无饲养层条件原代培养小鼠胚胎干细胞的实验研究

Culture of Mural Embryonic Stem Cells Under Feeder-free Condition

目的:对无饲养层条件下原代分离培养小鼠胚胎干细胞(ESC)的可行性进行研究。方法:利用微滴培养技术,将小鼠胚泡分别置于无饲养层条件及人胚胎成纤维细胞(HEF)饲养层上进行培养。观察胚泡的发育行为,并对两种不同条件下ESC的克隆率进行研究。结果:在无饲养层及HEF饲养层条件下小鼠囊胚均可以正常发育,并可以原代培养出ESC克隆,但无饲养层体系的胚泡48 h贴壁率(71.93%)及原代ESC克隆率(5.26%)低于HEF饲养层体系(88.71%,19.35%)(P<0.05)。结论:无饲养层条件下可以成功原代分离培养获得小鼠ESC,但该条件下ESC的分离效率仍需进一步研究。

Objective：To study the isolation and primary culture of mural embryonic stem cells （ESC）under feeder-free condition. Methods：With the microdrop culture technique, blastocysts were cultured under feeder-free condition and on HEF feeder layer respectively. Development of blastocysts and their behavior of praxiology were observed and the ESC isolation efficiency under feeder-free condition and on HEF feeder layer was studied simultaneously. Results：Blas- tocysts could normally develop under both feeder-free condition and on HEF feeder layer. Prima- ry ESC could be isolated by the two culture systems. When compared with the group of HEF feeder layer, attachment rate of blastocysts after 48 h culture and ESC isolation efficiency were significantly lower under feeder-free condition（P〈0.05）. Conclusion： ESC can be primarily obtained under feeder-free condition. How to improve the ESC isolation efficiency needs further studies.