Biomechanical Signaling in Vascular Smooth Muscle Cell Proliferative Disease Following Functional Loss of Elastin

项目总结 细胞外基质(ECM)蛋白弹性蛋白的丢失导致血管平滑肌的失控增殖 肌细胞(VSMC)。这种增殖伴随着血管平滑肌细胞和人类的收缩能力的丧失。 表现为主动脉瓣上狭窄(SVAS)。SVAS患者经历进展性 动脉腔闭塞，最终导致血管僵硬和心力衰竭，为此 机制在很大程度上仍不清楚。初步数据显示，硫酸软骨素和硫酸软骨素 和相关的蛋白多糖在弹性蛋白单倍体不足的小鼠模型的升主动脉中的变化，以及 两者都与弹力合成受损和过度增殖有关。此外，AS通过整合素β3和 FAK在SVAS中上调，而平滑肌收缩标记物的表达减少，I假设 弹性蛋白单倍性不足下的异常ECM成分是由内部识别和传递的 VSMC环境，导致从分化表型到增殖表型的转变。我会对此进行评估 使用诱导多能干细胞(IPSCs)作为模型平台的假说，因为IPSCs呈现无限供应 通过其他传统方法无法获得的患者特定细胞，并用一种 活体小鼠SVAS模型。在目标1中，我将验证IPSC来源的弹性蛋白单倍体不足的表型 神经脊谱系特异的VSMCs到表型SVAS，并试图通过 干扰凡西康在体内和体外的组装。在目标2中，我将研究生物力学信号转导。 参与弹性蛋白单倍体不足的VSMC从分化、收缩状态到表型转换的过程 在IPSC和SVAS小鼠模型中都处于去分化、增殖状态。这些目标包括活体研究。 小鼠模型、免疫荧光、免疫印迹、定量聚合酶链式反应、使用IPSCs的2D和3D建模。这个 这些目标的目的是查明在以下情况下发现的VSMCs无节制扩散的关键因素 弹性蛋白单倍体功能不全，以及挽救这种缺陷的方法。这项提案涉及重要的、未得到充分研究的 血管增殖的机制，并有可能促进治疗这些疾病的发展 目前是不治之症。

PROJECT SUMMARY Loss of the extracellular matrix (ECM) protein elastin leads to the uncontrolled proliferation of vascular smooth muscle cells (VSMCs). This proliferation is accompanied by a loss in contractility of VSMCs and in humans manifests as the condition supravalvular aortic stenosis (SVAS). SVAS patients experience progressive occlusion of their arterial lumen, ultimately leading to vessel stiffening and heart failure, for which the mechanisms remain largely unknown. Preliminary data has shown an upregulation of both chondroitin sulfate and the associated proteoglycan versican in the ascending aortas of elastin haploinsufficient mouse models, and both have been linked to impaired elastogenesis and hyperproliferation. Further, as signaling by integrin β3 and FAK is upregulated in SVAS, while expression of smooth muscle contractile markers is reduced, I hypothesize that abnormal ECM composition under elastin haploinsufficiency is recognized and transduced by the internal VSMC environment, leading to a shift from a differentiated to a proliferative phenotype. I will assess this hypothesis using induced pluripotent stem cells (iPSCs) as a model platform, as iPSCs present a limitless supply of patient-specific cells unattainable through other conventional methods and complement my findings with an in vivo mouse model of SVAS. In Aim 1, I will phenotypically validate elastin haploinsufficient iPSC-derived VSMCs specific to the neural crest lineage to phenocopy SVAS and try to rescue the phenotype through interfering with versican assembly in vivo and in vitro. In Aim 2, I will study the biomechanical signal transduction involved in the phenotypic switching of elastin haploinsufficient VSMCs from a differentiated, contractile state to a dedifferentiated, proliferative state in both iPSC and mouse models of SVAS. These aims involve in vivo studies of murine models, immunofluorescence, western blotting, qPCR, 2D- and 3D-modeling using iPSCs. The objectives of these aims are to identify key factors in the uncontrolled proliferation observed by VSMCs under elastin haploinsufficiency, and methods of rescuing this defect. This proposal addresses important, understudied mechanisms of vascular proliferation, and has the potential to facilitate the development of therapeutics for these currently incurable diseases.