间充质干细胞外泌体传递miR-125b调控ALI过度炎症反应的机制研究

Acute lung injury (ALI) is considered to be the major cause of respiratory failure in critically ill patients, which lacks effective intervention. Our recent study found mensenchymal stem cell (MSC) derived exosomes could restore ALI, However, the exact molecular mechanisms involved in this process remain unclear. Through microarrays single-channel fluorescence chip and bioinformatics analysis, we found that miR-125b was significantly increased in MSC derived exosomes and Traf6 was one of its target genes. It might be involved in the regulation of the inflammatory response of ALI. Using recombinant lentivirus vector technology, low and high miR-125b expression MSC derived exosomes will be built. The effects of MSC derived exosomes with different expression level of miR-125b on the lung injury induced by endotoxin will be observed in vitro and in vivo, respectively. Using luciferase reporter vector, we will demonstrate that miR-125b siginificantly block the expression of Traf6. Additionally, we will also explore the effect of miR-125b on the NF-κB signal pathway, which is downstream of Traf6. The present work will disclose the protective role and its underlying mechanisms of MSC derived exosomes on ALI induced by endotoxin and lay a foundation for the prevention and treatment of ALI.

急性肺损伤(ALI)是导致危重患者呼吸衰竭的主要原因之一,目前治疗效果欠佳。我们前期研究证实间充质干细胞（MSC）外泌体能修复ALI，但具体机制未明。我们采用miRNA芯片和生物信息学分析，发现MSC外泌体中miRNA-125b表达明显上调，且靶标于Traf6基因，推测其可能参与调控ALI的炎症反应。本项目拟通过慢病毒转染技术分别构建miR-125b过表达和miR-125b抑制表达的MSC外泌体，在LPS诱导的细胞和小鼠模型中，观察不同miR-125b表达水平的MSC外泌体对ALI过度炎症反应的影响；并通过荧光素酶报告基因分析技术验证miR-125b与Traf6的标靶关系，深入探讨miR-125b对Traf6下游NF-κB信号通路的影响，初步阐明MSC外泌体对ALI的修复机制，为临床防治ALI提供新的理论基础和实验依据。

间充质干细胞外泌体能抑制过度炎症反应，修复急性肺损伤时受损的肺组织。其发挥作用的机制与靶向传递功能性的miRNA密切相关。本课题通过体外研究发现小鼠骨髓间充质干细胞外泌体能降低LPS诱导的Raw264.7细胞促炎因子水平，升高抗炎因子水平。结合miRNA芯片分析,证实间充质干细胞外泌体与对照组成纤维细胞外泌体相比miR-125b水平明显增高。采用慢病毒感染间充质干细胞，构建miR-125b过表达和抑制表达的外泌体，在细胞水平和动物水平证实间充质干细胞外泌体通过分泌miR-125b传递至巨噬细胞，靶向结合Traf6，抑制NF-κB信号通路，逆转过度炎症反应，修复受损肺组织。综合上述结果，本研究为探讨间充质干细胞外泌体治疗急性肺损伤的机制奠定了坚实的工作基础，同时对于认识miR-125b和Traf6基因的标靶关系提供了重要的启示性线索。项目资助发表SCI论文3篇，欧洲呼吸年会论文1篇，中华结核呼吸杂志论文1篇，待发表SCI论文3篇。获得实用新型专利2项。培养硕士生2名。项目投入经费18万元，支出5.285万元，各项支出已根据实际情况进行调整。剩余经费计划用于本项目研究后续支出。