摘要
探讨绿色荧光蛋白(hrGFP)标记技术在组织工程骨体外构建中对大鼠骨髓间质干细胞(rBMSCs)的示踪作用。用293FT细胞包装含hrGFP基因的慢病毒载体,收集病毒液转染rBMSCs,转染后细胞持续表达绿色荧光,流式细胞仪检测转染率81.3%。将hrGFP标记的rBMSCs与生物玻璃-胶原蛋白-透明质酸-磷酸丝氨酸支架复合培养体外构建组织工程骨。荧光显微镜下观察到绿色荧光的细胞在支架表面很好地黏附、增殖。成骨诱导后,表达绿色荧光的细胞内部及支架表面均有COLⅠ的阳性表达。说明HrGFP标记技术能直观有效地观察组织工程骨体外构建过程中细胞的黏附、生长及成骨分化等行为。
This study was designed to label rat bone marrow mesenchymal stem cells(rBMSCs) with humanized renillar green fluorescent protein(hrGFP) for exploring the detectable effects of hrGFP on the construction of tissue-engineered bone in vitro.The hrGFP expression plasmid was packed into lentivirus by 293FT cells and transduced into rat BMSCs.After transduction,81.3% rBMSCs successfully expressed green florescence.The hrGFP-rBMSCs were statically loaded on Bioglass-Collagen-Hyaluronic Acid-Phosphatidylserine(BG-COL-HYA-PS) scaffold in complete L-DMEM medium or osteogenic medium for 14 days.At 6h,a number of cells expressing green fluorescence can be observed by fluorescent microscopy.At day 7 and 14 after co-culture,the number of cells on the scaffold gradually increased.After 14 days for osteogenic induction,the hrGFP-rBMSCs and the interior of scaffold can be detected the expression of type Ⅰ collagen.The results demonstrate that hrGFP labelling technique can detect visualizedly and effectively cell adhesion,proliferation and osteogenic differentiation in the construction of tissue-engineered bone in vitro.
出处
《生物医学工程学杂志》
EI
CAS
CSCD
北大核心
2011年第1期121-124,137,共5页
Journal of Biomedical Engineering
基金
国家自然科学基金重点项目资助(B09B5070230)
广东省自然科学基金博士启动项目资助(9451064101002989)
关键词
绿色荧光蛋白
骨髓间质干细胞
组织工程骨
体外构建
Green fluorescent protein(GFP)
Bone marrow mesenchymal stem cells(BMSCs)
Tissue-engineered bone
Construction in vitro