山东大学耳鼻喉眼学报 ›› 2015, Vol. 29 ›› Issue (4): 65-68.doi: 10.6040/j.issn.1673-3770.0.2015.137

• 论著 • 上一篇    下一篇

人眼眶脂肪基质细胞的分离培养、鉴定及成脂诱导分化的研究

赵平千, 张瑜, 杨文蕾   

  1. 上海交通大学医学院附属仁济医院眼科, 上海 200127
  • 收稿日期:2015-03-29 修回日期:2015-06-23 出版日期:2015-08-16 发布日期:2015-08-16
  • 通讯作者: 赵平千。E-mial:pingqianzhao@yahoo.com E-mail:pingqianzhao@yahoo.com
  • 作者简介:赵平千。E-mial:pingqianzhao@yahoo.com
  • 基金资助:
    上海交通大学医学院科技基金项目资助(13XJ10047)

Isolation, culture, identification and adipogenic differentiation of orbital adipose-derived stromal cells in human.

ZHAO Pingqian, ZHANG Yu, YANG Wenlei   

  1. Department of Ophthalmology, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127, China
  • Received:2015-03-29 Revised:2015-06-23 Online:2015-08-16 Published:2015-08-16

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摘要： 目的 探讨人眼眶脂肪基质细胞(hOADSC)体外分离培养、鉴定方法及其向脂肪细胞诱导分化的能力。方法 采用组织块培养法分离人眼眶脂肪基质细胞并在体外进行原代培养及传代扩增。取第二代hOADSC细胞, 用流式细胞仪检测细胞表面抗原CD29、CD31、CD44、CD45的表达情况;采用成脂诱导培养液体外诱导hOADSC细胞向成熟脂肪细胞分化并用油红O染色。倒置相差显微镜下观察hOADSC形态、测定细胞贴壁率、细胞倍增时间、CCK-8检测细胞增殖情况并绘制细胞生长曲线。结果 组织块培养法分离得到hOADSC, 细胞呈梭形或纺锤形生长, 传代后细胞增殖迅速。细胞表面标志物CD29、CD31、CD44、CD45的表达率分别为99.0%、1.5%、99.1%、1.2%。hOADSC在体外成脂诱导培养液的作用下可分化为成熟的脂肪细胞, 细胞内的脂肪滴被油红O染为红色。传代培养hOADSC细胞贴壁率为92.6%, 细胞倍增时间为1.98 d, 细胞可在体外生长传代扩增10余代。结论 采用组织块培养法成功地分离培养了hOADSC, 其在体外成脂诱导培养液的作用下可分化为成熟的脂肪细胞。

关键词: 眼眶, 脂肪基质细胞, 细胞培养

Abstract: Objective To investigate the method of isolation, culture and identification of human orbital adipose-derived stromal cells (hOADSCs) and it's ability of adipogenic differentiation in vitro. Methods The hOADSCs were isolated by tissue explant culture technique, then were primary cultured and subcultured in vitro. The second generation of hOADSCs was detached, and the expressions of cell surface antigen CD29, CD31, CD44 and CD45 were analyzed with flow cytometry to identify the cells. The hOADSCs were induced to be adipogenesis with the adipogenic induction medium in vitro and stained with oil red O. The morphology of the hOADSC was observed by phase contrast microscopy. The plating efficiency and doubling time of the hOADSCs were tested. The hOADSCs proliferation was detected by CCK-8, and the cell growth curve was made. Results The hOADSCs were isolated by tissue explant culture technique and exhibited a fibroblast- or spindle-like morphology. The hOADSCs grew rapidly after subculturing. The expression rate of cell surface antigen CD29, CD31, CD44 and CD45 was 99.0%, 1.5%, 99.1% and 1.2%, respectively. The hOADSCs were differentiated into adipocytes with the adipogenic induction medium in vitro and the intracellular lipid droplets were stained in red by oil red O. The plating efficiency and doubling time of the subcultured hOADSC were 92.6% and 1.98days, respectively. The hOADSCs could be subcultured for more than ten generations in vitro. Conclusion In this experiment, the hOADSCs were isolated, cultured by tissue explant culture technique and could be differentiated into mature adipocytes with the adipogenic induction medium in vitro.

Key words: Orbit, Adipose-derived stromal cell, Culture

中图分类号: 

  • Q813.1
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