mTOR coordinates cell metabolism, growth and survival in pulmonary hypertension

描述（由申请方提供）：肺动脉高压（PAH）是一种进行性致死性疾病，表现为肺动脉（PA）血管重塑、右心室后负荷升高、右心衰竭和死亡。肺动脉血管平滑肌细胞（PAVSMC）增殖增强和凋亡受损是PAH血管重塑的关键病理生理学组成部分，其分子机制尚未完全了解。该建议的重点是两个功能不同的复合物的哺乳动物雷帕霉素靶蛋白（mTOR），mTORC 1和mTORC 2，作为新的调节剂的PAVSMC的代谢，生长和生存的PAH。通过使用来自特发性PAH受试者（人PAH PAVSMC）和未使用的供体肺的PAVSMC，我们表明培养的人PAH PAVSMC保留了体内PAH肺中报告的分子和细胞异常，如增殖、存活和糖酵解代谢引起的细胞ATP水平改变，为翻译机制研究提供了独特的工具。我们发表的研究表明，慢性缺氧条件下PAVSMC增殖增加需要mTORC 1和mTORC 2的激活。我们的初步数据表明，mTORC 1和mTORC 2信号通路上调，在体内和体外的PAVSMC从小PA从受试者与特发性PAH和慢性缺氧诱导的肺血管重塑大鼠。我们的数据还表明，mTORC 1和mTORC 2促进增殖，但只有mTORC 2调节细胞能量水平和细胞存活，并表明抑制mTORC 1和mTORC 2是必需的，以抑制生长，增殖和促进人PAH PAVSMC的凋亡。基于已发表的研究和我们的数据，我们假设PAH中PAVSMC的增殖和存活增加需要mTORC 1（促进蛋白质合成和细胞生长）和mTORC 2（激活Akt，增加细胞能量水平，下调AMPK和增加细胞存活）的激活。我们还提出，在mTORC 1和mTORC 2中靶向mTOR对于抑制PAVSMC生长、增殖、促进细胞凋亡以及预防或消除PAH中的肺血管重塑是必要的。为了检验这一假设，在目标1中，将使用PAH患者和健康供体的肺组织样本以及特发性PAH患者和未使用供体肺的培养PAVSMC严格检测mTORC 1和mTORC 2信号通路的激活;在目标2中，我们将确定mTORC 1和mTORC 2是否采用差异分子机制来调节人PAH PAVSMC的生长、增殖和存活。将检查mTORC 1和mTORC 2在调节细胞生长、细胞ATP水平和细胞凋亡中的特定作用;在目标3中，我们将评估抑制mTORC 1和mTORC 2是否抑制生长、细胞能量水平在体外人PAH PAVSMC中增殖并诱导凋亡，并在PH大鼠慢性缺氧模型中预防或消除体内肺血管重塑。mTOR在调节PAH PAVSMC的能量水平、生长、增殖和存活中的作用，并将探索mTOR是否可以作为治疗人类PAH的潜在分子靶点。

DESCRIPTION (provided by applicant): Pulmonary arterial hypertension (PAH), a progressive fatal disease, manifests by vascular remodeling of pulmonary arteries (PA), elevated right ventricular afterload, right heart failure and death. Enhanced proliferation and impaired apoptosis of pulmonary arterial vascular smooth muscle cells (PAVSMC) are key pathophysiological components of vascular remodeling in PAH, the molecular mechanisms of which are not fully understood. This proposal focuses on two functionally distinct complexes of mammalian target of rapamycin (mTOR), mTORC1 and mTORC2, as novel regulators of PAVSMC metabolism, growth and survival in PAH. By using PAVSMC from subjects with idiopathic PAH (human PAH PAVSMC) and unused donor lungs, we show that cultured human PAH PAVSMC retain molecular and cellular abnormalities reported in PAH lungs in vivo, such as elevated proliferation, survival, and altered cellular ATP levels due to glycolytic metabolism that provides a unique tool for translational mechanistic studies. Our published study demonstrates that increased PAVSMC proliferation under chronic hypoxia requires activation of both mTORC1 and mTORC2. Our preliminary data show that mTORC1 and mTORC2 signaling pathways are up-regulated in vivo and in vitro in PAVSMC from small PA from subjects with idiopathic PAH and from rats with chronic hypoxia-induced pulmonary vascular remodeling. Our data also show that both mTORC1 and mTORC2 promote proliferation, but only mTORC2 modulates cellular energy levels and cell survival and suggest that inhibition of both mTORC1 and mTORC2 is required to suppress growth, proliferation and promote apoptosis in human PAH PAVSMC. Based on published studies and our data, we hypothesize that increased proliferation and survival of PAVSMC in PAH requires activation of both mTORC1 (promoting protein synthesis and cell growth) and mTORC2 (activating Akt, increasing cellular energy levels, down-regulating AMPK and increasing cell survival). We also propose that targeting mTOR in both mTORC1 and mTORC2 is necessary to inhibit PAVSMC growth, proliferation, promote apoptosis, and prevent or abrogate pulmonary vascular remodeling in PAH. To test this hypothesis, in Aim 1, activation of mTORC1 and mTORC2 signaling pathways will be critically tested using lung tissue samples from PAH patients and healthy donors and cultured PAVSMC from idiopathic PAH patients and unused donor lungs; in Aim 2, we will determine whether mTORC1 and mTORC2 employ differential molecular mechanisms to modulate growth, proliferation and survival of human PAH PAVSMC. Specific roles of mTORC1 and mTORC2 in regulating cell growth, cellular ATP levels and apoptosis will be examined; in Aim 3, we will evaluate whether suppression of both mTORC1 and mTORC2 inhibits growth, cellular energy levels, proliferation and induces apoptosis in vitro in human PAH PAVSMC and prevents or abrogates pulmonary vascular remodeling in vivo in a rat chronic hypoxia model of PH. Proposed studies will define the role of mTOR in regulating energy levels, growth, proliferation and survival of PAH PAVSMC and will explore whether mTOR could serve as a potential molecular target for treatment of human PAH.