中国组织工程研究 ›› 2014, Vol. 18 ›› Issue (19): 3005-3011.doi: 10.3969/j.issn.2095-4344.2014.19.009

• 干细胞培养与分化 stem cell culture and differentiation • 上一篇    下一篇

诱导多能干细胞建系及体外造血祖细胞定向分化体系的建立

沈  洁1,李金辉2,阮  静3,邱  云3,朱  殷3,崔大祥3,范志宏1,汪  铮2   

  1. 上海交通大学医学院附属仁济医院,1整形外科,2消化研究所,上海市  200127;3上海交通大学微纳米科学技术研究院生物纳米科学工程研究室,上海市  200030
  • 修回日期:2014-03-20 出版日期:2014-05-07 发布日期:2014-05-07
  • 通讯作者: 范志宏,博士,主任医师,上海交通大学医学院附属仁济医院整形外科,上海市 200030
  • 作者简介:沈洁,女,1969年生,上海市人,汉族,1995年上海第二医科大学毕业,硕士,副主任医师,主要从事眼科领域工作。

Systematic establishment for directed differentiation of induced pluripotent stem cells into hematopoietic progenitor cells in vitro

Shen Jie1, Li Jin-hui2, Ruan Jing3, Qiu Yun3, Zhu Yin3, Cui Da-xiang3, Fan Zhi-hong1, Wang Zheng2   

  1. 1Department of Plastic Surgery, 2Institute of Digestion, Renji Hospital, Medical School of Shanghai Jiao Tong University, Shanghai 200127, China; 3Biological Engineering Laboratory of Nanoscience, Micro-Nano Science and Technology Institute, Shanghai Jiao Tong University, Shanghai 200030, China
  • Revised:2014-03-20 Online:2014-05-07 Published:2014-05-07
  • Contact: Fan Zhi-hong, M.D., Chief physician, Department of Plastic Surgery, Renji Hospital, Medical School of Shanghai Jiao Tong University, Shanghai 200127, China
  • About author:Shen Jie, Master, Associate chief physician, Department of Plastic Surgery, Renji Hospital, Medical School of Shanghai Jiao Tong University, Shanghai 200127, China

摘要:

背景:目前,大量文献报道了诱导多能性干细胞系的建立,但大规模体外诱导分化造血祖细胞的研究还缺乏深入的探讨。
目的:建立诱导多能性干细胞体外定向诱导形成造血祖细胞的方法。
方法:采用慢病毒感染的方法将含有Oct4、Sox2、Nanog和Lin28全能性基因的慢病毒颗粒转导人皮肤成纤维细胞,获得了诱导多能性干细胞;在诱导分化体系中添加了Y-27632,克服干细胞扩增中的凋亡现象;运用OP9细胞产生的条件培养液建立诱导多能性干细胞体外定向分化形成造血祖细胞的分化体系。
结果与结论:①前3代细胞克隆传代时,诱导多能性干细胞发生凋亡的现象很多,很难大规模扩增培养。培养基中添加阻断ROCK活化的抑制剂,能够明显抑制胚胎干细胞的凋亡。②诱导多能性干细胞在OP9细胞条件培养液作用下,经过体外诱导分化,形成CD34+造血祖细胞。


中国组织工程研究杂志出版内容重点:干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程


全文链接:

关键词: 干细胞, 培养, 诱导多能性干细胞, 类胚体, 造血祖细胞, 体外分化

Abstract:

BACKGROUND: There have been a large number of reports on establishing induced pluripotent stem cell lines, but studies concerning large-scale in vitro induced differentiation of induced pluripotent stem cells into hematopoietic progenitor cells still have a lack of in-depth discussion.
OBJECTIVE: To develop methods to induce differentiation of induced pluripotent stem cells into hematopoietic progenitor cells in vitro.
METHODS: Using the method of infection with lentivirus particles containing four transcriptionfactor genes, which are Oct4, Sox2, Nanog and Lin28, human skin fibroblasts are transduced into induced pluripotent stem cells. In the induced differentiation system, Y-27632, a kind of tyrosine kinase inhibitor-ROCK (p160-Rho-associated coiled-coil kinase), was added, which obviously suppressed apoptosis of cells. Based on conditioned medium with OP9 cells, a differentiation system of inducing induced pluripotent stem cells differentiating into hematopoietic progenitor cells was established.
RESULTS AND CONCLUSION: (1) Apoptosis of induced pluripotent stem cells at the first three passages was very obvious, and the cells were difficult in a large-scale expansion. After Y-27632 was added, the apoptosis of embryonic stem cells was obviously inhibited. (2) During embryoid body differentiation, induced pluripotent stem cells cultured in OP9 conditional growth medium differentiated into hematopoietic progenitor cells in vitro that were positive for CD34.


中国组织工程研究杂志出版内容重点:干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程


全文链接:

Key words: induced pluripotent stem cells, embryoid bodies, hematopoietic stem cells, cell differentiation

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