摘要
分离19期鸡胚生殖嵴细胞,采用原始生殖细胞(primordial germ cells,PGCs)-体细胞共培养获得PGCs集落,借助PGCs特异性标记基因Dazl和CvhmRNA表达检测、过碘酸雪夫氏(periodic acid-Schiff,PAS)染色、阶段特异性胚胎抗原(stage-specific embryonic antigen,SSEA-1、SSEA-3、SSEA-4)免疫组化检测均表明所形成的细胞团为PGCs。RT-PCR检测及基因测序结果表明干细胞多能性相关基因PouV(Oct4同源基因)、Nanog和Sox2在鸡PGCs中均有显著表达。在去除饲养层、添加15%血清并采用悬浮培养的条件下,鸡PGCs在体外可分化形成类胚体(embryoid bodies,EBs)。在PGCs体外分化形成EBs第0d、3d、5d和10d分别检测Dazl、Cvh、PouV、Nanog和Sox2mRNA的表达。结果表明:PouV、Nanog和Sox2在体外培养的鸡PGCs中显著表达,并且在PGCs体外分化过程中,其表达量伴随PGCs标记基因Dazl和Cvh一起呈明显下调趋势,提示PouV、Nanog和Sox2在维持鸡PGCs多能性中起着重要的作用。
Primordial germ cells （PGCs） were isolated from the genital ridges of chicken embryos at the 19th stage and co-cultured with somatic cells to obtain multicellular PGCs colonies. These cells were identified by stem cell-specific markers SSEA-1, SSEA-3 and SSEA-4, expression of PGCs-specific marker genes such as Dazl and Cvh together with the pluripotency-associated genes such as chicken Oct4 homologue PouV （cPouV）, chicken Nanog （cNanog） and Sox2 genes. The expression profiles of Dazl, Cvh, cPouV, cNanog and Sox2 genes during the EBs development in vitro were also examined. Results showed that all of these genes were expressed in chicken PGCs, and were down-regulated during the formation of EBs, suggesting their presumable effects in their regulatory effects in maintaining the pluripotency during differentiation of chicken PGCs. Further research is required to study the mechanisms of these genes in regulating the pluripotency and self-renewal of chicken PGCs.
出处
《细胞生物学杂志》
CSCD
2009年第1期96-100,共5页
Chinese Journal of Cell Biology
基金
教育部新世纪人才支持计划(No.NCET-05-0514)
浙江省科技计划项目(No.2008C22040)资助~~
