小鼠精原细胞体外培养分化的研究

Study on in vitro Culture and Differentiation of Mouse Spermatogonia

目的探讨小鼠精原细胞的分离纯化及体外培养条件。方法采用组合酶消化法分离纯化小鼠精原细胞和支持细胞,在添加FSHC和TC的基本培养基中进行体外培养。采用反转录PCR方法检测精子细胞特异性基因过渡蛋白9(Tnp2)的表达以对分化细胞进行鉴定。结果培养5 d后有分化的圆形精子细胞出现,培养7d后收集的细胞可见Tnp2基因表达。结论通过体外培养小鼠精原细胞可迅速完成减数分裂过程和减数分裂后的分化过程,并可获得更接近成熟阶段的精子细胞。

Objective To study on the isolation and purification and in vitro culture of mouse spermatogonia. Methods The combined enzymatic digestion was used to separate and purify the germ cells from the testes of neonatal mouse, which were then in vitro cultured in DMEM/F12 medium with rFSH and testosterone. Reverse transcription-PCR （RT-PCR） was used to investigate the expression of transition protein 2 （Tnp2） , a spermatid specific gene,to identify the culture result. Results The round spermatids were generated after 5-day culture,and Tnp2 expression was detected in the collected cells after 7-day culture. Conclusion The mouse spermatogonia could perform meiosis and differentiation to generate spermatids after in vitro culture.