and LIMK to phosphorylate and inactivate cofilin to favor formation of actin networks and stress-fibers, with TG2 further crosslinking actin structures to make them more persistent; and 2) that LIMK activates MMP14 and leads to expression/secretion of MMP2 from VSMC. Then MMP2 through its elastolytic actions generates elastin peptides that activate VSMC to produce more elastin. This new elastin is incorporated in the IEL and reduces the size and number of fenestrae in the IEL. We expect this study will provide new insights on how cytoskeletal and IEL structures of resistance arteries are modified in hypertension. This knowledge should have a positive impact on strategies for preventing and treating hypertension, and the management of diseases associated with vascular remodeling.