HBx基因缺失突变体(HBx-d382)对L02细胞cyclinD1、cyclinG1和E2F1表达的影响

Influence of HBx gene deletion mutation(HBx-d382) on cyclin D1,cyclinG1 and E2F1 expression in L02 cells

目的前期的研究成功构建了稳定表达HBx基因及其缺失突变体(HBx-d382)的L02细胞,分别命名为L02/HBx和L02/HBx-d382,并证实其可导致肝细胞恶性转化。该研究进一步探讨HBx-d382对L02细胞G1期细胞周期调控相关基因cyclinD1,cyclinG1和E2F1表达的影响。方法实验分为L02/pcDNA3.0(稳定转染pcDNA3.0)、L02/HBx和L02/HBx-d382(分别稳定转染质粒pcDNA3.0/HBx和pcDNA3.0/HBx-d382)3组。通过实时定量PCR以及wester-blot检测转染HBx基因及其缺失突变体HBx-d382后L02细胞cyclinD1、cyclinG1和E2F1表达的改变。方法实时定量PCR以及wester-blot结果表明稳定表达HBx基因或HBx-d382的L02细胞cyclinD1、cyclinG1和E2F1表达上调,表达HBx-d382的L02细胞上调最为明显。结论 HBx-d382可能通过上调cyclinD1、cyclinG1和E2F1从而影响细胞G1期调控,进一步导致肝细胞恶性转化。

[Objective] Previously we had established L02 cell lines stably expressing HBx gene or its deletion mutant （HBx-d382）, which were shown to be implicated in hepatoeytes malignant transformation, named L02/HBx and L02/HBx-d382 respectively. This study further investigated the influence of HBx-d382 on cyclin D1, cyelin G1 and E2F1 expression associated with G1 cell cycle control in I1）2 cells. [ Methods] Cell lines LO2/pcDNA3, L02/ HBx and L02/HBx-d382 （stably transfeeted with plasmid pcDNA3.0, pcDNA3.0/HBx and pcDNA3.0/HBx-d382, respectively） were evaluated in this study. The mRNA and protein expression of eyclin D1, cyclin G1, E2F1 were detected by Real-time Quantitative PCR and Western blot respectively. [ Results ] Real-time Quantitative PCR and Western blot showed that the expression of cyclin D1, cyclin G1 and E2F1 were up-regulated in L02/HBx and L02/ HBx-d382 cell lines ,which was more significant in L02/HBx-d382 group. [ Conclusion ] HBx-d382 up-regulates the expression of cyclin D1, cyclin G1 and E2F1 in L02 cells which influences the G1 cell cycle control and may further contribute to liver carcinogenesis.