C57BL/6小鼠骨髓间充质干细胞的体外分离培养、分化及两种不同品牌血清对其增殖的影响  

Isolation, culture and differentiation of bone marrow mesenchymal stem cells from C57BL/6 mice in vitro and effects of two different brands of serum on their proliferation

作　　者：刘剑锋[1] 汤剑明[1] 阳莲 程建红 李露 洪莉[1] Liu Jianfeng;Tang Jianming;Yang Lian;Cheng Jianhong;Li Lu;Hong Li(Department of Obstetrics and Gynecology,Peoples Hospital of Wuhan University,Wuhan Hubei 430060,P.R.China)

机构地区：[1]武汉大学人民医院妇产科,湖北武汉430060

出　　处：《中国计划生育和妇产科》2022年第12期96-100,共5页Chinese Journal of Family Planning & Gynecotokology

基　　金：国家自然科学基金资助项目(项目编号:81971364);国家自然科学基金资助项目(项目编号:82001527)。

摘　　要：目的 研究C57BL/6小鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells, BMSCs)体外分离培养、分化及比较含两种不同品牌血清培养基对其生长的影响,探索体外培养C57BL/6小鼠BMSCs更优方法。方法 差速贴壁法体外分离提取BMSCs,分别用含10%Yeasen胎牛血清和10%Gbico胎牛血清的DMEM/F12培养基进行培养,分别在2 d、4 d、6 d(后)通过倒置显微镜观察两组BMSCs的细胞形态;CCK-8法检测其生长趋势;免疫荧光染色鉴定表型;不同诱导培养基诱导其成骨、成脂分化。结果 成功分离BMSCs,原代培养细胞呈长角形涡状排列生长,第二代BMSCs免疫表型鉴定结果显示CD44、CD90、CD106为阳性表达,CD34为阴性表达;成骨、成脂诱导分化后,细胞茜素红、油红染色均呈阳性;CCK-8检测显示含Gbico血清培养基所培养的BMSCs更容易形成细胞集落且数量更多,生长上升趋势更为明显。结论 两种血清都能促进BMSCs的体外增殖,Yeasen血清有着更高的性价比,Gbico血清则在促进增殖方面表现更加良好。Objective To study the isolation, culture and differentiation of C57BL/6 mouse bone marrow mesenchymal stem cells(BMSCs) in vitro and to compare the effects of two different serum media on their growth, and to explore a better method for in vitro culture of C57BL/6 mouse BMSCs.Methods BMSCs were isolated and extracted by differential adhesion method and cultured in DMEM/F12 medium containing 10% Yeasen fetal bovine serum and 10% Gbico fetal bovine serum respectively.The cell morphology of two groups of BMSCs was observed by inverted microscope after 2 days, 4 days and 6 days, the growth trend was detected by CCK-8 method, and the phenotype was identified by immunofluorescence staining.Osteogenic and adipogenic differentiation of BMSCs were induced by different induction media.Results BMSCs was isolated successfully, and the primary cultured cells grew in an angular vortex arrangement, and the immunophenotypic identification of the second generation BMSCs showed that CD44,CD90,CD106 was positive and CD34 was negative.After osteogenic and adipogenic induction and differentiation, the cells were positive for alizarin red and oil red staining.While CCK-8 detection showed that BMSCs cultured in the medium containing Gbico serum was easier to form cell colonies and more cells, and the growth trend was more obvious.Conclusion Both serums can promote the proliferation of BMSCs in vitro, Yeasen serum has a higher cost performance, and Gbico serum performs better in promoting proliferation.

关 键 词：血清 骨髓间充质干细胞 原代培养 CCK-8 成骨分化 成脂分化 

分 类 号：R332[医药卫生—人体生理学]