中华血液学杂志
中華血液學雜誌
중화혈액학잡지
Chinese Journal of Hematology
2012年
9期
715-719
,共5页
杨婷%陈广华%薛胜利%乔曼%刘慧文%田竤%乔淑敏%陈峰%陈志哲%孙爱宁%吴德沛
楊婷%陳廣華%薛勝利%喬曼%劉慧文%田竤%喬淑敏%陳峰%陳誌哲%孫愛寧%吳德沛
양정%진엄화%설성리%교만%류혜문%전굉%교숙민%진봉%진지철%손애저%오덕패
脐带间充质干细胞%胎牛血清%无血清培养基%异种蛋白
臍帶間充質榦細胞%胎牛血清%無血清培養基%異種蛋白
제대간충질간세포%태우혈청%무혈청배양기%이충단백
Mesenchymal stem cell%Umbilical cord%Fetal calf serum%Serum free medium%Xenogeneic protein
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aMEM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、CDl05、HLA-ABC抗原,不表达CD45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共培养的放射性核索每分钟闪烁计数(CPM)值分别为(4.57 4-0.14)× 104、(2.04±0.16)×104和(0.42±0.04)×104,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)×104、(2.55±0.15)×104和(0.52±0.03)×104,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.
目的 比較無血清及含胎牛血清培養基培養的人臍帶間充質榦細胞(UC-MSC)生物學特性,尋求無血清的UC-MSC培養方法.方法 採用膠原酶消化臍帶小塊.分彆採用MesenCult-XF無血清培養基和含胎牛血清的aMEM完全培養基培養UC-MSC.比較兩種方法培養的早期UC-MSC細胞形態、免疫錶型、增殖分化潛能、覈型及對T淋巴細胞增殖抑製作用的差異.結果 用無血清培養基培養製備的懸浮UC-MSC平均細胞直徑為26(18～39)μm,含血清完全培養基製備的懸浮UC-MSC平均細胞直徑為35(20～61)μm.早期UC-MSC採用無血清培養基培養時連續傳代可倍增(5.2±0.2)倍,而採用含胎牛血清的完全培養基培養時傳代可倍增(3.5±0.1)倍,UC-MSC採用無血清培養體繫傳代倍增數明顯高于含胎牛血清的培養體繫的傳代倍增數(P<0.05).UC-MSC錶達CD29、CD44、CD90、CD73、CDl05、HLA-ABC抗原,不錶達CD45及CD34等造血細胞抗原.無血清培養的UC-MSC按MSC:T細胞比例為1:100、1:10及1:5共培養的放射性覈索每分鐘閃爍計數(CPM)值分彆為(4.57 4-0.14)× 104、(2.04±0.16)×104和(0.42±0.04)×104,含胎牛血清製備的UC-MSC共培養對應的CPM值分彆為(5.29±0.18)×104、(2.55±0.15)×104和(0.52±0.03)×104,無血清培養的UC-MSC抑製T細胞增殖作用較含血清培養的UC-MSC抑製作用更明顯(P<0.05).結論 與含胎牛血清的完全培養基相比無血清培養的人UC-MSC細胞直徑小、早期傳代增殖潛能增加、無異種蛋白.無血清培養基製備的UC-MSC抑製T細胞增殖活性高于含胎牛血清培養基製備的UC-MSC.
목적 비교무혈청급함태우혈청배양기배양적인제대간충질간세포(UC-MSC)생물학특성,심구무혈청적UC-MSC배양방법.방법 채용효원매소화제대소괴.분별채용MesenCult-XF무혈청배양기화함태우혈청적aMEM완전배양기배양UC-MSC.비교량충방법배양적조기UC-MSC세포형태、면역표형、증식분화잠능、핵형급대T림파세포증식억제작용적차이.결과 용무혈청배양기배양제비적현부UC-MSC평균세포직경위26(18～39)μm,함혈청완전배양기제비적현부UC-MSC평균세포직경위35(20～61)μm.조기UC-MSC채용무혈청배양기배양시련속전대가배증(5.2±0.2)배,이채용함태우혈청적완전배양기배양시전대가배증(3.5±0.1)배,UC-MSC채용무혈청배양체계전대배증수명현고우함태우혈청적배양체계적전대배증수(P<0.05).UC-MSC표체CD29、CD44、CD90、CD73、CDl05、HLA-ABC항원,불표체CD45급CD34등조혈세포항원.무혈청배양적UC-MSC안MSC:T세포비례위1:100、1:10급1:5공배양적방사성핵색매분종섬삭계수(CPM)치분별위(4.57 4-0.14)× 104、(2.04±0.16)×104화(0.42±0.04)×104,함태우혈청제비적UC-MSC공배양대응적CPM치분별위(5.29±0.18)×104、(2.55±0.15)×104화(0.52±0.03)×104,무혈청배양적UC-MSC억제T세포증식작용교함혈청배양적UC-MSC억제작용경명현(P<0.05).결론 여함태우혈청적완전배양기상비무혈청배양적인UC-MSC세포직경소、조기전대증식잠능증가、무이충단백.무혈청배양기제비적UC-MSC억제T세포증식활성고우함태우혈청배양기제비적UC-MSC.
Objective To compare the differences of biological characteristics between human umbilical cord-derived mesenchymal stem cells (UC-MSCs) cultured by serum-free medium or fetal bovine serum-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 MesenCult-XF medium and FBS-based aMEM 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 medium 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, CD90, CD73, CD105 and HLA-ABC expressed on human UC-MSCs. Hematopoietic lineage markers CD34, CD45 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) ×l04, 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 ). Conclusion 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 xenogeneic proteins were presented in UC-MSCs preparations when cultured with serum-free medium. Human UC-MSCs suppressed T-eell proliferation in a dose-dependent manner. The immunosuppressive potential of serum-free medium cultured UC-MSCs was higher than FBS-based medium cultured ones.
