Ad-hBMP-2对促进软骨细胞分泌Ⅱ型胶原影响的实验研究  

The effect of Ad-hBMP-2 accelerating cartilage cells excreting collagen type Ⅱ

作　　者：王丽红[1] 何英[1] 赖国旗[2] 

机构地区：[1]重庆医科大学附属第一医院,重庆400016 [2]重庆医科大学实验动物中心

出　　处：《中国老年学杂志》2010年第18期2631-2634,共4页Chinese Journal of Gerontology

基　　金：重庆市卫生局基金资助项目〔2008〕38号

摘　　要：目的观察腺病毒-人骨形态发生蛋白-2-绿色荧光蛋白(Ad-hBMP-2-GFP)转染兔软骨细胞后,hBMP-2蛋白的表达和其对软骨细胞分泌Ⅱ型胶原功能的影响。方法包装hBMP-2-GFP腺病毒载体,测定病毒滴度。分离培养并鉴定原代兔软骨细胞,通过基因转染技术,转染Ad-hBMP-2-GFP至兔第三代软骨细胞,RT-PCR检测hBMP-2mRNA的表达,细胞免疫荧光法和Western印迹检测软骨细胞中hBMP-2蛋白的表达和Ⅱ型胶原的变化情况,用未转染组和空病毒组(Ad-GFP)做对照。结果转染后,hBMP-2的mRNA在48h有明显阳性条带表达,对照组呈阴性,hBMP-2蛋白在48、72h的表达量随时间而逐渐增强(P<0.05),同时,软骨细胞分泌Ⅱ型胶原的量也逐渐增加(P<0.05)。结论Ad-hBMP-2-GFP能成功转染兔软骨细胞,并能促进软骨细胞分泌Ⅱ型胶原,为软骨组织工程提供功能良好的种子细胞,且为hBMP-2基因治疗骨性关节炎提供实验依据。Objective To investigate the expression of hBMP-2 protein and secretion of collagen type Ⅱ in rabbit chondrocytes after transfection of the Adenovirus-human bone morphogenetic Ⅱ in rabbit chondrocytes after transfection of the Adenovirus-human bone morphogenetic protein-2-green fluorescent protein（Ad-hBMP-2-GFP）vector.Methods Virus titer was determined,rabbit＇s articular chondrocytes was primary cultured and used after the third passage,transfected with Ad-hBMP2-GFP.Non-transfected group and empty virus group were used as the controls.hBMP-2 mRNA was determined by RT-PCR,the expression of BMP2 and collagen type Ⅱ protein was detected by Western blot and immunofluorescence assay.Results After transfection,hBMP-2 mRNA expression was obviously positive at 48 hour while control groups were negative.hBMP-2 protein expression was gradually increased in 72 h after transfection（P0.05）.At the same time,secretion of collagen type Ⅱ increased（P0.05）.Conclusions hBMP-2 gene can be successfully transfected to articular chondrocytes and promote the secretion of collagen type Ⅱ,which provides the experimental evidence for chondrocytes as the better seeded cells of cartilage tissue engineering and the treatment of osteoarthritis with gene.

关 键 词：软骨细胞 人骨形态发生蛋白2 基因 转染 

分 类 号：R33[医药卫生—人体生理学]