猪羊水干细胞的分离培养及向脂肪细胞诱导分化  

Porcine Amniotic Fluid Stem Cells Isolation and Induction into Adipocytes

作　　者：卢智娟[1] 成德[1] 李军华[1] 张淑金[1] 陈家欢[1] 雷蕾[1] 陈帅[1] 高志敏[1] 王华岩[1] 

机构地区：[1]西北农林科技大学动物医学院,陕西省干细胞工程技术研究中心,陕西省农业分子生物学重点实验室,杨凌712100

出　　处：《农业生物技术学报》2010年第6期1054-1061,共8页Journal of Agricultural Biotechnology

基　　金：国家高技术研究发展计划(863)(No2008AA101005);国家重点基础研究发展规划(973)(No2009CB941002);转基因新技术新方法(No20082X0810-001)共同资助

摘　　要：羊水干细胞是一种存在于胎儿羊水中Oct4阳性的多能性干细胞。该细胞具有增殖能力强、分化潜能高和免疫原性低等特点,在生物学和医学研究上具有广泛的应用前景。本实验从不同胎龄的猪胎儿标本中分离猪羊水干细胞,并通过诱导其向脂肪细胞分化来检测其多向分化潜能。结果表明,从胎龄为30～70d的猪胎儿羊水中分离出的羊水干细胞贴壁率高,增殖能力强,活性好,并建立了两株猪羊水干细胞系。经流式细胞仪和RT-PCR检测,猪羊水干细胞表达CD117、CD44、CD166和CD90表面抗原,不表达CD45、CD54和CD34;稳定表达Oct4、Nanog、C-myc和HLA-abc干细胞特异标志基因;转染报告基因载体pOct4-EGFP和pNanog-EGFP后表达绿色荧光;此外,猪羊水干细胞还能够在体外诱导分化为脂肪细胞。研究结果为大动物羊水干细胞的研究提供了可借鉴的材料。Amniotic fluid stem （AFS） cells are the Oct4 positive multipotent stem cells derived from amniotic fluid. These cells possess the robust proliferation, multi-linear differentiation and lowness of immunity and have great promise in different fields of biology and clinical application. In this study, we isolated and characterized the porcine AFS （pAFS） cells devived from different gestational age of porcine fetus, and revealed the correlation between the gestational age of fetus and the proliferation and adhesion rate of pAFS cells. Further more, pAFS cells were able to differentiate into adipocytes when cultured in the special inducing medium. The results showed that pAFS cells derived from the fetus in 40～70 d range obtained the optimal proliferation and adhersion rate. Two porcine AFS cells lines were establishment so far. By FASC and RT-PCR analysis, pAFS cells expressed the following cell surface antigens including CD117, CD44, CD166 and CD90, but negatived for CD34, CD45 and CD54; These cells could be expressed constantly the stem cell special gene markers such as Oct4, Nanog, c-Myc and HLA-abc; The EGPF drived by promoters of Oct4 and Nanog were expressed in pAFS cells when transfected with pOct4-EGFP and pNanog-EGFP; Besides, pAFS cells were successful induced into adipocytes in vitro. This paper presents a method which is useful for the study of AFS cell from domestic animals.

关 键 词：猪 羊水干细胞 诱导 OCT4 脂肪细胞 

分 类 号：S852.2[农业科学—基础兽医学]