重型肝病患者血清诱导脐血间充质干细胞表达白蛋白和细胞角蛋白18

背景:脐血富含间充质干细胞,刺激后进入细胞周期的速度以及自分泌生长因子的能力均强于骨髓及外周血干细胞。目的:观察肝衰竭患者血清对人脐血间充质干细胞的诱导分化作用。方法:密度梯度离心和贴壁培养法相结合分离纯化脐血间充质干细胞并鉴定,以20%肝衰竭患者血清诱导培养,培养7,14,21d采用免疫组织化学方法检测白蛋白和细胞角蛋白18的表达。结果与结论:实验分离获得了高纯度贴壁生长的间充质干细胞,人脐血间充质干细胞高表达CD44和CD29,不表达CD34,体外可以分化为脂肪细胞;人脐血间充质干细胞在肝病患者血清的低糖DMEM培养基中形态发生明显改变,镜下观察细胞变得稍大而扁平,呈类上皮样细胞,SP染色显示实验组培养7d时仅少数细胞表达白蛋白和细胞角蛋白18,培养14,21d,白蛋白和细胞角蛋白18表达阳性率逐渐升高,与对照组相比差异有显著性意义(P<0.05)。提示密度梯度离心和贴壁培养法相结合可以获得纯度较高的脐血间充质干细胞,肝衰竭患者血清可诱导间充质干细胞表达白蛋白和细胞角蛋白18。

Expression of albumin and cytokeratin 18 in human umbilical cord blood mesenchymal stem cells induced by serum of patients with server liver diastase

BACKGROUND:Studies have found that the cord blood contains a large amount of mesenchymal stem cells(MSCs).These cells have quicker speed of getting into cell cycle and stronger autocrine ability of growth factors than those in the bone marrow or peripheral blood after activation.OBJECTIVE:To investigate the differentiation ability of human umbilical cord blood MSCs(HUMSCs) by serum of patients with severe liver disease.METHODS:HUMSCs were isolated and purified by density gradient centrifugation and adherent culture.HUMSCs were induced to differentiate with 20% serum of liver failure patients.On days 7,14,21 after induction,the expressions of albumin and cytokeratin 18 were identified by immunohistochemical method.RESULTS AND CONCLUSION:High purity HUMSCs were isolated from umbilical cord blood.HUMSCs strongly expressed CD44 and CD29,but did not express CD34.HUMSCs may differentiate into adipocytes in vitro.Morphological changes happened in HUMSCs cultured in L-DMEM medium containing 20% serum of patients with severe liver disease,the cells became larger and flat showing epithelioid cells type.Streptavidin-perosidase showed that only few cells were positive for albumin and cytokeratin 18 in the experimental group after 7 days induction.The positive rates of albumin and cytokeratin 18 were increased after 14 and 21 days induction,which were significantly different between the experimental group and control group(P < 0.05).It is indicated that high purity HUMSCs can be isolated by density gradient centrifugation and adherent culture,and serum of liver failure patients can induce the albumin and cytokeratin 18 expression in HUMSCs.