MSC释放的微粒改善ARDS肺微血管通透性的新机制 ——非S1P受体依赖的S1P-Rac1途径

Acute respiratory distress syndrome (ARDS) is characterized by the increase in lung microvascular permeability. Hence, it is critical to repair microvascular endothelial permeability in ARDS. Our previous study demonstrated that mesenchymal stem cell released microvesicle (MSC-MV) reduced inflammation, lung protein permeability and pulmonary edema in E.coli induced acute lung injury. Moreover, we found that hMSC-MV could restore protein permeability across injured human lung microvascular endothelial cell (HLMVEC). However, the underlying mechanisms for hMSC-MV restoring lung protein permeability were not fully understood. Our preliminary trials found that S1P (Sphingosine1-phosphate) levels in the HLMVEC and S1P related S1PK mRNA levels were elevated significantly to higher levels, but S1P levels in the cell culture medium was decreased after MSC-MV treatment, indicating that S1P may play an important role in the restoration of protein permeability across HLMVECs by hMSC-MV via a non-S1P-receptor dependent pathway: increase intracellular S1P and activate Rac1 signaling pathway. Therefore, we hypothesize that hMSC MV restore lung protein permeability via the Rac1 signaling pathway activated by intracellular S1P but not the S1P receptor -1 (S1PR1) dependent way. In this study, we investigate the role of S1P in hMSC MV treatment for ARDS and identify its mechanism whether it is associated with activation of Rac1 signaling pathway in endothelial cells in vitro study. What’s more, in vivo mouse model, we inject hMSC MV through tail to the S1PR1 ECKO (S1PR1f/f VE-cadherin-Cre-ERT2) mouse with LPS-induced lung injury, then we assess the lung permeability, the inflammation in lung tissue, lung injury and the activation of S1P pathway to demonstrate that the mechanism for MSC-MV to protect lung from ARDS is in part through S1P-Rac1 pathway but not relying on S1P receptor-1.

肺微血管内皮通透性增加是ARDS重要病理改变，保护内皮屏障、改善其通透性是治疗ARDS的关键。MSC释放的微粒（MSC-MV）能改善肺微血管通透性，但其机制不明。我们预实验发现MSC-MV增加内皮细胞内S1P水平却降低其培养上清液中S1P水平，加上胞内S1P-Rac1是S1P介导稳定内皮屏障的重要通路。推测MSC-MV可能通过非S1P受体依赖的S1P-Rac1途径发挥对内皮屏障的保护作用。本研究拟①体外在LPS诱导内皮细胞通透性模型之上，通过阻断内皮细胞S1PR1和干预其内Rac1表达，证实MSC-MV通过非S1P受体依赖的S1P-Rac1途径改善肺微血管通透性；②LPS气道滴入构建诱导性S1PR1基因敲除小鼠ARDS模型，观察MSC-MV对ARDS小鼠肺微血管通透性、肺部炎症、病理损伤及S1P通路活性的影响，阐明S1P-Rac1途径在MSC-MV改善肺微血管通透性和促进肺损伤修复中的意义

肺微血管内皮通透性增加是ARDS重要病理改变，保护内皮屏障、改善其通透性是治疗ARDS的关键。研究证实MSC释放的微粒（MSC-MV）能改善肺微血管通透性，然而其机制不明。本课题组体外在LPS诱导内皮细胞通透性模型之上，通过阻断内皮细胞S1PR1和干预其内Rac1表达，证实MSC-MV通过非S1P受体依赖的S1P-Rac1途径改善肺微血管通透性；此外，利用LPS气道滴入成功构建小鼠ARDS模型，进一步通过阻断S1P受体，明确了S1P-Rac1途径在MSC-MV改善ARDS小鼠肺微血管通透性和促进肺损伤修复中的作用和意义，为ARDS的治疗提供了新的靶点和方向。