Epigenetic control of vascular smooth muscle in cardiovascular disease

描述(申请人提供)：血管平滑肌细胞(VSMC)表现出显著的可塑性，允许可逆分化和去分化。这一特性不同于骨骼或心肌细胞，后者经历终末分化。平滑肌的可塑性在正常发育、生长和伤口愈合过程中非常重要，但也会导致动脉粥样硬化、内膜增生、高血压和移植动脉硬化等病理变化。由于VSMC表现出这种独特的可塑性，我们假设在其他类型的干细胞中发现的调控机制也可能参与VSMC的表型调节。事实上，我们的初步数据表明，去分化的VSMC表达多种干细胞基因，包括Oct4、Nanog和KLF4。由于染色质修饰蛋白Tet(Ten-Elevation)家族最近被认为是胚胎和造血干细胞多能性的重要调节因子，我们研究了Tet蛋白在VSMC中的作用。我们的初步数据表明，TET2在VSMC中高水平表达，并被分化剂诱导，但被PDGF-BB抑制。我们发现TET2是SMC分化的必要条件和充分条件，其中TET2与收缩基因启动子结合并增加可及性，但抑制干细胞基因启动子的染色质。我们进一步发现，TET2在正常血管中高水平表达，但在疾病状态下显著下调，包括人类动脉粥样硬化和小鼠内膜增生。重要的是，TET2的过度表达挽救了动脉损伤后的内膜增生。我们推测TET2是一种新的VSMC表型表观遗传主控调控因子。我们相信我们已经发现了一种基本的新机制，这是平滑肌可塑性的基础。因此，这项工作对多种血管疾病具有重要意义。在以下特定目标中，我们建议解决TET2的作用机制，并确定TET2在血管疾病模型中的作用。在具体目标1中，我们将使用尖端方法来确定TET2调节收缩和干细胞启动子的甲基化和基因表达的表观遗传学机制。在特定的目标2中，我们将使用小鼠模型来确定TET2在心血管疾病的发病机制中的作用，包括动脉粥样硬化、内膜增生和移植物动脉硬化。由于我们发现TET2的表观遗传调控位于VSMC表型调控的顶端，控制着其他已知的介质，如Myocardin、SRF和KLF4，我们认为在细胞和疾病水平上了解TET2的作用机制将为心血管疾病的治疗和预防创造新的治疗途径。

DESCRIPTION (provided by applicant): Vascular smooth muscle cells (VSMC) exhibit a remarkable plasticity that allows for reversible differentiation and de-differentiation. This propety is distinct from skeletal or cardiac myocytes, which undergo terminal differentiation. Smooth muscle plasticity is important in normal development, growth, and wound healing, but also contributes to pathologies including atherosclerosis, intimal hyperplasia, hypertension, and transplant arteriosclerosis. Because VSMC exhibit this unique plasticity, we hypothesized that regulatory mechanisms found in other types of stem cells might also be involved in VSMC phenotypic modulation. Indeed, our preliminary data demonstrate that de-differentiated VSMC express multiple stem cell genes, including Oct4, Nanog, and KLF4. As the TET (ten-eleven translocation) family of chromatin-modifying proteins has recently been implicated as essential regulators of pluripotency in embryonic and hematopoietic stem cells, we investigated the roles of TET proteins in VSMC. Our preliminary data reveal that TET2 is expressed at high levels in VSMC, and is induced by differentiating agents but repressed by PDGF-BB. We find that TET2 is necessary and sufficient for SMC differentiation, where TET2 associates with contractile gene promoters and increases accessibility, but represses chromatin at stem cell gene promoters. We further found that TET2 is expressed at high levels in normal vessels, but markedly downregulated in disease states, including human atherosclerosis and intimal hyperplasia in mice. Importantly, TET2 overexpression rescues intimal hyperplasia post-arterial injury. We hypothesize that TET2 is a novel epigenetic master regulator of VSMC phenotype. We believe we have uncovered a fundamental new mechanism that is the basis for smooth muscle plasticity. As such, this work has important implications for multiple vascular diseases. In the following specific aims, we propose to address mechanisms of action of TET2, and determine the role of TET2 in vascular disease models. In Specific Aim 1, we will use cutting edge methods to determine the epigenetic mechanisms by which TET2 regulates methylation and gene expression at contractile and stem cell promoters. In Specific Aim 2, we will use mouse models to determine the role of TET2 in the pathogenesis of cardiovascular diseases, including atherosclerosis, intimal hyperplasia, and graft arteriosclerosis. Because we have found that epigenetic regulation by TET2 lies at the apex of VSMC phenotypic modulation, controlling other known mediators such as myocardin, SRF, and KLF4, we propose that understanding the mechanisms of TET2 action at the cellular and disease level will generate new therapeutic approaches for treatment and prevention of cardiovascular disease.