HGF-ATX-LPA信号通路介导HGF基因修饰MSCs保护放射性肠损伤的机制研究

Effective therapeutic strategies to radiation-induced intestinal injury (RIII) are currently lacking. Mesenchymal stem cells engraftment played important roles in intestine injury. We have found that treatment of RIII with HGF-gene-modified MSCs (MSCs-HGF) reduces local inflammation and promotes the recovery of small intestinal histopathology in a mouse model but mechnisms remained unkown. Further studies have found that HGF could up-regulted the expression of autotaxin (ATX) and the experssion of lysophosphatidic acid receptoe 2 (LPA2) of intestine increased after MSCs-HGF treatment. But the specific mechnisms of HGF-ATX-LPA signal pathway on therapy required clarify. In this study, we will evaluate the regulation of HGF-ATX-LPA to the effect of MSCs-HGF on RIII and clarify the underling mechnism. The main reseach contents of this study including establish normal and LPA1-/- and LPA2-/- mouse RIII model; Construct ATX, LPA1 or LPA2 overexpression, shRNA, mutants and RFP co-expressed lentivirus vector and preparation virus; Evaluate the effects of HGF-ATX-LPA axis on the protective role of MSCs-HGF on radiation-injured intestine tissues by using western blot, immunostaining and FACS; By using signal pathway inhibitor and agonist to verify other signaling molecules involved in the network of HGF-ATX-LPA pathway. This study may be a breakthrough point for fundamentally improving the efficacy of MSCs to treat radiation-induced pulmonary injury and expanding its therapy range.

放射性肠损伤（RIII）无有效治疗策略，间充质干细胞（MSC）移植等生物治疗成为实验阶段主要防治措施，我们前期研究发现肝细胞生长因子基因修饰MSC（MSC-HGF）可通过调节免疫及促进肠组织恢复而发挥保护作用，HGF可上调自分泌运动因子（ATX）表达，细胞治疗后小肠组织溶血磷脂酸受体2 （LPA2）表达上调，但HGF-ATX-LPA信号通路的具体作用机制尚待阐明。针对上述科学问题，我们拟建立放射诱导正常及LPA受体敲除小鼠RIII模型；研究内容包括ATX、LPA1及LPA2高表达、干涉及基因突变慢病毒载体构建；通过免疫印迹、免疫组化及流式细胞术等阐明上述操作对于MSC-HGF保护RIII的影响；通过多种信号通路阻断剂确定参与该信号调控的网络。上述研究将阐明HGF-ATX-LPA信号在MSC-HGF保护RIII中的作用及机制。该科学问题的阐明将有助提高干细胞治疗放射损伤的疗效，并提供新药靶。

放射性肠损伤（RIII）无有效治疗策略，间充质干细胞（MSC）移植等生物治疗成为实验阶段主要防治措施。本项目中，我们首先阐明肝细胞生长因子（HGF）可以通过MAPK和AKT信号通路激活自分泌运动因子/溶血磷脂酸（ATX/LPA）轴；通过肠上皮细胞IEC-6体外照射模型，我们发现HGF可以保护60Co gamma射线照射引起的肠上皮细胞凋亡，对其增殖没有影响。携带ATX的重组腺病毒可以高效介导ATX在MSCs细胞高表达，ATX修饰不影响MSCs细胞的表型，可促进MSCs细胞增殖，保护由于缺氧与乏血清所导致的细胞凋亡，可通过LPAR2/3调控MSCs细胞迁移。ATX可以通过抑制铁死亡诱导剂Erastin诱导的脂质ROS聚集而抑制铁死亡。ATX来源的外泌体对放射损伤具有保护作用。通过建立放射性肠损伤动物模型，评价了HGF基因修饰MSCs的干预作用。发现HGF基因修饰的MSCs可以通过ATX/LPA轴调节免疫反应，减轻局部炎症，促进小肠组织修复，从而发挥救治作用。上述科学问题的阐明将有助提高干细胞治疗放射损伤的疗效，并提供新药靶。