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目的观察经全反式维甲酸(all-trans retinoic acid,ATRA)诱导后,单纯疱疹病毒胸苷酸激酶/丙氧鸟苷系统(HSV-TK/GCV)对人肝癌细胞SMMC-7721旁观者效应的影响。方法免疫组化法观察经全反式维甲酸诱导前后SMMC-7721细胞连接蛋白Cx32的表达;将含有HSV-TK基因的质粒通过静电吸附在脂质微泡表面上,采用超声辐照转染并用倒置显微荧光镜及RT-PCR观察TK基因的转入及表达;MTT法观察经全反式维甲酸诱导后HSV-TK/GCV对肝癌细胞SMMC-7721的旁观者效应。结果经全反式维甲酸诱导后SMMC-7721细胞膜上Cx32表达明显增加,而胞质内仅少量表达;通过超声辐照后HSV-TK基因可顺利转入SMMC-7721细胞中并稳定表达;与对照组相比,经全反式维甲酸诱导组可明显提高HSV-TK/GCV对肝癌细胞SMMC-7721的旁观者效应(P<0.05)。结论全反式维甲酸可提高SMMC-7721细胞中连接蛋白Cx32的表达且能正确定位于细胞膜上,这可能与其能增强HSV-TK/GCV系统对肝癌细胞SMMC-7721的旁观者效应有关。
Objective To observe the bystander effect of HSV-TK / GCV on human hepatoma SMMC-7721 cells induced by all-trans retinoic acid (ATRA) Impact. Methods Immunohistochemistry was used to observe the expression of connexin Cx32 in SMMC-7721 cells induced by all-trans retinoic acid. Plasmids containing HSV-TK gene were electrostatically adsorbed on the surface of lipid microbubbles and were transfected by using ultrasound irradiation Inverted microfluorimetry and RT-PCR were used to observe the transfection and expression of TK gene. The bystander effect of HSV-TK / GCV on SMMC-7721 cells was observed by MTT assay. Results The expression of Cx32 in SMMC-7721 cells was significantly increased after transfected with all-trans retinoic acid and only a small amount in cytoplasm. The HSV-TK gene was transfected into SMMC-7721 cells by ultrasound irradiation and stably expressed. Compared with the control group, all-trans retinoic acid induced group could significantly improve the bystander effect of HSV-TK / GCV on SMMC-7721 hepatoma cells (P <0.05). Conclusion All-trans retinoic acid enhances the expression of connexins Cx32 in SMMC-7721 cells and their localization in the cell membrane, which may be related to the enhancement of the bystander effect of HSV-TK / GCV system on SMMC-7721 hepatoma cells.