Mitochondrial network remodeling and the development of the hyper-proliferative and antiapoptotic endothelial phenotype.

项目摘要 肺血管疾病是导致婴儿和儿童常见的 先天性心脏缺陷导致肺血流量（PBF）和压力增加。都缺乏 有效的治疗，以限制共同的病理生理特征的内皮功能障碍和血管 重塑我们最近的研究表明，代谢重编程和线粒体功能障碍， 由机械应力介导，是这些儿童血管损伤的核心调节途径。 此外，我们最近发现存在过度增殖，抗凋亡的内皮细胞表型， 在我们的分流羔羊模型中，增加的PBF和压力参与血管生成反应并导致 肺小动脉数量增加。我们的数据表明，这种内皮细胞表型与 生存素（一种抗凋亡蛋白）表达增加，线粒体分裂增加， 自噬/线粒体自噬。这些过程与NO信号的丢失有关。在我们的研究中， 分流羔羊模型至少部分地通过ATP介导的hsp 90活化的减少而发生。大规模 与过度增殖相关的代谢需求需要大量消耗ATP。基于 这些数据我们的总体假设是，维持细胞过度增殖、抗肿瘤所需的ATP消耗量 与PBF和压力增加相关的凋亡、内皮表型在损失中起重要作用。 通过减弱热休克蛋白90的活性来抑制NO信号传导。我们的数据暗示RhoA/ROCK信号传导作为一个主调节器， 这些途径。我们实验室的研究表明，S1 PR 3受体的连接诱导Rho GT3信号转导， 和细胞骨架重塑。有趣的是，S1 PR 1受体，它对机械刺激产生保护作用， 应激，在我们的分流羔羊模型的肺中显著下调，而S1 PR 3受体表达在肺中显著下调。 增加我们假设机械应力介导的S1 PR 3受体-RhoA/ROCK的激活 轴负责线粒体分裂、自噬/线粒体自噬和细胞凋亡，协同产生 过度增殖、抗凋亡内皮表型和NO信号传导的丧失。三个具体目标 (SAs)来检验这一假设。在目标1中，我们将定义线粒体裂变在细胞分裂中的作用。 过度增殖的内皮细胞表型的发展，并确定这是如何调节NO信号。在 目的2，我们将描述自噬/线粒体自噬在条件下NO信号转导丢失中所起的作用。 增加PBF和压力，并研究抗凋亡因子生存素（Birc 5）所起的作用。在 目的3，我们将确定靶向线粒体分裂和自噬是否是潜在的治疗靶点 在我们的临床前分流羔羊模型。随着这项研究的完成，我们将大大增加我们的 理解线粒体网络重塑和自噬/线粒体自噬在细胞凋亡中的作用， 肺血管疾病的发病机制与PBF和压力增加有关，同时强调 新的治疗干预的应用。

Project Summary Pulmonary vascular disease is responsible for significant morbidity in infants and children with common congenital heart defects that result in increased pulmonary blood flow (PBF) and pressure. There is a lack of effective therapies to limit the shared pathophysiologic features of endothelial dysfunction and vascular remodeling. Our recent studies have demonstrated that metabolic reprogramming and mitochondrial dysfunction, mediated by mechanical stress, is a core regulatory pathway underlying the vascular injury in these children. Further, we have recently identified the presence of a hyper-proliferative, anti-apoptotic endothelial phenotype in our Shunt lamb model of increased PBF and pressure that is involved in an angiogenic response and results in an increase in pulmonary arteriole number. Our data indicate that this endothelial phenotype is associated with increased expression of survivin (an anti-apoptotic protein), mitochondrial fission and increased autophagy/mitophagy. These processes are linked to a loss of NO signaling. The decreased NO signaling in our Shunt lamb model occurs, at least in part, through a decrease in ATP-mediated hsp90 activation. The massive metabolic requirement associated with hyper-proliferation requires a significant consumption of ATP. Based on these data our overall hypothesis is that the ATP consumption required to maintain the hyper- proliferative, anti- apoptotic, endothelial phenotype associated with increased PBF and pressure plays a significant role in the loss of NO signaling by attenuating hsp90 activity. Our data implicate RhoA/ROCK signaling as a master-regulator of these pathways. Studies in our lab have shown that ligation of S1PR3 receptor, induces Rho GTPase signaling and cytoskeletal remodeling. Interestingly, S1PR1 receptor, which exerts a protective effect against mechanical stress, is significantly downregulated in the lungs of our Shunt lamb model, while S1PR3 receptor expression is increased. We hypothesize that a mechanical stress mediated activation of an S1PR3 receptor-RhoA/ROCK axis is responsible for the mitochondrial fission, autophagy/mitophagy, and apoptosis that synergize to produce the hyper-proliferative, anti-apoptotic endothelial phenotype and the loss of NO signaling. Three Specific Aims (SAs) are proposed to test this hypothesis. In Aim 1, we will define the role of mitochondrial fission in the development of a hyper-proliferative endothelial phenotype and determine how this modulates NO signaling. In Aim 2, we will characterize the role played by autophagy/mitophagy in the loss of NO signaling under conditions of increased PBF and pressure and investigate the role played by the anti- apoptotic factor, survivin (Birc5). In Aim 3, we will determine whether targeting mitochondrial fission and autophagy are potential therapeutic targets in our pre-clinical Shunt lamb model. With the completion of this study, we will significantly increase our understanding of the role played by mitochondrial network remodeling and autophagy/mitophagy in the pathogenesis of pulmonary vascular disease associated with increased PBF and pressure while highlighting the application of novel therapeutic interventions.