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目的观察急性白血病骨髓基质细胞(acute leukemia bone marrow stromal cells,ALBMSCs)经腺病毒介导的间隙连接蛋白43(Cx43)基因修饰后Cx43基因表达的变化以及对细胞间隙连接通讯(GJIC)功能的影响。方法用重组腺病毒Ad-Cx43-GFP转染ALBMSCs,荧光显微镜观察报告基因GFP的表达并计算转染效率,RT-PCR法、Westernblot法和免疫细胞化学法检测ABMSCs转染前后Cx43基因及蛋白表达情况,通过染料传输实验观察细胞间隙连接通讯功能的变化。结果荧光显微镜下观察可见ALBMSCs在转染Ad-Cx43-GFP后24h即有绿色荧光表达,计数绿色荧光细胞的百分率得出转染效率为(82.7±2.16)%;RT-PCR法、Westernblot法和免疫细胞化学法检测显示转染Ad-Cx43-GFP后ALBM-SCsCx43基因及蛋白表达较转染前增强(P<0·01);转染Ad-Cx43-GFP后ALBMSCs GJIC功能较转染前增强(P<0·01)。结论腺病毒介导Cx43基因修饰ALBMSCs后可上调其Cx43基因的表达并增强GJIC功能。
Objective To investigate the changes of Cx43 gene expression in acute leukemia bone marrow stromal cells (AMSCs) induced by adenovirus mediated gap junction protein 43 (Cx43) gene and its effect on the function of GJIC . Methods The recombinant adenovirus Ad-Cx43-GFP was used to transfect ALBMSCs. The expression of reporter gene GFP was observed by fluorescence microscopy and the transfection efficiency was calculated. The expression of Cx43 gene and protein was detected by RT-PCR, Western blot and immunocytochemistry The situation, through the dye transmission experiment observation cell gap junction communication function changes. Results The results of fluorescence microscopy showed that the expression of ALBMSCs was green fluorescence at 24h after transfected with Ad-Cx43-GFP, and the percentage of transfected cells was (82.7 ± 2.16)%. The RT-PCR, Western blot and The results of immunocytochemistry showed that the expression of ALBM-SCsCx43 gene and protein increased after transfection with Ad-Cx43-GFP (P <0.01). The GJIC function of ALBMSCs transfected with Ad-Cx43-GFP was enhanced (P <0.01). Conclusion Adenovirus-mediated Cx43 gene modified ALBMSCs can up-regulate the expression of Cx43 gene and enhance the function of GJIC.