外源性shh蛋白对c-kit^+造血祖细胞增殖分化作用研究  

Exogenous shh protein regulates the proliferation and differentiation of murine HPCS

作　　者：胡采红[1] 周琨[1] 孙汉英[1] 刘文励[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院血液内科,湖北武汉430030

出　　处：《中国现代医学杂志》2010年第14期2091-2095,共5页China Journal of Modern Medicine

基　　金：国家自然科学基金资助项目(No:30570773);教育部博士点基金资助项目(No:20060487061;No:200804870008)

摘　　要：目的探讨sonic hedgehog(Shh)信号对小鼠造血祖细胞的扩增分化调控作用。方法取孕14.5天Balb/C小鼠,分离培养小鼠胎肝基质细胞作饲养层。免疫磁珠法分选小鼠骨髓来源c-kit+造血祖细胞,将分选的造血祖细胞在不同实验组培养条件下培养7d(a.对照组;b.Shh组;c.以胎肝基质细胞为饲养层的Transwell共培养组;d.Shh+Transwell共培养组)。通过绘制细胞生长曲线,免疫表型鉴定及功能分析,观察胎肝基质细胞饲养层以及Shh活性肽对细胞增殖、分化、集落形成的调控作用。结果和对照组相比,胎肝基质细胞可以明显促进造血祖细胞增殖(P<0.05);单纯的外源性Shh蛋白可以同时促进造血祖细胞的增殖和分化(P<0.05);在与胎肝基质细胞共培养体系中,外源性Shh蛋白可以明显地促进细胞增殖,经过体外培养7d后,细胞数达80×105,其中c-kit+细胞占(84±9.44)%,集落形成能力亦明显增强。结论 Shh蛋白参与调控造血祖细胞的增殖与分化,在胎肝基质细胞的支持下,Shh蛋白对小鼠c-kit+造血祖细胞的扩增有明显的促进作用,表明Shh蛋白与胎肝基质细胞存在着某种整合作用,这种作用在一定程度上调控着造血祖细胞的增殖与分化。【Objectives】To explore the supportive and expansive effects of exogenous shh protein on the c-kit＋ murine hematopoietic progenitor cells（HPCs） in vitro.【Methods】Murine fetal liver stromal cells were separated from E14.5D Balb/C mice and cultured as the feeder layer.Murine c-kit＋ HPCs were separated by MACS system from bone marrows.Cultured c-kit＋ cells were divided into 4 different groups as follows：a,control group b,Shh protein treatment c,transwell co-cultured,combination of Shh treatment and transwell co-culture,and cultured in vitro for 7 days.The number of the expanded cells were counted every day.The expression of c-kit phenotype was detected by flow cytometry.The progenitor function was assessed by CFU assays.【Results】Cultured c-kit＋ progenitor cells were at a different expansion rate in different groups.FL derived stromal cells promoted the expansion of HPCs,comparatively,HPCs in the control group had a most poor expansion.Without co-culture with FL derived stromal cells,Shh protein not only promoted expansion but also differentiation of the murine c-kit＋ HPCs.In co-culture system with FL derived stromal cells,treatment with shh protein greatly induced the proliferation of the c-kit＋ HPCs.【Conclusion】These data suggest that there exist some integration mechanism between shh protein and fetal liver,and this connection may contribute to the HPCs being refrain from differentiation.

关 键 词：造血微环境 基质细胞 Shh通路 胎鼠 胎肝 造血祖细胞(c-kit+) 

分 类 号：R-332[医药卫生]