小鼠精原干细胞体外分化中Stra8、mina53表达的研究

A Primary Study of Gene Stra8 and Gene Mina53 in the Differentiation of mSSCs in Vitro

为检测Stra8、mina53在小鼠精原干细胞(mouse spermatogonial stem cells,mSSCs)体外分化中的表达,采用1)体外培养mSSCs并通过全反式视黄酸(ATRA,all-trans retinoic acid)进行诱导分化;2)通过间接免疫荧光反应鉴定mSSCs体外分化前后情况;3)通过RT-PCR检测mSSCs诱导分化前后(0h,2h,72h)Stra8、mina53mRNA的水平,初步了解Stra8、mina53的表达状况。结果表明:1)形态学变化显示ATRA使mSSCs向精母细胞方向发生了分化,并通过间接免疫荧光反应进行鉴定。2)Stra8在mSSCs诱导分化2h、72h的水平高于成年小鼠和未诱导分化的水平,其中72h时水平低于2h,各对照组间差异具有统计学意义。3)mina53的表达在诱导后增加,其水平高于成年小鼠和未诱导mSSCs的水平,但差异无统计学意义。这表明ATRA在体外使mSSCs向精母细胞方向分化,Stra8基因呈高表达,但mina53基因呈高表达不确定。

To study the expression of Stra8 and mina53 in orientation-induced differentiation in spermatogonial stem cells in vitro.Methods： 1）Separated and cultured spermatogonial stem cells in vitro.Induced mSSCs differentiate into spermatocyte orientation in vitro by ATRA.2）Markers（c-kit、Oct-4、GCNF） were detected by indirect immunofluorescene in mSSCs before and after induction.3）Spermatogonial stem cells were induced by ATRA for 2h and 72h.Meanwhile the levels of Stra8 and mina53 mRNA expression were detected by RT-PCR method.The results are as follows： 1）The morphology change of mSSCs indirect immunoinfluorescence showed cultured mSSCs had been differetioned by the induction of ATRA.2）The levels of stra8 mRNA of induced mSSCs for 2 h and 72 h were higher than those of adult mouse testis and uninduced mSSCs,however,the level that mSSCs had been induced for 72 h was below to which induced for 2 h.The differences between all groups were statistical（P0.05）.3）The expression of mina53 had been increased after mSSCs were induced.Surprisingly,the differnces between all groups weren,t statistical（P0.05） .The conclusion is that：ATRA might have the capability that induces mSSCs differentiate into spermatocyte orientation in vitro.The levels of stra8 mRNA of induced mSSCs is upgrated,but it is yet uncertain that the gene mina53 be high in vitro while those induced by ATRA.