体外诱导人胎盘来源间充质干细胞向肝细胞分化  

In vitro differentiation of human placenta-derived mesenchymal stem cells into hepatocytes

作　　者：梁婷婷 陈黎 范明松 张国英 张世昌 Liang Tingting;Chen Li;Fan Mingsong;Zhang Guoying;Zhang Shichang(Department of Laboratory Medicine,The First Affiliated Hospital of Nanjing Medical University,Nanjing 210029,Jiangsu Province,China;Department of Obstetrics,The First Affiliated Hospital of Nanjing Medical University,Nanjing 210029,Jiangsu Province,China)

机构地区：[1]南京医科大学第一附属医院检验学部,江苏省南京市210029 [2]南京医科大学第一附属医院产科,江苏省南京市210029

出　　处：《中国组织工程研究》2022年第24期3870-3874,共5页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81671836),项目负责人:张世昌。

摘　　要：背景:间充质干细胞是治疗肝功能衰竭的重要细胞来源,将其诱导分化为肝细胞后能提高肝功能衰竭的治疗效果。目的:探讨胎盘组织来源间充质干细胞在体外诱导分化为成熟肝细胞的可行性,为其用于肝功能衰竭治疗奠定基础。方法:从人胎盘组织中分离培养人胎盘来源间充质干细胞,采用流式细胞分析、多向诱导分化等方法鉴定。应用多细胞因子将胎盘间充质干细胞向肝细胞诱导分化22 d,采用免疫荧光染色检测肝细胞标志物的表达变化,糖原染色检测细胞的糖原合成能力。结果与结论:①倒置显微镜下可见分离培养的细胞为梭形或成纤维细胞样细胞,增殖迅速,传10代后细胞形态未见明显变化;②流式细胞分析结果显示CD73、CD90、CD105、CD44、CD166、CD29、CD54、HLA-ABC在胎盘间充质干细胞中阳性率超过95%,而CD34、CD45、CD117、CD14、CD19、CD133、CD31、HLA-DR均为阴性;③经成脂、成骨、成软骨细胞诱导分化后,油红O、茜素红、阿辛蓝染色均为阳性;④向肝细胞诱导分化22 d后,呈圆形或多角形、细胞界限明显的典型肝细胞形态,免疫荧光染色显示大量细胞表达肝细胞表面标志物白蛋白、细胞角蛋白18、HepPar1、CYP7A1,糖原染色为阳性;⑤结果表明,从人胎盘组织分离培养的细胞为间充质干细胞,人胎盘间充质干细胞在体外可诱导分化为成熟肝细胞,有望为肝功能衰竭治疗提供可靠的肝细胞来源。BACKGROUND:Mesenchymal stem cells are an attractive source of therapies for liver failure.The differentiation of mesenchymal stem cells into hepatocytes can improve the therapeutic effect of liver failure.OBJECTIVE:To investigate the feasibility of placenta derived mesenchymal stem cells to differentiate into mature hepatocytes in vitro for the treatment of liver failure.METHODS:Placenta-derived mesenchymal stem cells were isolated and cultured from human placenta tissue.The surface markers and differentiation potential of the isolated cells were identified by flow cytometry,and multidirectional induction.Placenta-derived mesenchymal stem cells were induced into hepatocytes by several cytokines for 22 days.Hepatic markers of induced cells were detected by immunofluorescence staining.Glycogen staining was used to detect the glycogen synthesis ability of cells.RESULTS AND CONCLUSION:(1)Under the inverted microscopy,the cultured cells were spindle or fibroblast-like cells,which proliferated rapidly.There was no obvious change in cell morphology after 10 passages.(2)Flow cytometry results revealed that the positive rate of CD73,CD90,CD105,CD44,CD166,CD29,CD54,and HLA-ABC in placenta-derived mesenchymal stem cells was more than 95%,while CD34,CD45,CD117,CD14,CD19,CD133,CD31,and HLA-DR were all negatively expressed.(3)The results of oil red O,alizarin red,and alcian blue staining were positive after adipogenic,osteogenic and chondrogenic differentiation of placenta-derived mesenchymal stem cells.(4)After 22 days of hepatic differentiation,these cells exhibited typical morphology of hepatocytes such as round or polygonal shape with obvious cell boundary.Immunofluorescence staining showed that a large number of differentiated cells expressed albumin,CK-18,HepPar1,and CYP7A1 after induction.The positive staining of differentiated cells was observed by glycogen staining.(5)Our results demonstrated that the cells isolated from placenta were mesenchymal stem cells,and placenta-derived mesenchymal stem cells can differentiate

关 键 词：干细胞 间充质干细胞 胎盘 肝细胞 肝衰竭 成熟肝细胞 体外诱导 

分 类 号：R459.9[医药卫生—治疗学] R318.1[医药卫生—临床医学]