CAJ | 학술논문

目的观察CD105-生物素-亲和素系统促进滑膜间充质干细胞的黏附和CD105阳性细胞的筛选,探讨软骨形成的效果.方法酶消化获取滑膜干细胞,应用流式细胞仪检测细胞表面CD29、CD34、CD44、CD45、CD90、CD105的表达.分3组(A:未处理+干细胞;B:亲和素化+生物素化干细胞;C:亲和素化+ CD105-生物素化干细胞),通过细胞吸光度值和显微镜下随机视野计数分别测定不同时间点细胞黏附数量;免疫荧光检测CD105阳性细胞的筛选.成软骨诱导3周后,通过苏木素-伊红(HE)染色、阿尔新蓝染色和Ⅱ型胶原免疫荧光观察3组软骨形成.结果滑膜干细胞贴壁生长,呈星形或梭形;CD29、CD44、CD90阳性表达,CD34、CD45阴性表达,CD105阳性率约为42％.C组和B组早期黏附细胞数量增加,与A组比较在1、2h差异有统计学意义(P＜0.05),B组虽然多于C组,但差异无统计学意义(P＞0.05).免疫荧光显示C组黏附的CD105阳性细胞在1、2h均明显高于B组和A组.软骨诱导培养后,3组均合成了蛋白聚糖和Ⅱ型胶原,但是C组软骨特异性细胞外基质最多.结论 CD105-生物素-亲和素系统既增加了滑膜干细胞的黏附,又筛选了CD105阳性细胞,更好的促进了软骨形成.
목적 관찰CD105-생물소-친화소계통촉진활막간충질간세포적점부화CD105양성세포적사선,탐토연골형성적효과.방법 매소화획취활막간세포,응용류식세포의검측세포표면CD29、CD34、CD44、CD45、CD90、CD105적표체.분3조(A:미처리+간세포;B:친화소화+생물소화간세포;C:친화소화+ CD105-생물소화간세포),통과세포흡광도치화현미경하수궤시야계수분별측정불동시간점세포점부수량;면역형광검측CD105양성세포적사선.성연골유도3주후,통과소목소-이홍(HE)염색、아이신람염색화Ⅱ형효원면역형광관찰3조연골형성.결과 활막간세포첩벽생장,정성형혹사형;CD29、CD44、CD90양성표체,CD34、CD45음성표체,CD105양성솔약위42％.C조화B조조기점부세포수량증가,여A조비교재1、2h차이유통계학의의(P＜0.05),B조수연다우C조,단차이무통계학의의(P＞0.05).면역형광현시C조점부적CD105양성세포재1、2h균명현고우B조화A조.연골유도배양후,3조균합성료단백취당화Ⅱ형효원,단시C조연골특이성세포외기질최다.결론 CD105-생물소-친화소계통기증가료활막간세포적점부,우사선료CD105양성세포,경호적촉진료연골형성.
Objective To observe the CD105-biotin-avidin promoting synovium-derived mesenchymal stem cells (SMSCs) adhesion and selecting CD105-positive cells in the cartilage tissue engineering,and evaluate its effects on cartilage formation.Methods SMSCs were obtained by enzyme digestion.Flow cytometry was used to detect markers on cell surface,including CD29,CD34,CD44,CD45,CD90 and CD105.With divided 3 groups (A:untreated + cells;B:avidin + biotin-cells;C:avidin + CD105-biotin-cells),adhesion cell number was counted by absorbance and random field count at different time point.CD105-positive cells were tested through immunofluorescence.After 3 weeks chondrogenic induction,hematoxylin-eosin (HE),alcian blue staining and immunofluorescence were used to assess chondrogenesis.Results SMSCs showed star or spindle shape with adherent growth.The expression of CD29,CD44 and CD90 was positive,while CD34 and CD45 negative.The positive rate of CD105 was about 42％.Compared with Group A,Group B and C significantly increased cells adhesion at 1 and 2 h (P ＜ 0.05).Although Group B had more cells than C,there was no statistical difference.Immunofluorescence showed CD105-positive cells in Group C were significantly higher than that in Group B and A.After chondrogenic induction,while proteoglycan and type Ⅱ collagen were observed in all groups,there was most cartilage specific extracellular matrix in Group C.Conclusion CD105-biotin-avidin system could not only enhance cells adhesion,but also select CD105-positive cells,which may promote chondrogenesis better.