NADPH氧化酶/血管过氧化物酶1途径介导的氧化应激促肺动脉高压血管重构的作用机制

NOX/VPO1途径介导的氧化应激可能通过促进内皮损伤和平滑肌细胞增殖等方式参与肺动脉高压(PAH)血管重构的发生发展。本项目率先探讨NOX/VPO1途径促PAH内皮损伤和平滑肌细胞增殖的信号通路及其表达调节机制：1) 建立低氧PAH大鼠模型，检测血管形态，ROCK、NOX活性，NOX2、NOX4、VPO1表达和NMLC20 磷酸化水平等指标, 结合药物干预和基因沉默，确定NOX/VPO1途径促PAH血管重构作用；2）建立低氧诱导的内皮(祖)细胞损伤模型或平滑肌细胞增殖模型，检测与细胞损伤或增殖等指标, 结合药物干预或基因沉默，确定NOX/VPO1途径促血管内皮损伤和平滑肌细胞增殖作用及机制；3）利用上述动物和细胞模型，结合染色质免疫沉淀、荧光素酶报告基因、寡核苷酸pull-down等方法，阐明NMLC20对NOX2、NOX4表达调控机制。本项目将为阐明PAH血管重构机制提供新的理论基础。

It is likely that NOX/VPO1 pathway -mediated oxidative stress is involved in vascular remodeling of pulmonary artery hypertension (PAH) through promotion of endothelium injury and proliferation of smooth muscle cell. It is the first time for this project to explore the signal transduction of NOX/VPO1 pathway in promotion of endothelium injury and proliferation of smooth muscle cell in PAH and the regulatory mechanisms of expression for this pathway as follows: 1) Establish a rat model of hypoxia-induced PAH and finish the measurements of vascular morphology, activities of ROCK and NOX, expression levels of NOX2,NOX4 and VPO1, and phosphorylation level of NMLC20, etc. Through combining the strategies of drug interference and gene silence, the role of and potential mechanisms of NOX/VPO1 pathway in vascular remodeling of PAH will be evaluated; 2)Establish a cell model of hypoxia-induced endothelial (progenitor) cells injury (apoptosis or senescence)or proliferation of smooth muscle cell and finish the measurements of the cell injury (apoptosis or senescence) or proliferation associated parameters. Through combining the strategies of drug interference or gene silence, the role of NOX/VPO1 pathway in promoting vascular remodeling of PAH will be confirmed; 3) By using the above-mentioned PAH model and cell injury or proliferation model and combining the strategies of chromatin immunoprecitation, agarose-oligonucleotide pull-down assay, co-immunoprecipitation, luciferase reporter genes, etc, the mechanisms for NMLC20 in regulating NOX2,NOX4 gene expression will be clarified. This project will lay a novel theoretic foundation for understanding the mechanisms underlying the vascular remodeling of PAH.