无血清和含胎牛血清培养基培养的脐带间充质干细胞生物学特性比较研究  

Comparison of the biological characteristics of serum-free and fetal bovine serum-contained medium cultured umbilical cord-derived mesenchymal stem cells

作　　者：杨婷[2] 陈广华[1] 薛胜利[1] 乔曼[1] 刘慧文[1] 田竑[1] 乔淑敏[1] 陈峰[1] 陈志哲[2] 孙爱宁[1] 吴德沛[1] 

机构地区：[1]苏州大学附属第一医院、江苏省血液研究所,卫生部血栓与止血重点实验室 [2]福建医科大学附属协和医院血液科,福建省血液病学重点实验室

出　　处：《中华血液学杂志》2012年第9期715-719,共5页Chinese Journal of Hematology

基　　金：江苏高校优势学科建设工程（2010）;福建省科厅重点项目（2009Y0022）;福建省科技厅重点项目（2009J1004）;新药研究开发关键技术研究项目（2009ZX09503）

摘　　要：目的比较无血清及含胎牛血清培养基培养的人脐带间充质干细胞（UC-MSC）生物学特性，寻求无血清的UC—MSC培养方法。方法采用胶原酶消化脐带小块。分别采用MesenCult—XF无血清培养基和含胎牛血清的αMEM完全培养基培养UC—MSC。比较两种方法培养的早期UC-MSC细胞形态、免疫表型、增殖分化潜能、核型及对T淋巴细胞增殖抑制作用的差异。结果用无血清培养基培养制备的悬浮UC—MSC平均细胞直径为26（18—39）μm，含血清完全培养基制备的悬浮UC—MSC平均细胞直径为35（20—61）μm。早期UC—MSC采用无血清培养基培养时连续传代可倍增（5．2±0．2）倍，而采用含胎牛血清的完全培养基培养时传代可倍增（3．5±0．1）倍，UC—MSC采用无血清培养体系传代倍增数明显高于含胎牛血清的培养体系的传代倍增数（P〈0．05）。UC—MSC表达CD29、CD44、CD90、CD73、CD105、HLA—ABC抗原，不表达CIM5及CD34等造血细胞抗原。无血清培养的UC—MSC按MSC：T细胞比例为1：100、1：10及1：5共培养的放射性核素每分钟闪烁计数（CPM）值分别为（4．57±0．14）×10^4、（2．04±0．16）×10^4和（0．42±0．04）×10^4，含胎牛血清制备的UC—MSC共培养对应的CPM值分别为（5．29±0．18）×10^4、（2．55±0．15）×10^4和（0．52±0．03）×10^4，无血清培养的UC—MSC抑制T细胞增殖作用较含血清培养的UC—MSC抑制作用更明显（P〈0．05）。结论与含胎牛血清的完全培养基相比无血清培养的人UC—MSC细胞直径小、早期传代增殖潜能增加、无异种蛋白。无血清培养基制备的UC—MSC抑制T细胞增殖活性高于含胎牛血清培养基制备的UC—MSC。Objective To compare the differences of biological characteristics between human um- bilical cord-derived mesenchymal stem cells （UC-MSCs） cultured by serum-free medium or fetal bovine ser- um-contained complete medium to establish a xenogeneic protein-free UC-MSCs culture system. Methods Healthy human umbilical cord segments were digested with collagenase. UC-MSCs were cultured by serum- free MesenCuh-XF medium and FBS-based αMEM complete medium, then analyzed the morphology, immu- nophenotype, expansion potential, trilineage differentiation potential, karyotype and immunosuppression of early passages. Results The average cell diameters of UC-MSCs in suspension cultured by serum-free medi- um and FBS-based medium were 26 （18 -39） μm and 35 （20 -61 ） μm, respectively. Cell expansion folds with serum free medium and FBS-based medium were （5.2 ± 0.2） and （3.5 ± 0.1 ） respectively, in the first five passages. The expansion potential of serum-free medium cultured UC-MSCs was significantly higher than FBS-based medium cultured ones （P 〈 0.05）. A panel of markers CD29, CD44, CDg0, CD73, CD105 and HLA-ABC expressed on human UC-MSCs. Hematopoietic lineage markers CD34, CIM5 and HLA-DR were not detectable on UC-MSCs. The cpm were （4.57 ±0.14） × 104, （2.04 ±0.16） × 104 and（0. 42 ± 0. 04） × 104 , respectively when serum-free medium cultured MSCs were added to the cultures at MSCs/T cell ratios of 1： 100, 1：10 and 1： 5. While the cpm was （4.57 ±0. 14） × 104, （2.04±0. 16） × 104 and （0.42± 0.04） × 104, respectively when serum-free medium cultured UC-MSCs were added to the cultures. The immunosuppressive potential of serum-free medium-cultured UC-MSCs was higher than serum-contained medium cultured ones at three different MSC/T cell ratios （ P 〈 0.05 ）. Cortdusion Compare with serum- contained medium cultured early passages of UC-MSCs, the cell diameter of serum-free medium cultured UC- MSCs was smaller with higher expansion potential. No xenogeneie

关 键 词：脐带间充质干细胞 胎牛血清 无血清培养基 异种蛋白 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]