HBV x基因真核表达载体的构建及对肝癌细胞恶性表型的影响

目的 构建乙型肝炎病毒x基因真核表达载体并在肝癌细胞中表达，以研究其对肝癌细胞恶性表型的影响。方法 应用PCR、基因克隆等方法构建乙型肝炎病毒x基因真核表达载体pcDNA3.1-HBX，用基因转染的方法将pcDNA3.1-HBX转染入肝癌细胞HCC－9204，筛选稳定表达乙型肝炎病毒X蛋白的肝癌细胞。MTT比色实验、平板集落形成实验检测肝癌细胞恶性表型。结果 成功构建了乙型肝炎病毒x基因真核表达载体pcDNA3.1-HBX并获得了稳定表达乙型肝炎病毒X蛋白的肝癌细胞系HCC－9204细胞克隆，且MTT比色实验、平板克隆形成实验显示稳定表达乙型肝炎病毒X蛋白的肝癌细胞恶性度增高。结论 乙型肝炎病毒X蛋白具有生长因子样作用，可刺激肝癌细胞生长，在肝癌细胞中稳定表达乙型肝炎病毒X蛋白可增加肝癌细胞恶性表型。

Construction and expression of eukaryotic expression vector of HBV x gene and its effect on the malignant phenotype of hepatocellular carcinoma cells

AIM To construct and express the eukaryotic expression vector of HBV x gene and study its effect on the malignant phenotype of hepatocellular carcinoma cells. METHODS Polymerase chain reaction (PCR) and other molecular biological methods were used to construct the eukaryotic expression vector of HBV x gene pcDNA3.1 HBX. Then gene transfection mediated by the lipofectamine was used to introduce the eukaryotic expression vector of HBV x gene into human hepatocellular carcinoma cell line HCC 9204. The selective medium containing G418 was used to select the cell clones which express the X protein of HBV constantly. MTT colorimetric analysis experiment and agarose colony forming were used to study the role of the X protein in the malignant phenotype and growth of hepatocellular carcinoma cells. RESULTS The eukaryotic expression vector of HBV x gene was successfully constructed. The cell clones which expressed the X protein constantly were obtained by the selective medium containing G418. Cells which constitutively expressed the X protein had a relatively higher malignant phenotype. CONCLUSION The X protein has growth factor like activities and can stimulate the growth of hepatocellular carcinoma cells and enhance its malignant phenotype.