NLRP3-EndMT途径参与机械通气相关性肺纤维化的机制研究

Mechanical ventilation is an indispensible treatment for patients with acute respiratory failure. However, it has recently been found that mechanical ventilation plays an important role in the pathogenesis of pulmonary fibrosis. In our preliminary study, we found that mechanical ventilation led to increased lung collagen content, increased expression of transforming growth factor-1 and mesenchymal marker (α-SMA). However, expression of endothelial marker VE-Cadherin decreased. Lung histology also demonstrated fibrosis formation and potential endothelial-mesenchymal transition (EndMT). In vitro direct mechanical stretch of primary cultured pulmonary vascular endothelial cells resulted in similar EndMT formation. On the other hand, we found that mechanical ventilation resulted in continuous activation of NLRP3 inflammasome. Knockdown of NLRP3 significantly reversed cyclic stretch-induced EndMT in primary cultured pulmonary vascular endothelial cells. On the basis of these results, the present proposal will firstly use genetically modified mice to verify that the NLRP3 inflammasome activation does contribute to mechanical ventilation-induced EndMT and pulmonary fibrosis. Furthermore, using co-immunoprecipitation (Co-IP) and high-throughput proteomic techniques, we tried to screen out the key proteins directly interacted with the activated NLRP3 protein in primary cultured pulmonary vascular endothelial cells exposed to mechanical stretch. The contribution of these key proteins in the pathogenesis of mechanical ventilation-induced EndMT and pulmonary fibrosis was finally clarified. The present study will provide reliable evidence to elucidate the mechanisms involved in the pathogenesis of mechanical ventilation-induced pulmonary fibrosis and provide a theoretical basis for the prevention and treatment of mechanical ventilation associated lung injury.

机械通气虽然可以提供有效的呼吸支持，但也可以促使肺间质发生纤维化病变。我们在机械通气诱导肺纤维化的小鼠模型中发现，机械通气可以诱导肺组织出现内皮-间质转化，同时伴随肺组织中NLPR3炎性体的持续激活。体外实验则发现，机械牵拉可以直接诱导肺血管内皮细胞中NLRP3炎性体激活、并发生内皮-间质转化，而敲低NLRP3表达则可显著逆转这一现象。在此基础上本项目将首先采用NLRP3基因修饰小鼠证实NLRP3炎性体激活在机械通气诱导内皮-间质转化和肺纤维化中的关键作用；然后将结合免疫共沉淀和高通量蛋白质组学研究方法，在机械牵拉处理的肺血管内皮细胞中筛选得到能够与激活的NLRP3相互作用的关键蛋白，最终验证该蛋白信号转导通路在机械通气诱导内皮-间质转化和肺纤维化中的作用。本研究将有助于阐明机械通气诱导肺纤维化的发生机制，为临床防治机械通气肺损伤提供相关理论依据。

机械通气虽然可以提供有效的呼吸支持，但也可以促使肺间质发生纤维化病变。我们在机械通气诱导肺纤维化的小鼠模型中发现，机械通气可以诱导肺组织出现内皮-间质转化，同时伴随肺组织中NLPR3炎性体的持续激活。体外实验则发现，机械牵拉可以直接诱导肺血管内皮细胞中NLRP3炎性体激活、并发生内皮-间质转化，而敲低NLRP3表达则可显著逆转这一现象。在此基础上本项目首先采用NLRP3基因修饰小鼠证实NLRP3炎性体激活在机械通气诱导内皮-间质转化和肺纤维化中的关键作用；然后结合免疫共沉淀和高通量蛋白质组学研究方法，在机械牵拉处理的肺血管内皮细胞中筛选得到能够与激活的NLRP3相互作用的关键蛋白，最终验证该蛋白信号转导通路在机械通气诱导内皮-间质转化和肺纤维化中的作用。本研究将有助于阐明机械通气诱导肺纤维化的发生机制，为临床防治机械通气肺损伤提供相关理论依据。

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