Smooth muscle cell integration of differentiation signals

描述（申请人提供）：血管平滑肌细胞（SMC）在病理性血管重塑过程中改变其分子和表型特征。Notch和TGF¿信号通路都促进成熟SMC的分化、收缩表型特征。然而，这些途径相互作用的机制才刚刚开始被理解。这两种途径的突变都是人类心血管疾病的原因，针对这些途径的治疗目前正在临床试验中。我们的实验室发现了SMC中Notch和Smad之间的调节相互作用，将Notch和TGF¿/BMP/Smad信号联系起来。Notch和TGF信号同时存在，可协同激活phosphoSmad （pSmad）转录活性和SMC标记物表达。本项目的目标是确定导致SMC分化的综合分子信号传导机制。本项目利用人原代血管细胞的分子和生化信号途径，以及小鼠转基因模型研究体内基因调控和功能。我们提出以下假设：1)通过典型的CBF-1介导途径，Jagged1激活SMC中的Notch信号通过转录抑制TGF¿共受体内啡肽。2)协调Notch和TGF -信号通过CBF1-和pSmad-转录复合物的相互作用导致SMC收缩基因的协同激活。3)血管损伤前SMC体内Notch的激活将维持分化表型，从而抑制内膜病变形成和动脉发生。具体目的1：验证TGF¿共受体内啡肽在SMC中受Notch信号的转录调节的假设。本研究将利用体外分子分析和体内转基因小鼠模型来表征Notch对SMC中内啡肽表达和TGF -信号传导的调节。特异性目标2：表征CBF1和Smad转录复合物对SMC收缩基因的相互作用。我们假设CBF1和pSmad转录复合物相互作用，转录调节包括平滑肌肌动蛋白（SM肌动蛋白）和calponin1在内的收缩基因。专项目的3：确定Notch对内啡肽和TGF -¿信号的调控如何影响病理性血管重构。建立的小鼠转基因菌株将在血管疾病模型中进行评估。

DESCRIPTION (provided by applicant): The vascular smooth muscle cell (SMC) alters its molecular and phenotypic characteristics during pathological vascular remodeling. Both Notch and TGF¿ signaling pathways promote a differentiated, contractile phenotype characteristic of mature SMC. However, mechanisms by which these pathways interact with each other are only beginning to be understood. Mutations in either pathway are causal for human cardiovascular disease, and therapies targeting these pathways are currently in clinical trials. Our laboratory discovered a regulatory interaction between Notch and Smad in SMC, which links Notch and TGF¿/BMP/Smad signaling. Concomitant Notch and TGF¿ signaling leads to synergistic activation of phosphoSmad (pSmad) transcriptional activity and SMC marker expression. The goal of this project is to define mechanisms of integrative molecular signaling leading to SMC differentiation. This project utilizes molecular and biochemical signaling approaches in human primary vascular cells, and mouse transgenic models to study in vivo gene regulation and function. We propose the following hypotheses: 1) Jagged1 activation of Notch signaling in SMC transcriptionally represses the TGF¿ co-receptor, endoglin, via the canonical CBF-1 mediated pathway. 2) Coordinate Notch and TGF¿ signaling leads to synergistic activation of SMC contractile genes via interaction of CBF1- and pSmad- containing transcriptional complexes. 3) Activation of Notch in vivo in SMC prior to vascular injury will maintain the differentiated phenotype, leading to suppressed neointimal lesion formation and arteriogenesis. Specific Aim 1: Test the hypothesis that the TGF¿ co-receptor, endoglin, is transcriptionally regulated by Notch signaling in SMC. This aim will utilize molecular assays in vitro and transgenic mouse models in vivo to characterize Notch regulation of endoglin expression and TGF¿ signaling in SMC. Specific Aim 2: Characterize the interaction of CBF1 and Smad transcriptional complexes on SMC contractile genes. We hypothesize that CBF1 and pSmad transcriptional complexes interact to transcriptionally regulate contractile genes including smooth muscle ¿-actin (SM actin) and calponin1. Specific Aim 3: Determine how the regulation of endoglin and TGF¿ signaling by Notch affects pathological vascular remodeling. Established mouse transgenic strains will be evaluated in vascular disease models.