猪滋养层干细胞样细胞的分离培养

›› 2013, Vol. ›› Issue (5): 156-160.

猪滋养层干细胞样细胞的分离培养

宋志强^1,2, 刘畅², 冯冲², 龙川², 赵勤丽², 刘忠华¹, 潘登科²

1. 东北农业大学动物医学学院, 黑龙江哈尔滨 150030;
2. 中国农业科学院北京畜牧兽医研究所, 北京 100193

收稿日期:2012-08-15 出版日期:2013-05-20 发布日期:2013-05-27
通讯作者: 潘登科(1972-),男,甘肃人,硕士生导师,副研究员,从事异种器官移植、基因修饰猪模型建立和猪干细胞研究。Tel:010-62818180;E-mail:pandengke2002@163.com;刘忠华(1972-),男,哈尔滨人,博士生导师,教授,从事基因修饰猪模型建立、猪早期胚胎发育的机制和猪干细胞研究。Tel:0451-55191729;E-mail:liu086@126.com E-mail:pandengke2002@163.com;liu086@126.com
作者简介:宋志强(1984-),男,黑龙江人,硕士生,研究方向:猪滋养层干细胞和猪胚胎干细胞。
基金资助:
国家自然科学基金面上项目资助(30972095);973"器官缺陷和异种移植医用猪的创建"(2011CBA01004)。

Isolation and Cultivation of Porcine Trophoblastic Stem-like Cell Lines

SONG Zhi-qiang^1,2, LIU Chang², FENG Chong², LONG Chuan², ZHAO Qin-li², LIU Zhong-hua¹, PAN Deng-ke²

1. College of Veterinary Medicine, Northeast Agricultural University, Harbin 150030, China;
2. Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China

Received:2012-08-15 Online:2013-05-20 Published:2013-05-27

摘要/Abstract

摘要： 滋养层干细胞(TSc)是形成胎盘组织细胞的前体细胞,与胚胎着床和胎盘形成密切相关。体外分离滋养层干细胞为研究滋养层的发育与功能提供了研究工具和基础。滋养层干细胞已经成功从小鼠附植前囊胚中获得,在猪上也有相关报道,但并没有关于6 d囊胚中分离到猪滋养层干细胞(pTSc)的报道。本试验通过对6 d孤雌囊胚透明带进行划口处理, 饲养层和基础培养液中附加碱性成纤维生长因子(bFGF)和人白血病抑制因子(hLIF)分离猪滋养层干细胞样细胞。同时,通过采用形态学和基因表达分析的方法对获得的猪滋养层干细胞样细胞进行了初步鉴定。结果显示,本试验分离得到的细胞呈现上皮样细胞形态,细胞间结合紧密,边缘光滑,核质比较大,RT-PCR结果显示表达滋养层干细胞标记基因Cdx2,体现了滋养层干细胞特征。结果表明,本试验成功在6 d孤雌囊胚分离得到猪滋养层干细胞样细胞,为后续研究猪滋养层发育提供了试验基础。

关键词: 猪; 滋养层干细胞; 体外囊胚; 分离培养

Abstract: Trophoblast stem cells (TSc) were the precursors of the differentiated cells of the placenta. It was associated with embryo implantation. Isolating trophoblast stem cells in vitro provided research foundation for trophoblastic development. TSc had been successfully derivated from preimplantation embryo, there were also many reasearches about porcine trophoblast stem cells(pTSc), but no reports indicated derivating 6 d pTSc. After broken the zona pellucida,we fixed the blastocyst on feeder under sterile condition. The porcine trophoblatic stem-like cell was intially analysed by morphogenesis and RT-PCR. We found the porcine trophoblastic stem-like cell combined tightly and present epithelioid-like cell. The clone was smooth,the ratio of the nucleus and cytoplasm was mass. The porcine trophoblastic-like cell was confirmed expressing Cdx2 gene by RT-PCR. We successfully derived the porcine trophoblastic stem-like cell from 6 d pathenogonidium blastcyst. The result provided some foundation for follow-up studies of porcine trophoblast development.

Key words: porcine; throphoblast stem cell; in vitro blastocyst; isolation and culture

中图分类号:

S813.1⁺1

宋志强, 刘畅, 冯冲, 龙川, 赵勤丽, 刘忠华, 潘登科. 猪滋养层干细胞样细胞的分离培养[J]. , 2013, (5): 156-160.

SONG Zhi-qiang, LIU Chang, FENG Chong, LONG Chuan, ZHAO Qin-li, LIU Zhong-hua, PAN Deng-ke. Isolation and Cultivation of Porcine Trophoblastic Stem-like Cell Lines[J]. China Animal Husbandry & Veterinary Medicine, 2013, (5): 156-160.

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