CAJ | 학술논문

背景：体外培养的脐带间充质干细胞在不同培养体系下生长状态差异显著，因此选取一种更适合的培养基相当必要。目的：对比观察人脐带间充质干细胞在3种培养基中的生长增殖情况，并检测细胞免疫表型以及间充质干细胞的诱导分化能力。方法：在无菌条件下用贴块法收获人脐带间充质干细胞，用T75培养瓶培养传代后，取第3代脐带间充质干细胞分别种入到含体积分数为5%胎牛血清的DMEM/F12培养基、含体积分数为10%胎牛血清的DMEM/F12培养基和Mesen PRO RSTM培养基中，培养第1，3，5，7天进行细胞计数，绘制细胞生长曲线。采用流式细胞仪分析第3代脐带间充质干细胞免疫表型，并检测其成骨及成脂肪诱导分化能力。结果与结论：培养出的第3代人脐带间充质干细胞高表达CD44、CD73、CD90、CD105，不表达CD29、CD31、CD34、HLA-DR。经成脂诱导后，油红O染色可见胞浆中有大量红色小脂滴；成骨诱导后，茜素红染色后镜下可见成骨样细胞团，说明脐带间充质干细胞在体外具有多向分化的潜能。在倒置显微镜下观察可见含体积分数为10%胎牛血清的DMEM/F12培养基中的细胞集落密集，形态均匀，而其他2种培养基中的细胞集落密集程度和形态都不如前者好。在培养传代细胞时，可优先选择含体积分数为10%胎牛血清的DMEM/F12培养基。
배경：체외배양적제대간충질간세포재불동배양체계하생장상태차이현저，인차선취일충경괄합적배양기상당필요。목적：대비관찰인제대간충질간세포재3충배양기중적생장증식정황，병검측세포면역표형이급간충질간세포적유도분화능력。방법：재무균조건하용첩괴법수획인제대간충질간세포，용T75배양병배양전대후，취제3대제대간충질간세포분별충입도함체적분수위5%태우혈청적DMEM/F12배양기、함체적분수위10%태우혈청적DMEM/F12배양기화Mesen PRO RSTM배양기중，배양제1，3，5，7천진행세포계수，회제세포생장곡선。채용류식세포의분석제3대제대간충질간세포면역표형，병검측기성골급성지방유도분화능력。결과여결론：배양출적제3대인제대간충질간세포고표체CD44、CD73、CD90、CD105，불표체CD29、CD31、CD34、HLA-DR。경성지유도후，유홍O염색가견포장중유대량홍색소지적；성골유도후，천소홍염색후경하가견성골양세포단，설명제대간충질간세포재체외구유다향분화적잠능。재도치현미경하관찰가견함체적분수위10%태우혈청적DMEM/F12배양기중적세포집락밀집，형태균균，이기타2충배양기중적세포집락밀집정도화형태도불여전자호。재배양전대세포시，가우선선택함체적분수위10%태우혈청적DMEM/F12배양기。
BACKGROUND:The growth of mesenchymal stem cells in vitro in different conditioned media is different evidently. So it is necessary to choose a more suitable medium. OBJECTIVE:To contrast and observe the proliferation of human umbilical cord mesenchymal stem cells in three kinds of media and to check the immunophenotype and differentiation ability of mesenchymal stem cells. METHODS:Human umbilical cord mesenchymal stem cells were col ected by explant method in sterile conditions. After subculturing by T75 incubation bottles, the third generation of mesenchymal stem cells were cultured in Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 medium containing 5%fetal bovine serum, Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 containing 10%fetal bovine serum and Mesen PRO RSTM medium. After 1, 3, 5, 7 days of culture, the cells were counted to draw a growth curve. Immunophenotype of the third generation of umbilical cord mesenchymal stem cells were determined by flow cytometry and the ability of osteogenic and adipogenic differentiation was also detected. RESULTS AND CONCLUSION:The third generation of cells cultured highly expressed CD44, CD73, CD90, CD105, but did not express CD29, CD31, CD34, HLA-DR. The oil red O staining showed a lot of little red lipid drops after adipogenic induction;alizarin red staining showed osteoblast-like cells after osteogenic induction, indicating umbilical cord mesenchymal stem cells in vitro have the potential of multi-directional differentiation. After observation and counting, the colony and shape of cells cultured in Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 containing 10%fetal bovine serum were superior to those cultured in the other two media. Therefore, it is concluded that Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 containing 10%fetal bovine serum is preferred for cellsubculture.