TGF-beta in intimal hyperplasia after vascular bypass

描述(申请人提供)：再狭窄，或血管介入治疗动脉粥样硬化后动脉管腔再次狭窄，对成千上万的人产生显著的死亡率和发病率。转化生长因子-β(TGF-2)是再狭窄的重要介质，但其对动脉壁的影响机制尚不清楚。在我们完成的方案中，我们探讨了这些机制，并假设抑制转化生长因子-2的S促纤维化作用和增强其对平滑肌细胞迁移和增殖的抑制作用将为控制内膜增生提供场所。我们的研究为了解转化生长因子-2的作用机制提供了新的见解。与我们最初的假设相反，转化生长因子-2不是通过产生细胞外基质而是通过信号分子SMAD3刺激SMC增殖而产生内膜增生。我们的数据还表明，转化生长因子-2可能通过SMAD3信号途径，通过募集骨髓前体细胞(BMPC)进入内膜病变，从而促进新生血管的增殖。最后，我们发现，转化生长因子-2通过Smad3，通过蛋白结缔组织生长因子(CTGF)介导，产生适应性重塑或动脉扩张。这些新发现产生了三个假说。在特定的目标I中，我们将检验这样的假设，即Smad3水平在动脉损伤中升高，持续高水平的Smad3通过将静止的Smad3转化为对转化生长因子-2具有增殖反应的表型而导致内膜增生。我们将首先测试阻断内源性Smad3的产生对体内新生内膜形成的影响。为了更好地了解Smad3激活的SMC从静止表型到增殖表型的转换，我们将研究Smad3与细胞周期蛋白依赖的激酶抑制物p27的关系。最后，我们将探讨转化生长因子-2和Smad3在人类再狭窄和动脉粥样硬化斑块中是否以相似的方式发挥作用。在特定的目标II中，我们将测试转化生长因子-2是否通过Smad3刺激动脉SMC产生BMPC的趋化物质而促进内膜增生。利用大鼠骨髓移植模型进行的体内研究，旨在验证转化生长因子-2和Smad3在祖细胞募集中的作用。此外，我们还将验证单核细胞趋化蛋白-1是介导转化生长因子-2和Smad3的S效应的趋化因子的假设。在特定的目的III中，我们将测试SMC中的转化生长因子-2/Smad3通路是否刺激CTGF的产生，而CTGF通过不同地调节I型和III型胶原的合成来刺激适应性重塑。我们将证明CTGF是适应性重塑的必要条件和充分条件，并探讨III型胶原在这一过程中的作用。通过这些研究，我们希望进一步了解转化生长因子-2及其在细胞增殖、骨髓基质细胞募集和动脉重塑中的作用。最终，我们的发现将导致抑制内膜增生和促进适应性重塑的治疗方法的开发，为毁灭性的再狭窄问题提供解决方案。在美国，动脉粥样硬化是主要的死亡原因。动脉粥样硬化的治疗方法很多；然而，动脉腔再狭窄或再狭窄的过程阻碍了它们的长期成功，血管成形术或支架置入术后30%-50%的患者会发生这种情况。我们建议的目标是建立在我们在过去几年中获得的关于转化生长因子-2的分子机制及其在再狭窄过程中所起作用的知识的基础上。长期目标是开发特定的治疗方法，防止或阻止动脉再狭窄的进展，从而减少每年数千人的发病率和死亡率。

DESCRIPTION (provided by applicant): Restenosis, or re-narrowing of the arterial lumen following vascular interventions to treat atherosclerosis, produces significant mortality and morbidity for thousands of individuals. Transforming growth factor-beta (TGF-2) is an important mediator of restenosis, although the mechanisms that contribute to its effects on the arterial wall remain unclear. In our completed proposal, we explored these mechanisms and hypothesized that inhibition of TGF-2's profibrotic effects and enhancement of its inhibitory effects on smooth muscle cell (SMC) migration and proliferation would provide a venue for controlling intimal hyperplasia. Our studies have provided new insights into the mechanism of action of TGF-2. Contrary to our original hypothesis, TGF-2 produces intimal hyperplasia not through the production of extracellular matrix but rather by stimulating SMC proliferation through the signaling molecule Smad3. Our data also suggest that TGF-2, through Smad3 signaling, may contribute to initmal hyperplasia through the recruitment of bone marrow progenitor cells (BMPC) into the intimal lesion. Lastly we found that TGF-2, through Smad3, appears to produce adaptive remodeling or arterial enlargement, mediated through the protein connective tissue growth factor (CTGF). These novel findings have generated three hypotheses. In Specific Aim I, we will test the hypothesis that Smad3 levels are elevated in arterial injury and that sustained high levels of Smad3 lead to intimal hyperplasia by converting quiescent SMCs to a phenotype that responds to TGF-2 with proliferation. We will begin by testing the effect of blocking endogenous Smad3 production on neointimal formation in vivo. To better understand the switch of Smad3 activated SMCs from a quiescent to a proliferative phenotype, we will study the relationship between Smad3 and the cyclin-dependent-kinase inhibitor p27. Finally, we will explore whether TGF-2 and Smad3 function in a similar fashion in human restenotic and atherosclerotic plaque. In Specific Aim II, we will test whether TGF-2, through Smad3, enhances intimal hyperplasia by stimulating arterial SMCs to produce a chemoattractant of BMPCs. In vivo studies, using a rat bone marrow transplant model, have been designed to verify the role in TGF-2 and Smad3 in progenitor cell recruitment. Moreover, we will test the hypothesis that monocyte chemoattractant protein-1 (MCP-1) is the chemoattractant that mediates TGF-2 and Smad3's effect. In Specific Aim III, we will test whether the TGF-2/Smad3 pathway in SMCs stimulates the production of CTGF, which in turn stimulates adaptive remodeling by differentially regulating synthesis of collagen types I and III. We will show that CTGF is necessary and sufficient for adaptive remodeling and explore the role of collagen type III in this process. With these studies, we expect to gain further insight into TGF-2 and its role in cell proliferation, BMPC recruitment, and arterial remodeling. Ultimately, our findings will lead to the development of therapies that can inhibit intimal hyperplasia and promote adaptive remodeling, providing a solution to the devastating problem of restenosis. Atherosclerosis is the leading cause of death in the United States. Treatments for atherosclerosis are numerous; however, their long term success is impeded by the process of restenosis or re-narrowing of the arterial lumen, which occurs in 30-50% of patients following angioplasty or stenting. The goal of our proposal is to build upon knowledge that we have acquired over the past several years about the molecular mechanisms that underlie TGF-2 and its role in the process of restenosis. The long-term goal is to develop specific therapies that prevent or halt the progression of arterial restenosis, thus reducing morbidity and mortality for thousands of individuals each year.