lncRNA-miRNA-HMGB1在脂肪干细胞外泌体治疗树鼩脓毒症中的调控作用及机制研究

The inflammatory cascade reaction is one of the main mechanisms of multiple organ damage caused by sepsis. Previous results showed that mesenchymal stem cells (MSCs) could inhibit the expression of high mobility group box protein B1 (HMGB1), which induced and aggravated the inflammatory cascade as a key proinflammatory factor, but the regulatory mechanism is unclear. As the crucial therapeutic unit for stem cells in the damage microenvironment, MSCs-derived exosomes play multiple biological effects by its miRNA regulating target factors. According to the theory of competitive endogenous RNA, lncRNA sponges miRNA to regulate its target gene and construct an across-talk network of lncRNA-miRNA-mRNA. This study intends to construct the lncRNA-miRNA-mRNA worknet and further discuss the regulatory mechanism of lncRNA-miRNA-HMGB1 in the treatment of sepsis in non-human primate tree threws with Adipose Derived Stem Cells-derived Exosomes in vitro and in vivo. The effects of associated MAPKs/NF-κB signaling pathways will be also elucidated. The experimental results will provide a new idea and theoretical basis for the new target of sepsis treatment.

炎症级联反应是脓毒症器官损伤的主要机制之一。前期结果发现间充质干细胞（MSC）抑制关键炎症因子高迁移率族蛋白B1（HMGB1）的表达抑制炎症级联反应，但其调控HMGB1机制不清。间充质干细胞外泌体在损伤微环境中发挥关键作用，通过miRNAs调节炎症反应。根据竞争性内源RNA理论，lncRNA可作为miRNA“分子海绵”对其靶基因进行调控，构建lncRNA-miRNA-mRNA的调控网络。本课题拟运用全转录组测序法结合生物学分析、qRT-PCR等方法构建脂肪干细胞外泌体治疗非人灵长类树鼩脓毒症的lncRNA-miRNA-mRNA调控网络，进一步确定lncRNA-miRNA-HMGB1轴并在体内、体外实验中证实其在脂肪干细胞外泌体治疗脓毒症中的炎症调节作用及下游MAPKs、NF-κB信号通路机制。实验结果为临床上脓毒症的治疗新靶点提供新的观点及理论基础。

炎症级联反应是脓毒症器官损伤的主要机制之一。前期结果发现间充质干细胞（MSC）可以抑制关键炎症因子高迁移率族蛋白B1（HMGB1）的表达抑制炎症级联反应，但其调控HMGB1机制不清。间充质干细胞外泌体在损伤微环境中发挥关键作用，可调节炎症反应。本项目：（1）完成了脂肪间充质干细胞（ADSC）细胞培养与鉴定及其来源的外泌体的提取与鉴定；（2）建立了树鼩盲肠结扎穿孔脓毒症模型及评价体系；（3）体内动物证实了ADSC-exo治疗脓毒症树鼩对HMGB1的调节作用及机制；（4）体外细胞实验证实ADSC-exo治疗内毒素诱导的RAW264.7体外炎症模型对HMGB1的调节作用及机制；（5）初步探讨ADSC-exo治疗脓毒症树鼩的转录组水平作用及机制，获得差异表达的lncRNA、miRNA、mRNA，构建了炎症调节的 lncRNA-miRNA-mRNA 调控网络，进一步确定了lncRNA-miRNA-HMGB1 轴。实验结果阐述了间充质干细胞外泌体调节脓毒症的作用及机制，为临床上脓毒症的治疗新靶点提供新的观点及理论基础。