Microenvironmental Endogenous Reprogramming of Differentiated Smooth Muscle Cells

描述（由申请人提供）：血管平滑肌细胞（SMC）是高度特化的细胞，收缩以维持血管张力，处于静止状态并表达高水平的SMC特异性蛋白，如平滑肌肌球蛋白重链（SMMHC）。然而，在病理条件下，SMC能够经历深刻的表型和功能变化，使其成为病理性新生内膜形成的主要贡献者。近年来的一个重要范式转变表明，具有分化为SMC和内皮细胞能力的血管祖细胞（VPC）位于出生后血管外膜内的专门生态位中。虽然新出现的数据支持外膜VPC的存在，但仍存在重要的未回答的问题，包括其起源及其对血管维护、修复和病理性病变形成的贡献。使用一种创新的体内命运映射方法，我们进行了意料之外的观察，成熟的SMC迁移到外膜，下调SM标志物的表达，并获得祖细胞标志物的表达。我们的初步研究结果表明，SMC衍生的Sca 1（+）细胞群体可以从表达功能性祖细胞表型并表现出分化为SMC的能力的成人血管中分离出来。一小部分保留了Sca 1的表达，表明有自我更新的潜力。我们的研究结果表明，居民VPC的出现，从重新编程的成熟SMC。我们的总体假设是，成熟的平滑肌细胞迁移到血管发育的后期阶段的外膜，并重新编程为VPC在响应特定的信号表达专门在外膜（例如微环境重编程）;重编程是依赖于诱导的转录因子，Klf 4。血管损伤后，这些细胞的募集有助于新生内膜的形成。这一发现可能对定义VPC形成和维持的分子机制产生深远的影响，并可能导致我们对体细胞重编程的理解取得进展，以改善临床治疗应用。成熟的SMC作为常驻VPC的来源是一个反教条的理论，将挑战现有的概念。提出了两个目的，以确定SMC衍生的AdvSca 1细胞在正常血管和血管损伤中的重要性和作用。在目标一中，我们将表征和定义成熟SMC对外膜VPC池的贡献，我们将定义SMC衍生的VPC的体外功能特征，我们将定义Klf 4对SMC重编程的体外和体内作用。对于目标二，我们将确定SMC衍生的VPC对导丝诱导的血管损伤的反应。

DESCRIPTION (provided by applicant): Vascular smooth muscle cells (SMCs) are highly specialized cells that contract to maintain vascular tone, are quiescent and express high levels of SMC-specific proteins, such as smooth muscle myosin heavy chain (SMMHC). Under pathological conditions, however SMCs are capable of undergoing profound phenotypic and functional changes making them major contributors to pathological neointima formation. An important paradigm shift in recent years suggests that vascular progenitor cells (VPCs) with the capacity to differentiate into SMCs and endothelial cells reside in a specialized niche within the adventitia of postnatal vessels. While emerging data support the existence of adventitial VPCs, important unanswered questions remain, including their origin and their contribution to vessel maintenance, repair, and pathological lesion formation. Using an innovative in vivo fate-mapping approach, we made the unanticipated observation that mature SMCs migrate into the adventitia, downregulate SM marker expression, and gain expression of progenitor cell markers. Our preliminary findings demonstrate that a population of SMC-derived Sca1(+) cells can be isolated from adult vessels that express a functional progenitor cell phenotype and exhibit the capacity to differentiate into SMCs. A small percentage retained expression of Sca1, indicating the potential for self-renewal. Our findings suggest the emergence of resident VPCs from reprogrammed mature SMCs. Our overall hypothesis is that mature SMCs migrate to the adventitia during the later stages of vascular development and are reprogrammed to VPCs in response to specific signals expressed exclusively in the adventitia (e.g. microenvironmental reprogramming); reprogramming is dependent on induction on the transcription factor, Klf4. After vascular injury, recruitment of these cells contributes to neointima formation. Such a finding could have a profound impact on defining the molecular mechanisms underlying the formation and maintenance of VPCs and could lead to advances in our understanding of somatic cell reprogramming to improve clinical therapeutic applications. Mature SMCs as the source of resident VPCs is an anti-dogmatic theory that would challenge existing concepts. Two Aims are proposed to establish the importance and the role of SMC-derived AdvSca1 cells in normal vessels and in the setting of vascular injury. In Aim One, we will characterize and define the contribution of mature SMC to the adventitial VPC pool, we will define the in vitro functional characteristics of SMC-derived VPCs, and we will define the in vitro and in vivo role of Klf4 on SMC reprogramming. For Aim Two, we will determine the fate of SMC-derived VPCs in response to wire-induced vascular injury.