人参皂苷诱导骨髓间充质干细胞对急性肝功能衰竭大鼠肝组织再生及Wnt/β-catenin信号表达的影响

目的:分析人参皂苷诱导骨髓间充质干细胞（BMSC）对急性肝功能衰竭（ALF）大鼠肝组织再生及Wnt/β-catenin信号表达的影响。方法:选取SPF级Wistar雄性大鼠66只，其中1只行BMSCs培养，随机选取50只大鼠制备ALF模型，24 h后随机选取其中5只断颈处死，依据大鼠肝功能与组织病理学判别其是否成功造模。剩余15只大鼠将等剂量0.9%氯化钠溶液腹腔注入，作为正常组。成功造模的45只大鼠随机分成3组，分别为BMSCs组、模型组及人参皂苷诱导BMSCs组，每组各15只。造模后6 h，模型组和正常组大鼠由尾静脉注入1 mL细胞培养液，BMSCs组注入1 mL BMSCs悬液（细胞浓度2.0×109/L），人参皂苷诱导BMSCs组注入1 mL人参皂苷诱导分化BMSCs悬液（细胞浓度2.0×109/L），连续给药10 d。观察大鼠肝脏组织病理状况，血清总胆红素（TBil） 、AST、ALT、TNF-α、IL-6含量，肝组织内Wnt7b、Wnt7a及Wnt2基因表达。结果:正常组大鼠肝细胞无坏死、变性，整齐排列，肝小叶结构清晰，汇管区内没有炎性细胞浸润；模型组大鼠肝细胞表征为广泛坏死，肝小叶内全部肝细胞溶解坏死，网状的支架发生塌陷，汇管区和周边存在大量粒细胞、单核细胞与淋巴细胞的浸润，残余肝细胞变性、肿胀且伴随胆汁淤积；BMSCs组及人参皂苷诱导BMSCs组大鼠肝小叶结构趋向完整，肝细胞坏死、变性数量减少，汇管区内有少许炎性细胞浸润，人参皂苷诱导BMSCs组又优于BMSCs组。和正常组相比，模型组大鼠血清TBil、AST、ALT、IL-6及TNF-α含量上升；和模型组相比，BMSCs组、人参皂苷诱导BMSCs组大鼠血清TBil、AST、ALT、IL-6及TNF-α含量降低；和BMSCs组相比，人参皂苷诱导BMSCs组大鼠血清TBil、AST、ALT、IL-6及TNF-α含量降低，差异有统计学意义（P＜0.05）。和正常组相比，模型组大鼠肝组织内Wnt7b、Wnt7a、Wnt2 mRNA相对表达量降低；和模型组相比，BMSCs组、人参皂苷诱导BMSCs组大鼠肝组织内Wnt7b、Wnt7a、Wnt2 mRNA相对表达量升高；和BMSCs组相比，人参皂苷诱导BMSCs组大鼠肝组织内Wnt7b、Wnt7a、Wnt2 mRNA相对表达量升高，差异有统计学意义（P＜0.05）。结论:人参皂苷诱导BMSCs可显著改善ALF大鼠肝功能，其作用机制可能为激活Wnt/β-catenin信号通路加速肝脏再生。

Effects of ginsenoside-induced bone marrow mesenchymal stem cells on liver regeneration and Wnt/β-catenin signaling in rats with acute liver failure

AIM:To analyze the effects of ginsenoside-induced bone marrow mesenchymal stem cells (BMSC) on liver regeneration and Wnt/β-catenin signaling in rats with acute liver failure (ALF). METHODS:Sixty-six Wistar male rats of SPF grade were selected. One of them was cultured with BMSCs. Fifty rats were randomly selected to prepare ALF model.After 24 hours,5 of them were randomly selected and sacrificed.The success of the rat model was judged by liver function and histopathology.The remaining 15 rats were intraperitoneally injected with an equal dose of 0.9% sodium chloride solution as a normal group.Forty-five rats were randomly divided into three groups:BMSCs group,model group and ginsenoside-induced BMSCs group,with 15 rats in each group.The model group and the normal group were injected with 1 mL of cell culture medium from the tail vein and the BMSCs group was injected with 1 mL of BMSCs suspension (cell concentration:2.0×109/L).The ginsenoside-induced BMSCs group was injected with 1 mL of ginsenoside to induce differentiation of BMSCs (cell concentration 2.0×109/L).The solution was administered continuously for 10 days.The pathological conditions of liver tissue were observed, serum TBil,AST,ALT,TNF-α,IL-6 levels,Wnt7b,Wnt7a and Wnt2 gene expression in liver tissue.RESULTS:The normal group of rats had no necrosis,degeneration,neatly arranged,the structure of hepatic lobule was clear,and there was no inflammatory cell infiltration in the portal area.The hepatocytes in the model group were characterized by extensive necrosis,and all hepatocytes in the hepatic lobules were necrotic.The stent collapsed, and there were a large number of granulocytes, monocytes and lymphocytes infiltrated in the portal area and surrounding areas.The residual hepatocytes were degenerated,swollen and accompanied by cholestasis.The BMSCs group and ginsenoside-induced BMSCs group showed that the hepatic lobule structure became more intact.The number of hepatocyte necrosis and degeneration was reduced,and there was a little inflammatory cell infiltration in the portal area. The ginsenoside-induced BMSCs group was superior to BMSCs.Compared with the normal group,serum TBil,AST,ALT,IL-6 and TNF-α level in the model group increased. Compared with the model group, serum TBil,AST,ALT,IL-6 and TNF-α level in the BMSCs group and ginsenoside-induced BMSCs group decreased.Compared with BMSCs group,the levels of serum TBil,AST,ALT,IL-6 and TNF-α in ginsenoside-induced BMSCs group were significantly lower (P<0.05).Compared with the normal group,the relative expression of Wnt7b,Wnt7a and Wnt2 mRNA in the liver tissue of the model group was decreased.Compared with the model group,the relative expression of Wnt7b,Wnt7a and Wnt2 mRNA in the BMSCs group and ginsenoside-induced BMSCs group increased.Compared with BMSCs group,the relative expression of Wnt7b,Wnt7a and Wnt2 mRNA in the liver tissues of rats in ginsenoside-induced BMSCs group increased,the difference was statistically significant (P<0.05). CONCLUSION:Ginsenoside-induced BMSCs can significantly improve liver function in ALF rats,and its mechanism may be related to activating Wnt/β-catenin signaling pathway to accelerate liver regeneration.