腺病毒介导的碱性成纤维细胞生长因子对人骨关节炎软骨细胞的作用

Effects of Adenovirus-Mediated bFGF Gene Transfer on Human Osteoarthritis Chondrocytes in Vitro

目的:探讨腺病毒介导的人碱性成纤维细胞生长因子(bFGF)对体外培养的人骨关节炎(OA)软骨细胞的转染及其作用。方法:人OA软骨细胞分为3组:OA对照组、绿色荧光蛋白(EGFP)对照组及bFGF转染组。采用含bFGF的腺病毒载体转染人OA软骨细胞,48 h后通过流式细胞术分析腺病毒转染软骨细胞的效率,绘制转染效率曲线。6 d后检测培养基中bFGF的浓度及软骨细胞增殖;观察软骨细胞蛋白多糖及Ⅱ型胶原的表达。结果:腺病毒介导的bFGF可高效转染人OA软骨细胞并在细胞外表达。与OA对照组相比,bFGF转染后可明显促进软骨细胞的增殖(P<0.05);同时增加了软骨细胞蛋白多糖和Ⅱ型胶原的生物合成(P<0.05)。结论:腺病毒介导的bFGF可促进OA软骨细增殖,增强细胞基质的合成,对软骨细胞表型的维持具有重要作用。

Objective： To investigate the effect of recombinant adenovirus-mediated basic fibroblast growth factor（bFGF） gene transfer on the human osteoarthritis（OA） chondrocytes.Methods： Monolayer cultures of human OA chondrocytes were divided into three groups： ①OA control group（chondrocytes without transfection）;②OA chondrocytes transfected with AdEGFP;③OA chondrocytes transfected with AdbFGF.Human OA chondrocytes were infected with recombinant adenovirus containing human bFGF gene.The rate of transfection was analyzed by flow cytometry and the diagram of expression was drawn after 48 hours.Six days later,levels of gene expression in culture supernatant were detected.The proliferation of chondrocytes was analyzed by methyl thiazolyl tetrazolium（MTT）.The biosynthesis of proteoglycans and type Ⅱ collagen was observed by toluidine blue staining and immunohistochemistry examination.Results： The expression level of bFGF was high following adenoviral transfection in culture supernatant.The delivery of bFGF promoted the cell proliferation（P〈0.05） and resulted in a significant enhanced biosynthesis of proteoglycans and type Ⅱ collagen of chondrocytes（P〈0.05）.Conclusion： Recombinant adenovirus-mediated bFGF gene transfection can greatly promote the cell proliferation and increase matrix synthesis in vitro,which is important to keep the phenotype of chondrocytes.