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目的 :研究人胚肺成纤维细胞 (humanEmbryonicLungFibroblast,HELF)饲养层对人胚胎干细胞 (HumanEmbryonicStemCells ,hES细胞 )生长的作用 ,摸索适宜的hES细胞培养与识别方法。方法 :以流产胎儿肺组织制备HELF饲养层 ,取生殖嵴或背肠系膜及其周围组织 ,获得由人原始生殖细胞 (humanPrimordialGermCells,hPGCs)转化成的hES细胞。比较其在不同条件下的增殖行为 ,并通过生物学特性鉴别该细胞系。结果 :HELF饲养层在LIF存在的条件下有利于支持hES细胞的体外生长 ;hES细胞碱性磷酸酶组织化学染色及端粒酶ELISA检测阳性 ;具有正常的核型 ;在体外发展为类胚体。结论 :HELF饲养层和LIF的协同作用能支持hES细胞的生长 ;建立通过生物学特性鉴别hES细胞的方法。
OBJECTIVE: To study the effect of human embryonic lung fibroblast (HELF) feeder layer on the growth of human embryonic stem cells (hES) and explore the suitable method of hES cell culture and identification. Methods: Human fetal primordial germ cells (hPGCs) were transformed into hES cells by making HELF feeder layer from aborted fetus lung tissue and taking genital ridge or dorsal mesangial membrane and its surrounding tissues. The proliferative behavior under different conditions was compared and the cell line was identified by biological characteristics. RESULTS: The HELF feeder layer was able to support the in vitro growth of hES cells in the presence of LIF. The hES cells were positive for alkaline phosphatase staining and telomerase enzyme-linked immunosorbent assay (ELISA), with normal karyotypes, and developed in vitro as embryoid bodies . CONCLUSION: The synergistic effect of HELF feeder layer and LIF can support the growth of hES cells. A method of identifying hES cells by biological characteristics was established.