目的 探讨肝细胞生长因子的浓度和每日诱导时间对人骨髓间充质干细胞向肝细胞分化的影响.方法 密度梯度离心结合贴壁培养分离纯化骨髓间充质干细胞,将10、20、40和80 ng/mL肝细胞生长因子以每天0、3、6、12和24 h诱导骨髓间充质干细胞至第21天,荧光定量PCR检测白蛋白mRNA相对表达量,脲酶法检测培养上清液尿素浓度,过碘酸希夫试验进行糖原染色.结果 肝细胞生长因子浓度≥20 ng/mL且每日诱导时间≥6h的诱导组可检测到白蛋白mRNA表达、尿素分泌以及糖原合成.白蛋白表达与肝细胞生长因子浓度和每日诱导时间呈正相关(r=0.89,t=6.97,P＜0.05;r=0.77,t=5.85,P＜0.05).尿素分泌与肝细胞生长因子浓度和每日诱导时间呈正相关(r=0.91,t=7.27,P＜0.05；r=0.79,t=6.99,P＜0.05).糖原合成与肝细胞生长因子浓度和每日诱导时间呈正相关(r =0.93,t=8.33,P＜0.05；r=0.88,t=6.92,P＜0.05).结论 肝细胞生长因子体外诱导骨髓间充质干细胞向肝细胞方向分化以浓度≥20 ng/mL且每日诱导时间≥6h为宜,分化效果与肝细胞生长因子的浓度和每日诱导时间呈正向依赖性.%Objective To explore the influences of hepatocyte growth factor's concentration and induction time on the differentiation capability of human marrow mesenchymal stem cells (MSCs) into hepatocyte-like cells.Methods MSCs were isolated and purified by density gradient centrifugation and in plastic adherent culture.MSCs were induced by HGF of different concentrations and induction time per day until 21th day.The relative level of albumin mRNA was detected by quantitative PCR.The level of urea in supernatant was determined using urease method.Staining for glycogen was conducted with periodic acid Schiff test.Results Albumin mRNA,urea and glycogen synthesis were detected in groups with ≥20ng/ml HGF concentration and ≥6h induction time per day and none were detected in other groups.The expression of albumin had a positive correlation with HGF concentration and induction time per day (r =0.89,t=6.97,P＜0.05;r=0.77,t =5.85,P＜0.05).The secretion of urea had a positive correlation with HGF concentration and induction time per day (r=0.91,t=7.27,P＜ 0.05;r=0.79,t =6.99,P＜0.05).The synthesis of glycogen had a positive correlation with HGF concentration and induction time per day (r=0.93,t=8.33,P＜0.05;r=0.88,t=6.92,P＜0.05).Conclusion MSCs could be induced into hepatocyte-like cells by HGF under conditions of ≥20ng/ml HGF concentration and ≥6h induction time per day.The differentiation effects showed positive dependence on HGF concentration and induction time per day.
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