绵羊羊膜上皮细胞分离培养及鉴定

Isolation,culture and identitication of Ovis aries amnion epithelial cells

背景:目前有研究发现人羊膜和大鼠羊膜分离消化培养后,可成功获得具有干细胞特性的人羊膜上皮细胞和鼠羊膜上皮细胞,在体外一些外源因子作用下具有向三胚层细胞分化的潜能。目的:获取绵羊羊膜上皮细胞,并对其干细胞特性进行鉴定。方法:采用机械方法剥离绵羊羊膜组织,运用低速旋转TrypLE(胰酶替代物)消化法对绵羊羊膜进行分离培养获得绵羊羊膜上皮细胞。结果与结论:细胞免疫荧光检测结果表明绵羊羊膜上皮细胞表达Oct-4、SSEA-1、SSEA-3、SSEA-4、TRA-1-60、TRA-1-81等胚胎干细胞标记蛋白;RT-PCR技术鉴定结果显示绵羊羊膜上皮细胞的Oct-4、Sox-2、Rex-1全能性相关基因表达,而Nanog基因不表达。检测结果提示实验成功获得具有干细胞特性的绵羊羊膜上皮细胞。

BACKGROUND：The amnion epithelial cells（AECs） with stem cells properties have been successfully obtained after isolating,digesting and culturing human and rat amnion and AECs have the potential to differentiate into cells of the endoderm（liver,lung,epithelium）,mesoderm（bone,fat）,and ectoderm（neural cells）,OBJECTIVE：To obtain the Ovis aries AECs and detect the stem cell characteristics.METHODS：Ovis aries amnion tissue was peeled off by mechanical way and the low velocity-trypLE（pancreatic enzyme replacement） digestion method was used to isolate and culture the Ovis aries amnion,and the Ovis aries AECs were obtained.RESULTS AND CONCLUSION：The immunofluorescence observation showed that the labeled protein of AECs such as Oct-4,SSEA-1,SSEA-3,SSEA-4,TRA-1-60,TRA-1-81 could be expressed on Ovis aries AECs.RT-PCR experiments confirmed that the genes of Oct-4,Sox-2 and Rex-1 were expressed and Nanog was not expressed.The results indicate that the Ovis aries AECs with stem cells properties have been achieved successfully.