脐带间充质干细胞移植对急性肝功能衰竭的修复及移植途径的优化

Therapeutic effects and approach optimizing of human umbilical cord mesenchymal stem cells transplantation on rat acute liver failure model

目的探讨脐带间充质干细胞(hUCMSC)移植对急性肝功能衰竭大鼠的治疗作用,并优化移植治疗途径。方法采用组织块贴壁法收集hUCMSC,流式细胞术检测细胞表面标志物。将SD大鼠随机分为经尾静脉注射移植治疗组,经肝叶注射移植治疗组,模型对照组,空白对照组。急性肝衰竭动物模型采用一次性腹腔注射2.5 mL/kg CCl4橄榄油(1:1)混合溶液建立,24 h后移植治疗组分别经尾静脉和肝叶注射hUCMSC细胞悬液;治疗后0h、24 h、48 h、72 h、96 h和1周收集血清分析肝功能恢复情况;治疗后3 d、1周、2周留取肝脏标本,观察病理学恢复情况,用Real-time PCR检测鼠肝中人CK8、CK18和AFP基因mRNA转录水平,免疫组化法检测人CK18的表达。结果移植治疗组TBil和ALT含量较模型对照组有明显恢复(P<0.05),肝细胞再生程度提高,炎细胞减少,肝脏病理学修复作用增强。在造模后1周、2周时,经尾静脉移植治疗组和经肝叶注射移植治疗组的肝组织中,CK8、CK18和AFP基因mRNA的相对转录量与模型对照组相比差异有统计学意义(P<0.05)。经尾静脉注射移植治疗组和经肝叶移植治疗组中,移植后的hUCMSC在鼠肝中诱导分化后表达人肝细胞相关蛋白CK18。经尾静脉和经肝叶注射移植治疗组的肝功能、移植细胞分化程度比较差异无统计学意义(P>0.05)。结论采用hUCMSC移植治疗急性肝衰竭大鼠模型后,能促进肝功能和肝脏病理学修复。移植后的hUCMSC自身能分化为具有肝细胞功能的类肝样细胞。经尾静脉与经肝叶注射移植疗效相似。

Objective To explore treatment effect and optimize the approach of human umbilical cord mesenchymal stem cells （hUCMSCs）transplantation on acute hepatic failure in rats.Methods hUCMSCs were isolated from umbilical cord with tissue explants adherent method,and the surface antigens of which were tested by flow cytometry.Forty-eight male Sprague Dawley rats were randomly divided into four groups,including transplantation treatment via tail vein group, transplantation treatment via hepatic lobe group,liver failure animal model group,and control group.Animal model of acute liver failure was induced by injecting intraperitoneally with 50% olive oil solution of carbon tetrachloride 2.5 ml·kg-1 ,then treatment groups were injected with hUCMSCs suspension separately through tail vein or injection directly into the liverhepatic lobe after 24h post-animal model built 24h.Blood serum and liver tissues were collected in several times to analyze liver function improvement and histological repair.Real-time PCR was used to detect the expression of human CK8, CK18 and AFP mRNA in liver tissues.Immunohistochemistry was used to detect the expression of human CK18 in liver tissues.Results There were statistically significant differences among liver function indexes after transplantation （P 0.05）.Conclusion There were statistically significant differences among liver function indexes after transplantation （P 0.05）.