胎儿肺组织来源间充质干细胞的分离、培养、鉴定及重编程初探

Isolation, Proliferation, Identification and Reprogramming of Mesenchymal Stem Cells Derived from Fetal Lung

该文主要研究将进行胎儿肺部组织来源的间充质干细胞(mesenchymal stem cells derived from fetal lung,FL-MSCs)转变成为诱导多能干细胞(induced pluripotent stem cells,iPS细胞)。首先使用酶消化法对胎儿肺部组织进行分离,然后采用常规方法进行培养并成功获得成纤维细胞样细胞。使用共聚焦技术检测获得的细胞,发现角蛋白表达呈阴性;共聚焦技术检测c-Myc、Oct4、Nanog以及Nestin四个干性相关因子,发现它们呈阳性;检测成纤维细胞样细胞的免疫表型,符合间充质干细胞的表型判断标准;然后进行诱导分化实验,发现这些细胞可以向成脂、成骨细胞分化,经过以上实验鉴定获得的成纤维细胞为FL-MSCs。使用Yamanaka四因子体系对FL-MSCs进行诱导,可以形成类似人胚胎干细胞(human embryonic stem cells,hES细胞)的克隆,采用核型分析、STR检测分析以及畸胎瘤形成实验初步验证获得的克隆为iPS。

The research is about to reprogramme mesenchymal stem cells derived from fetal lung （FL- MSCs） into induced pluripotent stem cells （iPS cells）. At first, we used trypsin to digest the tissue block derived from fetal lung, and then we obtained fibroblast-like cells through the conventional culture method. Detected the immunophenotype and differentiation potential of fibroblast-like cells to confirm that they were FL-MSCs. The expression level of keratin was demonstrated by confocal immunoflurescence, and it was negative. The expression levels of c-Myc, Oct4, Nanog and Nestin were demonstrated by confocal immunoflurescence, and they were all positive. Then we used Oct4, Sox2, Klf4 and c-Myc to transfect FL-MSCs and got the human embryonic stem cells-like （hES-like） clones. We used karyotype and STR analysis to prove that these clones were derived from the FL-MSCs, and then we got the preliminary evidence about that these clones were iPS by teratoma experiment.