腺病毒5型早期区1A基因对人肝癌细胞株HepG2辐射敏感性的影响及其作用机制

Effect of adenovirus type 5 early region 1A gene on radiosensitivity of human hepatoma cell line HepG2 and its mechanism

目的探讨腺病毒5型早期区1A(E1A)基因对人肝癌细胞HepG2辐射敏感性的影响及其作用机制。方法取对数生长期HepG2细胞分为HepG2-E1A组、HepG2-vect组、HepG2组(未转染的HepG2细胞)。HepG2-E1A组、HepG2-vect组以聚乙烯亚胺纳米凝胶(PEI)为载体,分别将含E1A基因的重组质粒psv-E1A、未携带E1A基因的质粒转染进入HepG2细胞中。检测转染后3组HepG2细胞中E1A mRNA及蛋白的表达情况,以及细胞周期分布情况。3组细胞均分别接受0、1、2、4和8 Gy的6 MV-X线照射,72 h后检测3组的细胞存活率,24 h及72 h后检测3组的细胞凋亡率。结果在HepG2-E1A组的细胞中可见E1A mRNA、E1A蛋白表达。当辐射剂量为2 Gy及以上时,HepG2-E1A组细胞的存活率均低于其他两组细胞(P<0.05);以1 Gy、2 Gy、4 Gy、8 Gy的剂量分别照射24 h及72 h后,HepG2-E1A组细胞的凋亡率均高于其他两组(P<0.05)。与HepG2-vect组和HepG2组相比,HepG2-E1A组G2/M期细胞比例升高,而S期细胞比例降低(均P<0.05),但3组G1期细胞比例比较差异无统计学意义(P>0.05)。结论 PEI能将质粒psv-E1A转染至肝癌细胞。EIA基因能够明显提高人肝癌细胞对辐射的敏感性,其作用机制可能与E1A促进凋亡并导致S期抑制、G_2期阻滞有关。

Objective To investigate the effect of adenovirus type 5 early region 1 A（ E1 A） gene on radiosensitivity of human hepatoma cell line HepG2 and its mechanism. Methods HepG2 cells in logarithmic phase were divided into HepG2-E1 A group,HepG2-vect group and HepG2 group（ HepG2 cells without being transfected）. Recombinant plasmid psv-E1 A containing E1 A gene and plasmid without E1 A gene were transfected into HepG2 cells of the HepG2-E1 A group and the HepG2-vect group by the vector of polyethylenimine（ PEI） respectively. The expressions of E1 A mRNA and protein in HepG2 cells,and the cell cycle distribution were detected in the three groups after transfection. The cells in the three groups received radiation of 6 MV-X ray at the doses of 0,1,2,4 and 8 Gy,respectively,then the cell survival rate was assessed in the three group after 72 hours of detection. The cell apoptosis rate was analyzed in the three group after 24 and 72 hours of detection. Results The cells of the HepG2-E1 A group expressed E1 A mRNA and protein. The cell survival rates of the HepG2-E1 A group were lower than those of the other two groups at the radiation dose of 2 Gy and above（ P〈0. 05）. After 24 and 72 hours of radiation at the dose of 1,2,4 and 8 Gy,the cell apoptosis rates of the HepG2-E1 A group were higher than those of the other two groups（ P〈0. 05）. In the HepG2-E1 A group,the proportion of cells in G2/M phase increased and the proportion of cells in S phase decreased compared to the HepG2-vect group and the HepG2 group（ all P〈0. 05）. There was no significant difference in the proportion of cells in G1 phase among the three groups（ P〈0. 05）.Conclusion PEI can transfect plasmid psv-E1 A gene into hepatoma cells. E1 A gene can effectively enhance radiosensitivity of human hepatoma cells,which may be related to its effects on promoting apoptosis,and lead to S phase suppression and G_2 phase arrest.