Epigenetic control of smooth muscle cell phenotype during microvascular remodeling

项目总结 外周动脉疾病(PAD)是一种导致肢体动脉衰弱的闭塞性疾病。 由于适当的血管形成和有效血管的缺陷而引起的并发症(如跛行、截肢) 改建。最近通过血管内皮生长因子治疗促进PAD患者治疗性血管生成的尝试已经 失败，可能是因为这些策略只针对平滑肌细胞(SMC)，即 对毛细血管和末梢小动脉重塑(即动脉生成)以增加血液分布是必要的 到缺血区。动脉形成依赖于SMC的可塑性和可逆表型 在他们瞬时下调收缩装置的地方切换，参与毛细血管投资，以及 然后重新分化回收缩状态。然而，对控制机制的理解 SMC在体内重新分化和保留谱系记忆的能力是有限的。我们之前确定了 SMC谱系特有的表观遗传学特征，包括组蛋白3的赖氨酸4的二甲基化 (H3K4me2)在SMC标记基因的启动子(简称SMC-H3K4me2)上 SMC去分化。这些观察结果表明，SMC-H3K4me2可能在维持中发挥关键作用 SMC的身份和血统记忆的保留，这可能是控制SMC参与的关键机制 在动脉形成中。这项提议中的研究将检验H3K4me2控制SMC的中心假设 分化状态，并使SMC参与侧支毛细血管投资和肌化 通过促进TET2在SMC标记基因上的募集而发生动脉形成。与这一假设相一致，我们的 利用一种新的位点特异性表观基因组编辑系统特异性去除H3K4me2的初步研究 在SMC标记基因上，提供了SMC-H3K4me2密切调控SMC分化状态和SMC分化的证据 发现H3K4me2与介导DNA去甲基化的关键酶Ten-Eleven易位(TET2)相互作用 是SMC分化的主要调节者。我们的中心假设将通过解决以下问题来进一步检验 遵循特定的目标。目标1将确定SMC-H3K4me2丢失对SMC参与的影响 动脉形成。SMC-H3K4me2去除对SMC表型的功能影响，他们的能力 投资毛细血管，以及组织再灌流将被评估。目标2将检验H3K4me2的假设 作为TET2在SMC标志物上的对接位点，在促进SMC分化方面发挥了关键作用 基因。我们期望这些研究的完成将导致新的控制机制的确定 平滑肌细胞的特性、可塑性和谱系记忆及其对有益微血管的参与 动脉化和成熟化。这些结果可以通过制定新的战略来促进 PAD患者的治疗性血运重建。

PROJECT SUMMARY Peripheral Artery Disease (PAD) is an occlusive disease of the lower extremity arteries leading to debilitating complications (e.g. claudication, amputation) due to defect in proper vascularization and efficient vascular remodeling. Recent attempts to promote therapeutic angiogenesis by VEGF therapies in patients with PAD have failed, perhaps because these strategies have only not targeted smooth muscle cells (SMC), cells that are essential for remodeling of capillaries and terminal arterioles (i.e. arteriogenesis) to increase blood distribution to ischemic regions. Arteriogenesis relies on the ability of SMC to be plastic and undergo a reversible phenotypic switching where they transiently downregulate their contractile apparatus, participate in capillary investment, and then re-differentiate back to the contractile state. However, the understanding of the mechanisms controlling SMC ability to re-differentiate and the retention of their lineage memory in vivo is limited. We previously identified an epigenetic signature specific to the SMC lineage consisting of the dimethylation of the lysine 4 of histone 3 (H3K4me2) on the promoters of the SMC marker genes (referred as SMC-H3K4me2) that is retained during SMC dedifferentiation. These observations suggest that SMC-H3K4me2 could play a pivotal role in maintenance of SMC identity and retention of lineage memory and could be a key mechanism controlling SMC participation in arteriogenesis. Studies in this proposal will test the central hypothesis that H3K4me2 controls the SMC differentiation state and enables SMC involvement in collateral capillary investment and muscularization during arteriogenesis by promoting the recruitment of TET2 on SMC marker genes. Consistent with this hypothesis, our preliminary studies, utilizing a novel locus-specific epigenome editing system to remove specifically H3K4me2 on the SMC marker genes, provide evidence that SMC-H3K4me2 tightly regulates SMC differentiation state and show that H3K4me2 interacts with Ten-eleven translocation (TET2), a key enzyme mediating DNA demethylation and a master regulator of SMC differentiation. Our central hypothesis will be further tested by addressing the following specific aims. Aim 1 will determine the impact of loss of SMC-H3K4me2 on SMC participation to arteriogenesis. The functional consequences of SMC-H3K4me2 removal on SMC phenotype, their ability to invest capillaries, as well as tissue reperfusion will be evaluated. Aim 2 will test the hypothesis that H3K4me2 plays a key role in promoting SMC differentiation by serving as a docking site for TET2 on the SMC marker genes. We expect the completion of these studies will lead to the identification of novel mechanisms controlling smooth muscle cell identity, plasticity, and lineage memory and their participation to beneficial microvascular arterialization and maturation. These results could serve from the development of new strategies for enhancing therapeutic vascularization in patients with PAD.