骨髓间充质干细胞促进大鼠80%肝切除后残肝再生及修复作用的研究

Effects of Bone Marrow Mesenchymal Stem Cells on Liver Regeneration and Repair after 80% Hepatectomy in Rats

目的:观察骨髓间充质干细胞(BMSCs)对大鼠自体80%肝切除后残余肝脏再生及修复的作用。方法:采用密度梯度离心联合贴壁筛选法分离纯化鉴定BMSCs,取6～8周体质量200～220g的大鼠,建立大鼠80%肝切除模型,实验组16只,术后即刻经门静脉注射BMSCs;对照组20只,注射等量生理盐水。采用生物化学方法检测肝功能,流式细胞仪和免疫组化检测肝细胞周期和增殖细胞核抗原(PCNA)。结果:体外成功分离培养、扩增BMSCs;实验组术后生存15只,死亡1只,对照组生存15只,死亡5只;肝脏组织显示,实验组术后1、3和7d时肝窦轻度扩张,较少的炎性细胞浸润,少见坏死,坏死程度明显轻于对照组,术后3d差异尤为明显;实验组术后1、3和7d时血ALT和AST水平较对照组明显降低,差异有统计学意义(P<0.05);实验组术后1、3和7d时肝细胞周期中S期细胞比例与对照组相比均明显增高(P<0.05);残肝免疫组化显示实验组PCNA表达水平明显高于对照组(P<0.05)。结论:BMSCs可以促进大鼠80%肝切除后残余肝脏细胞的再生及修复。

Objective： To investigate the effects of rat bone marrow mesenchymal stem cells （BMSCs） on liver regeneration and repair after 80% hepatectomy.Methods： BMSCs were isolated and purified by density gradient centrifugation combined adherent method.The model of 80% hepatectomy was established with rats aged 6-8 weeks and weighted 200220 g.Experimental group contained 16 rats with BMSC injection via the portal veins,and control group included 20 rats with saline injection by the same way.The liver functions were tested by biochemical methods.The flow cytometry and immunohistochemistry were used to detect the hepatocyte cell cycle and proliferation cell nuclear antigen （PCNA）.Results： BMSCs were isolated,cultured and proliferated successfully in vitro.The survival rate was 93.7% after operation in the experimental group and 75% in control group.The liver tissues displayed that hepatic sinusoid mild expansion,less inflammatory cell infiltration and rare necrotic lesions after 1 d,3 d and 7 d operation,in the experimental group than those of the control group,especially 3 d after operation.The levels of aspartate aminotransferase （ALT） and aspartate aminotransferase （AST） were decreased more significantly in experimental group than those in control group （P 〈 0.05）.The S phase of hepatocyte cell cycle was significantly higher in experimental group than those in control group （P 〈 0.05）.The remnant liver immunohistochemistry showed that PCNA expression was significantly higher in the experimental group than that in the control group （P 〈 0.05）.Conclusion： BMSCs can improve rat liver regeneration and repair significantly after 80% hepatectomy.