Role of PTEN in Hypoxia-Induced Vascular Remodeling, Raphael Nemenoff

缺氧诱导的肺动脉高压（PAH）的发病机制的特点是： 血管收缩和血管重塑导致肺血管阻力增加， 右心衰竭这些反应的特点是在驻地血管壁细胞的功能变化 包括平滑肌（SMC）、内皮和成纤维细胞，以及循环祖细胞的募集 和炎症细胞。我们的初步数据表明，缺氧导致平滑肌细胞Akt的快速激活， 肺部脉管系统PTEN是PI 3-激酶/Akt/mTOR信号传导的负调节剂，并且是PI 3-激酶/Akt/mTOR信号传导的抑制剂。 SMC增殖。我们已经证明，SMC特异性靶向PTEN突变小鼠（PTEN KO） 自发地发展为肺动脉高压。PTEN KO小鼠还表现出血管周围和血管外的血管密度增加。 血清趋化因子水平、循环祖细胞增加以及祖细胞向主要器官的运输 血管和肺。其他研究者在该PPG中的工作表明，罗格列酮，一种特异性 核受体PPARy的激活剂，减弱缺氧诱导的肺血管重塑。在 在其他系统中，PPARy抑制细胞增殖和调节抗炎反应的能力是 在很大程度上通过上调PTEN表达和/或活性介导。基于这些 综合观察，我们假设肺动脉SMC中PTEN的失活将诱导 慢性缺氧导致重度PAH。这种反应将通过对SMC的直接作用介导 增殖以及SMC产生趋化因子，这将参与招募 的祖细胞/促炎细胞，也可以自分泌的方式对SMC本身。 相反，激活PPARy将至少抑制低氧诱导的肺血管重构。 部分通过上调PTEN表达。该项目将采用体内和体外方法， 测试这个模型。提出了两个具体目标。Aiml将使用一种新的，tampxifen诱导的PTEN 基因敲除小鼠模型，以检查小鼠中SMC特异性缺失PTEN对趋化因子的影响。 诱导的SMC增生和祖细胞和炎性细胞的募集 肺血管重构该模型将允许中膜SMC和骨髓的命运映射- 在PAH发病过程中衍生的祖细胞，提供了关于 这些细胞在肺血管重塑中的作用。体外研究将使用shRNA沉默， 肺动脉SMC中的PTEN以确定介导这些反应的下游效应物。目标2将 采用类似的策略在体内特异性删除PPARy并与PTEN比较反应 缺陷小鼠体外实验将确定PTEN在介导PPARy作用中的作用。 最后，研究了PTEN在介导罗格列酮和吡格列酮（两种良好的抗肿瘤药物）的保护作用中的作用。 将检查表征的PPARy活化剂。 相关性（参见说明）： 介导缺氧诱导的PAH的分子途径涉及多种细胞类型。这个项目是 旨在专门研究SMC对这一过程的贡献。新的小鼠模型，其中 信号通路可以以时间依赖的方式被操纵，特别是在SMC中， 的SMC。体外方法将描绘控制生长、表型调节 以及这些细胞的细胞因子产生。调节这些途径的药物代表了新的 用于治疗和预防PAH的治疗剂。

The pathogenesis of hypoxia-induced pulmonary arterial hypertension (PAH) is characterized by vasoconstriction and vascular remodeling contributingto increased pulmonary vascular resistance leading to right heart failure. These responses are characterized by functional changes in resident vascular wall cells including smooth muscle (SMC), endothelial,and fibroblast, as well as recruitment of circulating progenitor and inflammatory cells. Our preliminary data indicatethat hypoxia leads to rapid activation of Akt in SMC of the lung vasculature. PTEN is a negative regulator of PI3-kinase/Akt/mTOR signaling, and an inhibitor of SMC proliferation. We have shown that SMC-specific, targeted PTEN mutant mice (PTEN KO) spontaneously develop pulmonary hypertension. PTEN KO mice also exhibit increased perivascular and serum chemokine levels, increases in circulating progenitor cells, and trafficking of progenitor cells to major vessels and the lung. Work by other investigators in this PPG demonstrated that rosiglitazone, a specific activator of the nuclear receptor PPARy, attenuates hypoxia-induced pulmonary vascular remodeling. In other systems, the ability of PPARy to inhibit cell proliferation and regulate anti-inflammatory responses is mediated, in large part, through the upregulation of PTEN expression and/or activity. Based on these collective observations, we hypothesize that inactivation of PTEN in pulmonary arterial SMC will induce severe PAH in response to chronic hypoxia. This response will be mediated by direct effects on SMC hyperplasia as well as the production of chemokines by SMC which will be involved in the recruitment of progenitor/pro-inflammatory cells and may also act in an autocrine fashion on the SMC themselves. Conversely, activation of PPARy will inhibit hypoxia-induced pulmonary vascular remodeling at least in part through the upregulation of PTEN. This project will employ both in vivo and in vitro approaches to test this model. Two specific aims are proposed. Aiml will use a novel, tampxifen-inducible PTEN knockout mouse model to examine the effects of SMC-specific deletion of PTEN in mice on chemokine- induced SMC hyperplasia and recruitment of progenitor and inflammatory cells during hypoxia-induced pulmonary vascular remodeling. This model will allow fate-mapping of medial SMC and bone marrow- derived progenitor cells during the pathogenesis of PAH, providing clear information regarding the contributions of these cells in pulmonary vascular remodeling. In vitro studies will use shRNA silencing of PTEN in pulmonary artery SMC to define downstream effectors mediating these responses. Aim 2 will employ an analogous strategy to specifically delete PPARy in vivo and compare responses with PTEN deficient mice. In vitro experiments will establish the role of PTEN in mediating the effects of PPARy. Finally, the role of PTEN in mediating the protective effects of rosiglitazone and pioglitazone, two well- characterized PPARy activators, will be examined. RELEVANCE (See instructions): The molecular pathways mediating hypoxia-induced PAH involve multiple cell types. This project is designed to specifically examine the contribution of SMC to this process. Novel mouse models in which signaling pathways can be manipulated in a time-dependent fashion specifically in SMC will define the role of SMC. In vitro approaches will delineate molecular pathways controlling growth, phenotypic modulation and cytokine production by these cells. Pharmacological agents regulating these pathways represent novel therapeutic agents for treatment and prevention of PAH.