红细胞裂解法及不同培养条件对兔骨髓间充质干细胞体外增殖的影响  

Red Blood Cell Lysis Method and Effects of Different Culture Conditions on Proliferation in Vitro of Rabbit Bone Marrow Mesenchymal Stem Cells

作　　者：吕婧玉[1] 和小娥[1] 张志平[1] 许晓婷[1] 冯建昆[1] 王新庄[1] 

机构地区：[1]河南农业大学牧医工程学院,河南郑州450002

出　　处：《江西农业学报》2012年第11期132-135,共4页Acta Agriculturae Jiangxi

基　　金：国家自然科学基金(31001096);河南省基础与前沿技术研究计划(092300410081)

摘　　要：研究采用红细胞裂解液法从兔骨髓中分离BMSCs,相差显微镜观察其形态,四甲基偶氮唑盐法(MTT法)筛选基础培养液和血清浓度,绘制生长曲线,用流式细胞仪检测细胞周期,免疫细胞化学鉴定表面抗原。分离的细胞在形态学观察与生长动力学上均符合BMSCs特征;在实验范围内DMEM/F12+20%FBS是体外培养兔BMSCs的最佳体系;细胞周期结果显示,BMSCs平均82%处于G0/G1期,18%处于S+G2期;免疫细胞化学结果显示所分离的BMSCsCD90、CD44阳性表达,CD34阴性表达。本研究结果表明,通过红细胞裂解液法分离的兔BMSCs,其具有体外增殖和多向分化的潜能,可以作为组织工程的种子细胞。The red blood cell lysis method was used to isolate the bone marrow mesenchymal stem cells （BMSCs） of rabbits here. We also observed its pattern with inverted phase contrast microscope. Basic medium and serum concentration for BMSCs culture were optimized with MTr [ 3 - （4,5） - dimethyhhiahiazo （ - z - yl） - 3,5 - di - phenytetrazoliumromide ] reduction assay. The growth curves for BMSCs were made, and its cell cycles were detected by flow cytometry. Then CD34, CD44 and CD90 antigens of BMSCs were identified by immunocytochemistry. Results showed that the isolated and cultured cells corresponded with BMSCs in morphology and growth kinetics. DMEM/F12 ＋20% FBS was the best system for rabbit BMSCs in vitro based on our results. The cell cycle analy- sis showed that 82% cells were in GO/G1 phase, and 18% cells were in S ＋ G2 phase. The immunocytochemistry result showed that the surface antigen CDgO and CD44 were positive, while CD34 was negative. The results also indicated that BMSCs isolated by the red blood cell lysis method in vitro had better potentiality of proliferation and differentiation. They could be used as the seed cells of tissue engineering.

关 键 词：骨髓间充质干细胞 兔 分离培养 红细胞裂解法 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]