Endothelium-derived Extracellular Vesicles, MicroRNAs and Pulmonary Hypertension

摘要 我们的长期目标是了解肺血管平滑肌细胞 在肺动脉高压（PH）的血管重构背景下调节PASMC（PASMC）增殖。在 肺血管内皮细胞（PVEC）调节肺血管内皮细胞的功能和增殖。 平滑肌细胞（PASMC）通过各种生物活性剂进入PASMC， 肌内皮连接和/或细胞外囊泡（EV）。核心假设是在缺氧时，PVEC- 衍生的EV含有不仅促进PASMC增殖而且抑制增殖的microRNA 正是这两种microRNA之间的不平衡导致了疾病。这项补助金的目的是 是为了表征EV及其microRNA货物在缺氧中的作用，在调节肺动脉粥样硬化的进展中， 我们的具体目标是：1）确定PVEC源性EV的作用以及 他们的两种miRNA货物miR-212- 5 p和miR-210- 3 p在调节缺氧诱导的血管重塑中的作用。 博士2)确定miR-212- 5 p抑制PASMC增殖的机制，特别是miR-212- 5 p的作用。 TNF受体相关因子3（TRAF 3）和TGF-β 1通路以及KLF 4在miR- 212- 5 p对PASMC增殖的影响。此外，miR-212- 5 p和miR-210- 3 p的新靶点将通过使用 iTRAQ质谱/蛋白质组学分析。3)提供概念验证，证明工程EV可用于 我们将设计EV携带miR-212- 5 p和anti-miR-210- 3 p，并确定它们是否可以 这一贡献是重要的，因为它将建立一个关键的作用， PVEC衍生的EV及其microRNA货物在PH发病机制中的作用;将描述miR-212- 5 p的作用， 肺血管重构和PH的重要负调节剂，并将测试利用 工程EV治疗严重PH。拟议的研究是创新的，知识来自 这些建议应该为进一步探索开辟新的领域，并使我们能够找到新的治疗战略， 重度PH

ABSTRACT Our long term goals are to understand the mechanisms by which pulmonary vascular smooth muscle cell (PASMC) proliferation is regulated in the context of vascular remodeling in pulmonary hypertension (PH). In the lung, the pulmonary vascular endothelial cell (PVEC) regulates the function and proliferation of the underlying smooth muscle cell (PASMC) by various bioactive agents that get to the PASMC via into the circulation, myoendothelial junctions and/or extracellular vesicles (EV). The central hypothesis is that in hypoxia, PVEC- derived EV contain microRNA that not only promote PASMC proliferation but also those that inhibit proliferation and it is the imbalance between these two types of microRNAs that leads to disease. The objective of this grant is to characterize the role of EV and their microRNA cargo in hypoxia, in modulating the progression of pulmonary vascular remodeling and PH. Our specific aims are 1) To determine the role of PVEC-derived EV and the roles of two of their miRNA cargo, miR-212-5p and miR-210-3p, in modulating vascular remodeling in hypoxia-induced PH. 2) To determine the mechanisms by which miR-212-5p inhibits PASMC proliferation; specifically the roles of TNF Receptor Associated Factor 3 (TRAF3) and the TGF- pathway and KLF4 in the inhibitory effect of miR- 212-5p on PASMC proliferation. In addition, novel targets of miR-212-5p and miR-210-3p will be identified using iTRAQ mass spectrometry/proteomics analysis. 3) To provide proof of concept that engineered EV can be used to treat severe PH. We will engineer EV to carry miR-212-5p and anti-miR-210-3p and determine if they can ameliorate PH in two rodent models of PH. This contribution is significant since it will establish a critical role for PVEC-derived EV and their microRNA cargo in the pathogenesis of PH; will delineate the role of miR-212-5p as an important negative regulator of pulmonary vascular remodeling and PH and will test the possibility of utilizing engineered EV for treatment of severe PH. The proposed research is innovative and knowledge derived from the proposal should open new areas for further exploration and enable us to find new treatment strategies for severe PH.