肝匀浆上清液诱导人脐带间充质干细胞分化为肝样细胞

摘要

BACKGROUND:Preliminary experiments have demonstrated that rat liver homogenate supernatants can induce the morphological changes of human umbilical cord mesenchymal stem cells. However, little is known about whether the induced cells have some phenotypic and functional features of hepatocytes. OBJECTIVE:To investigate whether human umbilical cord mesenchymal stem cells have some phenotypic and functional characteristic of hepatocytes after being induced by liver homogenate supernatants. METHODS:Passage 3 human umbilical cord mesenchymal stem cells were used and divided into control group (cells were cultured in basic culture medium) and liver homogenate supernatant group (cells were cultured in liver homogenate supernatants for 3, 5, 7 days). Meanwhile, positive control group (QSG-7701 human liver celllines) and negative control group (simple liver homogenate supernatants) were set up. The protein and mRNA level of hepatocyte markers, alpha-fetoprotein, cytokeratin 18 and tryptophan 2,3-dioxygenase enzyme, were detected at different time points. RESULTS AND CONCLUSION:After inducement, the stem cells of fusiform shape began to lose their sharp edges and progressively shrunk, and then they changed into hepatocyte-like cells with the morphous of triangle, polygon and anomalism shape. Compared with the control group, the protein and mRNA level of alpha-fetoprotein, cytokeratin 18 and tryptophan 2,3-dioxygenase enzyme significantly increased time dependently after inducement with liver homogenate supernatants (P<0.01). This study demonstrated that human umbilical cord mesenchymal stem cells are able to differentiate into hepatocyte-like cells in vitro that possess some functions of liver cells.%背景：前期实验证明大鼠肝组织匀浆上清液能诱导人脐带间充质干细胞发生形态学变化，但诱导后细胞是否具有肝细胞的表型和功能特征尚不清楚。　　目的：以人脐带间充质干细胞为研究对象，对肝组织匀浆上清液定向诱导分化后细胞是否具有肝细胞的表型和功能特性进行初步验证。　　方法：选取第3代人脐带间充质干细胞，采用肝组织匀浆上清液作为诱导培养基，进行如下分组：单纯肝组织匀浆上清液作为阴性对照组，QSG-7701人肝细胞株作为阳性对照组，基础培养的人脐带间充质干细胞作为标准对照组；肝组织匀浆上清液处理3，5，7 d诱导组；在蛋白水平和转录水平对肝细胞特异性标志物甲胎蛋白、角蛋白18和色氨酸2，3-加双氧酶进行检测。　　结果与结论：经肝组织匀浆上清液诱导后，细胞形态发生了显著变化，随诱导时间延长梭形干细胞逐渐减少，细胞呈三角形，多角形或不规则形。肝组织匀浆上清液诱导干细胞3，5，7 d后肝细胞标志物甲胎蛋白、人细胞角蛋白18和色氨酸2，3-加双氧酶在蛋白和mRNA水平均显著高于标准对照组(P<0.01)。实验结果提示在体外经肝组织匀浆上清液诱导后，人脐带间充质干细胞能够分化为具有肝细胞特异性标记的肝样细胞，并具有部分肝细胞功能。