羊水间充质干细胞体外培养方案的优化及生物学特性

背景：目前干细胞工程种子来源多样化,羊水中存在胎儿脱落细胞,可分离培养出间充质来源干细胞。目的：探索不同成分培养基对人羊水间充质干细胞（human amniotic fluid-derived mesenchymal stem cells,h-AFMSCs）体外培养的影响,并分析最适培养基体外培养的h-AFMSCs的生物学特性,建立优化的h-AFMSCs培养方案。方法：孕16～24周的孕妇产前检查中获得羊水进行原代及传代培养并采用6种不同的培养基成分对h-AFMSC进行培养。结果与结论：从孕中期羊水中可分离出h-AFMSCs,体外使用低糖DMEM培养基（L-DMEM）＋体积分数10%胎牛血清与合成培养基MESEN PRO体积比1：1配制的培养基对其扩增促进作用最强。h-AFMSCs高表达CD29、CD73、CD90、CD105、CD166,低表达CD14、CD34、CD45及HLA-DR;分离培养的h-AFMSCs高表达的干细胞基因为OCT-4和Nanog;h-AFMSCs体外具有向成骨、成脂细胞分化的潜能且具有抑制淋巴细胞增殖的作用。提示孕中期羊水是低免疫原性的h-AFMSCs的良好细胞来源。

Biological characteristics and optimal strategy of human amniotic fluid-derived mesenchymal stem cells cultured in vitro

BACKGROUND：At present,seed sources of stem cells engineering are diverse,and mesenchymal stem cells can be isolated from exfoliated cells in fetal amniotic fluid.OBJECTIVE：To investigate the effects of the media with different elements on human amniotic fluid-derived mesenchymal stem cells（h-AFMSCs） cultured in vitro,and to analyze the biological characterization of h-AFMSCs at optimum medium cultured in vitro in order to optimize the culture strategy of h-AFMSCs.METHODS：Amniotic fluid was extracted from pregnant women during 16 to 24 weeks of gestation,and then was used for primary culture and subculture.h-AFMSCs were cultured in media containing six different elements.RESULTS AND CONCLUSION：h-AFMSCs were isolated from amniotic fluid in mid-trimester.The proliferative ability of h-AFMSCs was improved to a maximal extent in the mixed medium of low glucose Dulbecco＇s modified eagle medium containing 10% fetal bovine serum and MESEN PRO medium at a mixed ratio of 1：1.h-AFMSCs highly expressed CD29,CD73,CD90,CD105,CD166 and lowly expressed CD14,CD34,CD45,HLA-DR.Stem cells gene OCT-4 and Naong highly expressed in h-AFMSCs cultured in vitro.h-AFMSCs showed a higher osteogenic and adipogenic differentiation potential,and they could inhibit the proliferation of lymphocytes in vitro.It is indicated that mid-trimester amniotic fluid is a good cell source of h-AFMSCs with lower immunogenicity.