2种饲养层对牛iPSCs从始发态向初始态转化的影响  

Impacts of two types of feeder layer cells ontransformation of bovine iPSCs from primed state to naive state

作　　者：陈斓歆 姜禹 杨蕊 朱春玲 张博洋 王玥琦 张言 赵彦森 唐博 张学明 CHEN Lanxin;JIANG Yu;YANG Rui;ZHU Chunling;ZHANG Boyang;WANG Yueqi;ZHANG Yan;ZHAO Yansen;TANG Bo;ZHANG Xueming(College of Veterinary Medicine,Jilin University,Changchun 130062,China;College of Animal Science and Veterinary Medicine,Henan Institute of Science and Technology,Xinciang,Henan 453003,China)

机构地区：[1]吉林大学动物医学学院,吉林长春130062 [2]河南科技学院动物科技学院,河南新乡453003

出　　处：《中国兽医学报》2024年第2期335-340,355,共7页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(32172803,31872434);广东省清远市科技计划资助项目(2018A018)。

摘　　要：饲养层细胞对获得多能干细胞(pluripotent stem cells,PSCs)非常重要,小鼠胚胎成纤维细胞(mouse embryonic fibroblasts,MEF)已被广泛用作培养和产生多种胚胎干细胞(embryonic stem cells,ESCs)及诱导PSCs(induced PSCs,iPSCs)的饲养层,但由于种间差异,MEF可能不适于家畜ESCs和iPSCs。研究表明,PSCs可被分为至少2种多能性状态:初始态(naive state)和始发态(primed state)。本室前期用MEF饲养层从牛睾丸支持细胞(sertoli cells,SCs)获得了牛的始发态iPSCs。本研究基于初始态培养液(na6ve medium,NM),比较了MEF饲养层和牛胚胎成纤维细胞(bovine embryonic fibroblasts,BEF)饲养层对牛iPSCs多能性和形态维持及其从始发态向初始态样转化的影响。结果显示,MEF+NM(MNM)和BEF+NM(BNM)均能在3~4 d内将牛iPSCs从始发态转化为初始样状态。克隆分析显示,BNM培养中牛iPSCs的克隆数、碱性磷酸酶活性和转化效率均高于MNM培养中的指标。这些克隆在长期培养中均能表达初始态iPSCs的多能性分子标记。结果表明,采用NM条件,在MEF和BEF饲养层上均可产生牛初始态样iPSCs,但在BEF上转化效率较高。Feeder cells are essential to derive pluripotent stem cells(PSCs).Mouse embryonic fibroblasts(MEF)are widely used as feeder to generate and culture embryonic stem cells(ESCs)and induced PSCs(iPSCs)in many species.However,it may not be suitable for livestock ESCs/iPSCs due to interspecies difference.Studies indicate that PSCs can be classified into at least two pluripotent states,the naive and the primed states.Previously bovine primed iPSCs were derived from bovine Sertoli cells using MEF feeder in our laboratory.Here we compared the effects of MEF feeder and bovine embryonic fibroblasts(BEF)feeder on the maintenance of bovine iPSCs pluripotency and morphology as well their contributions to the na6ve-like conversion,based on a na6ve medium(NM).The results showed successful conversion of the primed iPSCs to the naive-like iPSCs within 3-4 d both on MEF feeder and BEF feeder in NM(termed as MNM and BNM respectively).The colony analysis demonstrated that the colony number,colony alkaline phosphatase,and the conversion efficiency were all higher under BNM condition compared with MNM condition.During the long-term cultivation,these bovine iPSCs colonies retained the pluripotent markers of naive iPSCs.Together,bovine naive-like iPSCs can be generated both on MEF and BEF feeder in NM condition.The BNM condition is better to improve the conversion efficiency.

关 键 词：牛 牛胚胎成纤维细胞 诱导多能干细胞 小鼠胚胎成纤维细胞 初始态 始发态 

分 类 号：S814.8[农业科学—畜牧学]