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目的用重组质粒pBudCE4.1_Cx43转染大鼠骨髓间充质干细胞(BMSCs),观察缝隙连接蛋白43(connexin43,Cx43)的mRNA和蛋白的表达及细胞间隙连接通讯功能的变化。方法用脂质体转染的方法将重组表达质粒pBudCE4.1_Cx43转染细胞;用逆转录聚合酶链式反应(RT-PCR)检测Cx43mRNA表达,免疫细胞化学法检测Cx43蛋白的表达,划痕染料示踪法检测细胞间通讯功能。结果转染Cx43基因后的细胞在形态学、生长能力等方面的特征与未转染的细胞比较没有明显变化;Cx43mRNA与蛋白的相对表达量和对照组相比差异显著(P<0.01);转染组细胞传递的荧光强度与对照组相比亦差异显著(P<0.01)。结论转染Cx43基因的BMSCs能表达有生物学活性的基因产物,改善细胞间通讯功能,能为细胞性心肌成形术提供较理想的基因工程细胞。
OBJECTIVE: To investigate the expression of connexin43 (Cx43) mRNA and the changes of cell gap junctional intercellular communication in rat bone marrow mesenchymal stem cells (BMSCs) transfected with recombinant plasmid pBudCE4.1_Cx43. Methods The recombinant plasmid pBudCE4.1_Cx43 was transfected into the cells by liposome transfection. The expression of Cx43 mRNA was detected by reverse transcriptase-polymerase chain reaction (RT-PCR), the expression of Cx43 protein by immunocytochemistry, Tracer method to detect intercellular communication function. Results Compared with untransfected cells, the expression of Cx43 gene in Cx43 cells showed no significant difference in morphological and growth characteristics. The relative expression of Cx43 mRNA and protein was significantly different from that in control group (P <0.01) Fluorescence intensity of transfected cells was also significantly different from that of the control group (P <0.01). Conclusion BMSCs transfected with Cx43 gene can express biologically active gene products, improve the intercellular communication function, and provide ideal gene engineering cells for cardiomyoplasty.