不同细胞因子组合对c-kit^+Lin^-细胞的体外扩增研究  

In vitro expansion of c-kit^+Lin^- cells of bone marrow in presence of several cytokines

作　　者：张守华[1] 廖彩仙[1] 张春兴[1] 苏俊[1] 赖勇强[1] 周杰[1] 

机构地区：[1]南方医科大学南方医院肝胆外科,广东广州510515

出　　处：《中国现代医学杂志》2008年第12期1652-1654,1659,共4页China Journal of Modern Medicine

基　　金：国家自然科学基金(30371385);广东省自然科学基金(04020419)

摘　　要：目的探讨在无血清无基质条件下,不同细胞因子组合对c-kit+Lin-细胞增殖效应以及最佳扩增时间。方法采用免疫磁珠法分离小鼠骨髓c-kit+Lin-细胞,将干细胞因子(stem cell factor,SCF),FLt-3配基(FLt-3 ligand,FL),血小板生成素(thrombopoietin,TPO),白细胞介素-3(interleukin,IL-3),肝细胞生长因子(hepatocyte graoth factor,HGF)等细胞因子进行不同组合,体外扩增干细胞14d,于不同时间检测细胞总数和c-kit+Lin-细胞数及比例,并在第10天通过细胞凋亡早期膜变化标记FIFT-Annexin-V,检测细胞凋亡率。结果各组细胞因子均能显著扩增c-kit+Lin-细胞,扩增倍数3～19倍不等。在SCF+FL+TPO中加入IL-3或HGF,其效果更明显,c-kit+Lin-细胞在第6天分别扩增4.71和5.15倍,当同时加入时,两者有协同作用。其中SCF+FL+TPO+IL-3+HGF在第10天扩增程度最为显著,并且凋亡率最低为6.59%。结论在体外短期培养过程中上SCF+FL+TPO+IL-3+HGF组合能产生大量的c-kit+Lin-细胞,并且最佳培养时期为第10天,为肝干细胞提供种子细胞。[Objective] To explore the regulation and the best time of c-kit^＋Lin^-cell proliferation from mouse bone marrow by various combinations of cytokines without serum and matrix. [Methods]c-kit^＋Lin^-cells were isolated from mouse bone marrow by using a high-gradient magnetic cell sorting system （MACS） and expanded for 14 days under different combinations of stem cell factor（SCF）, FLt-3 ligand （FL）, thrombopoietin （TPO）, hepatic growth factor （HGF）. At the different time points total cells and c-kit^＋Lin^-cells were counted. FITC-Annexin-V was applied to detect apoptosis rate on the tenth day in each group. [Results] Cytokines in each group could expand c-kit^＋Lin^-cells 3-19 times as many as before with the combination of SCF, FL TPO, IL-3 and HGF. While IL-3 or HGF were added to the combination of SCF, FL and TPO, c-kit^＋Lin^-cells were increased significantly, and the apoptosis rate decreased. When the two cytokines were added together, they showed synergistic actions with SCF, FL and TPO to expand c-kit^＋Lin^-cells. SCF＋FL＋TPO＋IL-3＋HGF expanded c-kit^＋Lin^-cells most obviously on the tenth day and Annexin-V positive cells were only 6.59%. [Conclusion] SCF＋FL ＋TPO＋HGF＋IL-3 show a stronger potential for expanding c-kit^＋Lin^-cells in vitro without serum and matrix. And the best time to expand the cells is ten days.

关 键 词：无血清培养 骨髓c-kit^+Lin^-细胞 肝干细胞 扩增 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]