以人皮肤成纤维细胞作为饲养层细胞培养人胚胎干细胞  

Culture of human embryonic cells by using human dermal fibroblasts as feeder cells

作　　者：杨俊林[1] 刘雄昊[1] 喻姣玲[1] 冯劢[1] 谭杨[1] 李卓[1] 吴奔[1] 高庭[1] 潘盛[1] 邬玲仟[1] 梁德生[1] 夏家辉[1] 

机构地区：[1]中南大学医学遗传学国家重点实验室,湖南长沙410078

出　　处：《现代生物医学进展》2009年第16期3001-3005,共5页Progress in Modern Biomedicine

基　　金：国家重点基础研究发展计划(2004CB518800);国家高技术研究发展计划(2007AA021002和2007AA021206);国家自然科学基金(30700458和30971298)

摘　　要：目的:比较人皮肤成纤维细胞(human dermal fibroblasts,HDFs)与小鼠胚胎成纤维细胞(Mouse embryonic fibroblasts,MEFs)的增殖能力及研究人皮肤成纤维细胞作为饲养层支持人胚胎干细胞(human embryonic stem cells,hESCs)未分化生长的能力。方法:利用组织贴壁法从人皮肤中分离出HDFs,通过细胞形态的观察和生长曲线的绘制比较HDFs与MEFs的体外增殖能力。将HDFs作为饲养层细胞与hESCs共培养,传代12代后,检测hESCs碱性磷酸酶(AKP)、表面特异性标志及胚胎干细胞特异性转录因子。结果:HDFs可连续传代培养15代以上,10代以下的HDFs增殖迅速,而MEFs自第4代起,增殖能力就明显下降;hESCs在HDFs饲养层上可传代培养12代以上,克隆边界清晰,细胞排列紧密,碱性磷酸酶、表面标志物检测均呈阳性,表达了hESCs特异性转录因子。结论:HDFs比MEFs具有更强的增殖能力;HDFs可作为培养hESCs的饲养层细胞。Objective： To compare the proliferative potential of human dermal fibroblasts（HDFs） and mouse embryonic fibroblasts （MEFs）, and to explore the feasibility for HDFs as feeder cells to support the growth of human embryonic cells （hESCs）. Methods： HDFs were obtained from human skin by using the method of tissue-adherence. The proliferative capacities of HDFs and MEFs were compared by cell morphology and growth curve. HDFs were used as feeder cells to culture hESCs, After growing on HDFs for 12 passages, the hESCs were tested for the expressions of alkaline phosphatase （AKP）, the cell surface markers and cell-specific transcription factor of hESCs. Results： HDFs could be subcultured for more than 15 passages, and retained strong proliferative potential before 10 passages, while it decreased obviously in MEFs after subcultured for 4 passages. The hESCs can passaged on HDFs feeder cells for 12 passages with tightly packed cells and clear boundaries of clone, showing the positive results of alkaline phosphatase （AKP） and the cell surface markers and the expression of hESCs-specific transcription factor. Conclusion： HDFs showed a stronger proliferation ability than MEFs; HDFs can support the growth of hESCs.

关 键 词：人胚胎干细胞 人皮肤成纤维细胞 饲养层细胞 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]