人的类胚胎干细胞的分离和培养

ISOLATION AND CULTURE OF HUMAN EMBRYONIC STEMLIKE CELLS

人的类胚胎干细胞的分离和培养①徐令１黄绍良１李树浓２周灿权３乔春平２张敏芳３（中山医科大学１孙逸仙纪念医院儿科２病理生理教研室３附属第一医院生殖中心；广州，５１０１２０）主题词胚胎；干细胞中图号Ｒ３２９．１自从Ｅｖａｎｓ和Ｋａｕｆｍａｎ［１，２］于１...

ince mouse embryonic stem (ES) cell line was first established by Evans and Kaufman in 1981,especially after success in gene transfer and nuclear transfer of animal, the study of ES cells become popular. ES cells are undifferentiated noncommitted cells from inner cell mass of early embryos and have the developmental potential to give rise to all cell types. Human ES cells also have the developmental potential to give rise to all adult cell types. They will have several beneficial uses, not only as a model in studying the events involved in cell differentiation and development in vitro, but also will supply materials of organ transplantation, organ and tissue repair. Human ES cells would offer exciting new possibility for transplantation medicine because any disease resulting from the failure of specific cell types would be treatable through the transplantation of differentiated cells derived from ES cells. Therefore, it has great value in studying human ES cells.Only Bongso has reported that inner cell mass cells from human blastocysts were isolated and cultured in human leukaemia inhibitory factor (LIF), but the cells died after two passages. They defeated to establish human ES cell line. We cultured the fertilize ovum (from a volunteer which is supplied by reproduction center of Sun Yatsen University of Medical Sciences) in Earle′s medium until morula (Fig.1) The zona pelluceda was removed and the cells which dispersed from the morula were grown in Earle′s medium with LIF for several days and cocultured on mouse embryonic fibroblasts in DMEM with BRL (Buffalo Rat Liver Cells) for about 10 days. The cells were grown very fast and tightly packed in large nest and colonies in which it was difficult to recognize individual cells. Each cell was small, round or fusiform, with large nucleus and minimal cytoplasm, and the nuclei contained one or more prominent nucleoli. The cells were coincided with mouse ES cells (Fig.2) and were continually grown after 5 passages. AKP was positive (Fig.3) and it suggests that the ESlike cells were undifferentiated. The method avoids culturing the fertilized ovum to blastocysts and is easy to isolate the earlier ES cells. It has benefit to establish human ES cell line.