中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2013年
7期
1357-1359
,共3页
肖军%马德彰%杨钟华%张山锋%刘军
肖軍%馬德彰%楊鐘華%張山鋒%劉軍
초군%마덕창%양종화%장산봉%류군
成人骨髓间充质干细胞%淫羊藿苷%微团培养%软骨分化
成人骨髓間充質榦細胞%淫羊藿苷%微糰培養%軟骨分化
성인골수간충질간세포%음양곽감%미단배양%연골분화
Adults' mesenchymal stem cells%Icariin%Pellet culture%Chondrogenic differentiation
目的 探讨应用淫羊藿苷促进人骨髓间充质干细胞(hMSCs)成软骨分化的可行性及效果.方法 体外分离培养hMSCs,取第3代细胞实验.将hMSCs分别用无血清高糖培养基(H-DMEM)和含1×10-8、1×10-7、1×10-6、1×10-5mol/L淫羊藿苷的软骨诱导液(CM)诱导,显微镜下观察细胞形态变化,噻唑蓝(MTT)法检测各组细胞的增殖.以5×109/L的细胞密度离心构建微团,诱导培养3周.逆转录-聚合酶链反应(RT-PCR)检测各组蛋白多糖和Ⅱ型胶原mRNA的表达.免疫组织化学、甲苯胺蓝染色检测Ⅱ型胶原和蛋白多糖表达.结果 5组微团分别由无血清H-DMEM、含1 × 10-8、1×10-7、1×10-6、1×10-5mol/L淫羊藿苷的CM诱导3周后,除H-DMEM组无Ⅱ型胶原和蛋白多糖表达外,其他各组Ⅱ型胶原蛋白和蛋白多糖的表达呈浓度依赖性增加,ImagePro Plus 6.0软件测得各组Ⅱ型胶原平均吸光度(A)值分别为0.0214±0.0035、0.1182 ±0.0105、0.1886 ±0.0184、0.2903±0.0261、0.3476±0.0287,各组差异均有统计学意义(P＜0.05).结论 淫羊藿苷能促进hMSCs成软骨分化,软骨分化程度呈浓度依赖性增加.
目的 探討應用淫羊藿苷促進人骨髓間充質榦細胞(hMSCs)成軟骨分化的可行性及效果.方法 體外分離培養hMSCs,取第3代細胞實驗.將hMSCs分彆用無血清高糖培養基(H-DMEM)和含1×10-8、1×10-7、1×10-6、1×10-5mol/L淫羊藿苷的軟骨誘導液(CM)誘導,顯微鏡下觀察細胞形態變化,噻唑藍(MTT)法檢測各組細胞的增殖.以5×109/L的細胞密度離心構建微糰,誘導培養3週.逆轉錄-聚閤酶鏈反應(RT-PCR)檢測各組蛋白多糖和Ⅱ型膠原mRNA的錶達.免疫組織化學、甲苯胺藍染色檢測Ⅱ型膠原和蛋白多糖錶達.結果 5組微糰分彆由無血清H-DMEM、含1 × 10-8、1×10-7、1×10-6、1×10-5mol/L淫羊藿苷的CM誘導3週後,除H-DMEM組無Ⅱ型膠原和蛋白多糖錶達外,其他各組Ⅱ型膠原蛋白和蛋白多糖的錶達呈濃度依賴性增加,ImagePro Plus 6.0軟件測得各組Ⅱ型膠原平均吸光度(A)值分彆為0.0214±0.0035、0.1182 ±0.0105、0.1886 ±0.0184、0.2903±0.0261、0.3476±0.0287,各組差異均有統計學意義(P＜0.05).結論 淫羊藿苷能促進hMSCs成軟骨分化,軟骨分化程度呈濃度依賴性增加.
목적 탐토응용음양곽감촉진인골수간충질간세포(hMSCs)성연골분화적가행성급효과.방법 체외분리배양hMSCs,취제3대세포실험.장hMSCs분별용무혈청고당배양기(H-DMEM)화함1×10-8、1×10-7、1×10-6、1×10-5mol/L음양곽감적연골유도액(CM)유도,현미경하관찰세포형태변화,새서람(MTT)법검측각조세포적증식.이5×109/L적세포밀도리심구건미단,유도배양3주.역전록-취합매련반응(RT-PCR)검측각조단백다당화Ⅱ형효원mRNA적표체.면역조직화학、갑분알람염색검측Ⅱ형효원화단백다당표체.결과 5조미단분별유무혈청H-DMEM、함1 × 10-8、1×10-7、1×10-6、1×10-5mol/L음양곽감적CM유도3주후,제H-DMEM조무Ⅱ형효원화단백다당표체외,기타각조Ⅱ형효원단백화단백다당적표체정농도의뢰성증가,ImagePro Plus 6.0연건측득각조Ⅱ형효원평균흡광도(A)치분별위0.0214±0.0035、0.1182 ±0.0105、0.1886 ±0.0184、0.2903±0.0261、0.3476±0.0287,각조차이균유통계학의의(P＜0.05).결론 음양곽감능촉진hMSCs성연골분화,연골분화정도정농도의뢰성증가.
Objective To explore the feasibility and effect of icariin on promoting chondrogenic differentiation of human bone marrow mesenchymal stem cells (hMSCs).Methods hMSCs were isolated and cultured in vitro,and the cells from Passage 3 were used in this study.hMSCs were randomly grouped into five according to the kinds of induction medium-serum-free DMEM-H,chondrogenic medium supplemented with 1 × 10-8,1 × 10-7,1 × 10-6,1 × 10-5 mol/L icariin.The proliferation of hMSCs was investigated by methyl thiazol tetrazolium (MTT) assay.The cells were resuspended and pellets were constructed by centrifugation at a density of 5 × 109/L,and then continued to be kept for three weeks.Reverse transcriptase-polymerase chain reaction (RT-PCR) was performed to detect the expression of mRNA of type Ⅱ collagen and aggrecan.Immunohistochemistry and toluidine blue staining were performed to detect the expression of type Ⅱ collagen and proteoglycan.Results After three weeks' induction,Group A did not express mRNA of type Ⅱ collagen and aggrecan,the mRNA transcripts of type Ⅱ collagen and proteoglycan in other four groups kept increasing in a concentration-dependent manner.The mean absorbance (A) values of collagen type Ⅱ were,respectively,(0.0214 ± 0.0035),(0.1182 ± 0.0105),(0.1886 ±0.0184),(0.2903 ±0.0261),(0.3476 ±0.0287) with significant differences (P＜0.05).Conclusion Icariin can promote chondrogenic differentiation of hMSCs.Treated with a certain concentration of icariin,the expression of type Ⅱ collagen and proteoglycan increases in a concentration-dependent manner.
