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目的研究将人骨形成蛋白３（ＢＭＰ３）的ｃＤＮＡ构建于真核，表达载体ＰｃＤＮＡ３，形成重组ＤＮＡ ＰｃＤＮＡ３－ｈＢＭＰ３，转染兔关节软骨细胞，以探讨基因转染对关节软骨细胞增殖的影响。方法利用重组ＤＮＡ和基因克隆技术构建重组ＤＮＡ；用细胞培养和基因转染技术体外转染兔关节软骨细胞；用核酸杂交和蛋白电泳检测细胞ＢＭＰ３的表达，最后通过ＦＣＭ、ＤＮＡ和葡糖醛酸含量检测以及Ⅱ型胶原探针原位杂交分析其对增殖的影响。结果软骨细胞６次传代后，Ⅱ型胶原原位杂交的灰度值降低，同样条件下转染的软骨细胞仍保持较高水平Ⅱ型胶原的表达；转染的软骨细胞经流式细胞仪的分析，软骨细胞Ｓ期细胞比例增多，说明细胞ＤＮＡ的合成增加；经ＤＮＡ和葡糖醛酸含量测定，转染的软骨细胞ＤＮＡ和葡糖醛酸含量较转染前有明显提高。结论ｈＢＭＰ３对维持软骨细胞的表型具有十分重要的作用，在脂质体的介导下，ＤＮＡ ＰｃＤＮＡ３－ｈＢＭＰ３转染兔关节软骨细胞获得成功。
Objective To study the effect of gene transfection on the proliferation of articular chondrocytes by constructing the cDNA of human bone morphogenetic protein-3 (BMP3) in eukaryotic and expression vector pcDNA3 to form recombinant pcDNA3-hBMP3. Methods Recombinant DNA was constructed by recombinant DNA and gene cloning technique. Rabbit articular chondrocytes were transfected by cell culture and gene transfection in vitro. The expression of BMP3 was detected by nucleic acid hybridization and protein electrophoresis. Finally, FCM, DNA and glucuronic acid Content detection and type Ⅱ collagen probe in situ hybridization analysis of its proliferation. Results After 6 passages of chondrocytes, the gray value of type Ⅱ collagen in situ hybridization decreased. Under the same conditions, the transfected chondrocytes still maintained high levels of type Ⅱ collagen. The transfected chondrocytes were transfected by flow cytometry Analysis, the proportion of cells in S phase of chondrocytes increased, indicating that the synthesis of DNA increased; DNA and glucuronic acid content of DNA and glucuronic acid content of transfected chondrocytes significantly increased compared with before transfection. Conclusion hBMP3 plays an important role in maintaining the phenotype of chondrocytes. Under the guidance of liposome, hBMP3 transfected rabbit articular chondrocytes successfully.