一种简单经济的小鼠骨髓源性肥大细胞的培养与鉴定  

A simple and economical method for murine bone marrow mast cell cultivation and identification

作　　者：王春锋[1] 李济宇[1] 

机构地区：[1]上海交通大学医学院附属新华医院普外科,上海200092

出　　处：《中国免疫学杂志》2011年第10期910-912,921,共4页Chinese Journal of Immunology

基　　金：国家自然科学基金资助项目(30600598);上海市科学技术委员会资助项目(09411952500);上海市科委青年科技启明星后资助项目计划(10QH1401800);上海市教育委员会曙光计划(10SG20)

摘　　要：目的:通过一种简单经济的方法在体外获得大量高纯度的肥大细胞。方法:将小鼠骨髓细胞直接冲出加入到含有IL-3和SCF的培养基中培养,每周换一次液,四到六周后收集培养细胞,通过光镜和电镜下观察肥大细胞的表面和内膜结构特征,流式细胞术检测表面CD117和FITC-FCεRⅠα的表达情况,利用甲苯胺蓝检测其功能。结果:四周后收集培养的细胞,其大小均一,内部结构符合肥大细胞的特征,电镜下可见肥大细胞内部含有丰富的高电子密度的颗粒,其中有些可见脱颗粒的空泡。流式检测显示CD117和FITC-FCεRⅠα单阳性的表达率都在90%以上,双阳性的表达率也在80%以上。甲苯胺蓝染色发现收集的肥大细胞核被染成深蓝色,胞质染成淡紫色。通过这种方式我们获得了大量的肥大细胞,其成熟时间较短,所需培养基和相关细胞因子量较少,而且寿命较长,纯度较高。结论:在IL-3和SCF的刺激下可以诱导骨髓干细胞向肥大细胞定向分化,通过这种方式可以快速经济的获得大量高纯度的肥大细胞,用于体外更好的研究其在移植免疫领域的作用。Objective：To obtain lots of high pure mast cells simply and economically in vitro.Methods：The bone marrow cells isolated from BALB/c mice were flushed into the culture medium contained IL-3 and stem cell factor.The culture medium were changed once a week.The cultured cells were harvested and observed their surface and internal structure under light and transmission electron microscope after four to six weeks.The expression of surface marker-CD117 and FCεRⅠα were detected by flow cytometry.The cultured cell function was analyzed by toluidine blue staining.Results：The four-week cultured cells were in uniform size and their internal structures were in accordance with the characteristic of mast cells.The cultured mast cells were rich in high electron-density particles,some of which degranulation vacuoles might be seen under transmission electron microscope.The single-positive CD117 and FCεRⅠα expression were more than 90%,and double-positive cells were more than 80% detected by flow cytometry.The nucleus of harvested cells was stained dark blue and their granules were stained purple by toluidine blue.We can obtain more longevity and high purity mast cells through this pathway in a comparably short time economically.Conclusion：The bone marrow stem cells can be induced into mast cells under IL-3 and stem cell factor stimulation.We can obtain high purity mast cells simply in order to explore their potential role in organ transplantation in vitro.

关 键 词：骨髓 肥大细胞 培养 移植 

分 类 号：Q291[生物学—细胞生物学]