超声靶向微泡破坏技术介导基因JNK1在小鼠肝癌细胞株中表达及对细胞迁移和侵袭作用的研究

摘要

Objective To explore the effect on the expression of JNK1 in mouse hepatocellular carcinoma cell lines Hca-P and cell migration and invasion mediated by ultrasound-targeted microbubble Destruction (UTMD).Methods The vector of pCDNA3.1-JNK1 was built and identified.The hepatocellular cell lines Hca-P were cultured in vitro and divided into the following groups:Group A (control group),group B (plasmid group),group C (lipofection group),group D (ultrasound microbubbles combined with ultrasound irradiation group),group E (lipofection combined with ultrasound microbubbles and ultrasound irradiation group).The transfection rate was observed by inverted fluorescence microscope.The expression levels of JNK1 mRNA and protein were evaluated by fluorescence quantitative PCR and Western Blot respectively.The cell activity was detected by CCK-8.The ability of migration and invasion in vitro was detected by transwell assay.Results The vector of pCDNA3.1-JNK1 was built.The transfection rate of group E was higher than those of group B,C and D (all P＜0.05);there was no significant difference between group C and D (P＞0.05).The expression levels of JNK1 mRNA and protein were highest in group E (all P ＜0.05).The cell viability,the abilities of migration and invasion increased most in group E (all P＜0.05).Conclusions The transfection rate of gene JNK1 can be improved through the combination of lipofection and UTMD in mouse hepatocellular cell lines,therefore enhancing the up-regulation of gene JNK1 expression.The effects on the cell viability,migration and invasion can also be enhanced.%目的 探讨应用超声靶向微泡破坏(UTMD)技术介导小鼠肝癌细胞株Hca-P JNK1基因的表达以及肝癌细胞迁移和侵袭.方法 构建并鉴定pCDNA3.1-JNK1载体.将小鼠肝癌细胞株Hca-P分组如下:A组(空白对照组),B组(质粒组),C组(脂质体+质粒组),D组(超声微泡+质粒+超声辐照组),E组(脂质体+超声微泡+质粒+超声辐照组).采用倒置荧光显微镜观察各组细胞的转染率,荧光定量PCR和Western Blot法检测JNK1的基因和蛋白质表达水平,以CCK-8法检测5组细胞的细胞活性,应用Transwell实验检测各组细胞的体外迁移和侵袭能力.结果 成功构建了pCDNA3.1-JNK1载体.各组转染率比较:E组均大于B、C、D组(P均＜0.05),C、D组转染率均高于B组(P均＜0.05),C、D组之间差异无统计学意义(P＞0.05).E组JNK1 mRNA和蛋白表达水平均高于其他各组(P均＜0.05);E组细胞活性、迁移和侵袭能力均高于其他各组(P均＜0.05)结论 UTMD技术结合脂质体转染法可以提高pCDNA3.1-JNK1载体转染小鼠肝癌细胞株Hca-P的效率,加大基因JNK1表达的上调幅度,增强细胞活性、迁移和侵袭能力.