中文摘要：

目的 通过观察转染生肌调节因子（myoblast determination，MyoD）基因和连接蛋白43（Connexin）基因的大鼠真皮成纤维细胞（demml fibroblast，DFs）生物学功能的变化，探讨Myo和Cx43基因转染DFs细胞的意义。方法采用Gateway技术，构建真核质粒表达载体，利用慢病毒（LV）表达系统，将大鼠MyoD cDNA和Cx43 cDNA转入大鼠成纤维细胞中，经Blasticidin筛选培养，通过RT-PCR，Western blot及膜片钳技术等方法检测MyoD及Cx43 mRNA及蛋白表达，并检测离子电流变化，通过显微镜观察转染后生长情况，免疫组织化学检测肌动蛋白和结蛋白的表达。结果RT-PCR，Westem blot检测出MyoD及Cx43的mRNA及相应蛋白表达，膜片钳检测到转染后钙离子电流，显微镜观察到基因转染筛选后，培养1周细胞的融合现象，并有多核肌管形成。免疫组织化学检测可见其下游蛋白Desmin及α-actin呈现阳性表达。结论MyoD和Cx43基因转染使DFs分化为成肌细胞，为进一步研究基因治疗心力衰竭奠定基础。

英文摘要：

Objective To study changes on the differentiation growth and biological function of cultured rat fibreblasts （FB） transfected by myoblast determining （MyoD） and connexin 43（Cx43） genes in order to explore prehminarily the possible mechanism and way by which MyoD and Cx43 genes on treatment of heart failure （HF）. Methods Gene cloning technology was used to construct the eukaryotic expressed plasmid vector pLenti6/V5-DEST-MyoD and cells were pLenti6/V5-DEST-Cx43 in which MyoD cDNA or Cx43 cDNA was inserted. DFs transfected with exogenetic MyoD cDNA or Cx43 cDNA via hpofectamine, and followed by Blasticidin （50 μg/ml） selection, according to lentiviral expression system （ViraPower） protocol. Expression and its biological fuctions of MyoD and Cx43 in the transfectants were testified by RT-PCR, Western blot, molecular and clamp patch methods. The mophological structure changes of cells before and after transfection were observed with microscopy. Immunotfistochemical methods indicated the expression of skeletal α-actin and desmin respectively in these cells. Results The expression of MyoD and Cx43 was detected in the MyoD and Cx43 genes transfected DFs with RT-PCR and Western blot. Myotube were found from cultures incubated a week in differentiation medium, which the transfected cells were of characteristic of filaments in their cytoplasm and presented myoblast morphlogy. Conclusion MyoD cDNA could induce cultured DFs to diferentiate into myoblasts and Cx43 cDNA could enhance gap junctional intercellular communication between cell and cell, furtherly providing an experimental foundation for the therapy of HF.