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目的 探讨外源性小鼠Cbfa1基因在兔皮肤成纤维细胞中获得瞬时表达的可行性。方法 在阳离子脂质体介导下 ,将含外源性小鼠Cbfa1基因的 pSG5 Cbfa1真核表达质粒导入原代培养的第 2代兔皮肤成纤维细胞内。采用RT PCR及Western Blot法检测Cbfa1基因在兔皮肤成纤维细胞内的瞬时表达。同时采用RT PCR法检测细胞内骨钙素及碱性磷酸酶基因的表达情况。结果 转染pSG5 Cbfa1真核表达质粒的兔皮肤成纤维细胞内有大量小鼠Cbfa1mRNA转录及蛋白瞬时表达 ,同时诱导骨钙素mRNA及碱性磷酸酶mRNA大量表达。结论 在阳离子脂质体介导下 ,小鼠Cbfa1基因能够导入兔皮肤成纤维细胞内并获得瞬时表达 ,同时诱导细胞内成骨特异性骨钙素基因及碱性磷酸酶基因的转录
Objective To investigate the feasibility of exogenous mouse Cbfa1 gene transient expression in rabbit skin fibroblasts. Methods The pSG5 Cbfa1 eukaryotic expression plasmid containing exogenous mouse Cbfa1 gene was introduced into primary cultured rabbit skin fibroblasts, which were induced by cationic liposome. The transient expression of Cbfa1 gene in rabbit skin fibroblasts was detected by RT PCR and Western Blot. At the same time, the expression of osteocalcin and alkaline phosphatase in cells was detected by RT PCR. Results A large number of mouse Cbfa1 mRNA transcripts and protein transient expression were observed in rabbit skin fibroblasts transfected with pSG5 Cbfa1 eukaryotic expression plasmids. Osteocalcin mRNA and alkaline phosphatase mRNA were also induced in large numbers. Conclusion Under the condition of cationic liposome, mouse Cbfa1 gene can be introduced into rabbit dermal fibroblasts and transiently expressed, meanwhile, osteoblast-specific osteocalcin and alkaline phosphatase gene transcription can be induced