天津医药

天津医药 ›› 2015, Vol. 43 ›› Issue (6): 611-615.doi: 10.11958/j.issn.0253-9896.2015.06.009

• 实验研究 • 上一篇    下一篇

大鼠骨髓间充质干细胞的分离鉴定及免疫调节性能初探

张志强， 刘义， 李克秋， 李光   

  1. 天津医科大学基础医学院生物学教研室


  • 收稿日期:2014-09-30 修回日期:2014-12-19 出版日期:2015-06-15 发布日期:2015-06-10
  • 通讯作者: 张志强 E-mail:qqzzq8821@163.com
  • 基金资助:
    国家高技术研究发展计划（863计划）资助项目（2012AA021003）； 国家自然科学基金资助项目（21177091）； 天津市科技支撑计
    划资助项目 （12ZCZDSY03400）

Isolation of rat bone marrow-derived mesenchymal stem cells and explore its role in#br# immunomodulation

ZHANG Zhiqiang, LIU Yi, LI Keqiu, LI Guang   

  1. Department of Biology, Basic Medical College,Tianjin Medical University

  • Received:2014-09-30 Revised:2014-12-19 Published:2015-06-15 Online:2015-06-10
  • Contact: Zhi-Qiang ZHANG E-mail:qqzzq8821@163.com

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摘要： 目的 分离培养大鼠骨髓间充质干细胞 （BMSCs）， 鉴定其向脂肪细胞和软骨细胞的分化潜能， 并探讨其免疫调节性能。方法 无菌条件下摘取 Sprague-Dawley（SD） 大鼠股骨和胫骨， 全骨髓贴壁法培养 BMSCs， 胰酶消化传代培养。待培养至第 3 代时进行流式鉴定， 检测分离的 BMSCs 分化为脂肪细胞的潜能。与 Wistar 大鼠脾脏来源 T细胞进行共培养， 分为直接接触组和 Transwell 共培养组， 探讨其对 T 细胞亚群的影响， 并研究其相关机制。结果 BMSCs 呈梭形生长， 生长至第 3 代的细胞经检测发现几乎不表达 CD34 和 CD45， 高表达 CD29、 CD44 和 CD90， 且经诱导后细胞可分化为脂肪细胞和软骨细胞。与脾脏来源 T 细胞共培养可有效降低 CD8+的效应 T 细胞（Teffs）的比例， 并且增加 CD4+CD25+双阳性的调节性 T 细胞 （Tregs） 数量， 具有一定的免疫调节性能。与 T 细胞共培养后 BMSCs的 IL-10 和 TGF-β1 基因表达显著增强， 其中与 T 细胞直接接触组效果更为明显。结论 BMSCs 可能通过直接接触以及分泌细胞因子来发挥免疫调节功能。

关键词: 骨髓, 间质干细胞, 细胞分化, 神经免疫调节, 大鼠, Sprague-Dawley, 大鼠, Wista

Abstract: Objective To explore the immunomodulation property of bone marrow-derived mesenchymal stem cells(BMSCs) from Sprague-Dawley (SD) rats after they are isolated, cultured and identified by surface marker and differentiation potential examination. Methods BMSCs were isolated from femur and tibia of SD rats and passaged by trypsinization. The surface markers of the 3rd passage BMSCs were detected by flow cytometry and the capacity of their adipocyte and cartilage differentiation were examined. In order to explore the immunomodulation property of BMSCs, allogeneic spleen T cells of Wistar rats were co-cultured with BMSCs through either cell-to-cell contact or transwell, then its effect on the T cell subsets and related mechanism was also examined. Results BMSCs were mainly spindle-shaped in culture. Surface marker detection showed that BMSCs expressed high levels of CD29, CD44 and CD90 but no CD34 nor CD45 at the third generation. Under specific condition, BMSCs could differentiate into adipocytes and chondrocytes. The CD8+ effector T cells (Teffs) decreases effectively and the CD4+CD25+ regulatory T cells (Tregs) increased remarkably when BMSCs were co-cultured with allogeneic spleen T cells for 48 hours. The expressions of IL-10 and TGF-β1 of BMSCs significantly increased after co-culture with T cells, and this effect was more obvious in cell-to-cell contact group. Conclusion The immunomodulation property of BMSCs were presumably function through cell-to-cell contacts and cytokine secretion.

Key words: bone marrow, mesenchymal stem cells, cell differentiation, neuroimmunomodulation, rats, Sprague-Dawley, rats, Wistar