维甲酸对MESPU35胚胎干细胞系神经定向分化的影响

Effect of retinoic acid on differentiation of embryonic stem cell line MESPU35

目的 了解维甲酸 (Retinoicacid ,RA)对MESPU3 5胚胎干细胞系诱导分化的影响。方法 小鼠胚胎成纤维细胞作为饲养层 ,培养MESPU3 5细胞系 ,4 4+法诱导其神经定向分化。结果 相差显微镜观察RA诱导MESPU3 5细胞系出现神经样细胞拟胚体百分率随分化时相点和RA浓度升高而升高 ,免疫细胞化学观察分化的NF2 0 0和GFAP阳性细胞随分化时相点和RA浓度升高而增加 ,NF2 0 0阳性细胞由无突起变为多极细胞 ,GFAP阳性细胞突起由短变长 ,最后联结成网状。流式细胞仪检测分化的GFAP、NF2 0 0阳性细胞比例变化与免疫细胞化学结果相符。结论 RA诱导MESPU3 5细胞系神经定向分化的调节以浓度和时间依赖模式进行 。

QIN Mao lin, CAI Wen qin, YAO Zhong xiang (Department of Histology and Embryology, Third Military Medical University, Chongqing 400038, China) Abstract: Objective To explore the effect of retinoic acid(RA) on neural differentiation of embryonic stem cell line MESPU35. Methods MESPU35 ES cells cultured on feeder cells (mouse embryonic fibroblast) were induced by RA(4 /4 + method). Results By using RA treatment, it was found by phase contrast microscope that the percentage of outgrowths of embryonic bodies containing neuron like cells increased as the concentration of RA and days of differentiation increased. Immunocytochemistry results showed numbers of NF200 positive cells and GFAP positive cells increased with the increases of the concentrations of RA and time of differentiation. NF200 positive cells changed from spherical shape to multipolar shape. Processes of GFAP positive cells became longer, connected with each other and formed a network. The results by flow cytometry were in conformity with those by immunocytochemistry. Conclusion The differentiation of ES cells can be induced by RA in dose dependent manner and time dependent manner, suggesting that MFSPU35 cells can provide a source of differentiated nerve cell types that can be used for transplantation.