论文部分内容阅读
目的研究腺病毒5型早期区1A(E1A)基因对人肝癌细胞的放射增敏作用及其机制。方法利用多聚乙烯亚胺(PEI)载体将E1A基因转染进人肝癌细胞(7721系)中。7721细胞、转染空载体的细胞(7721-vect)及转染EIA基因的细胞(7721-E1A)分别接受0、2、4、6和8 Gy的6 MV-X线照射,用MTT法检测细胞存活率,观察EIA基因对7721细胞放射敏感性的影响;采用流式细胞术检测E1A基因对细胞凋亡以及细胞周期分布的影响;采用免疫组化染色(SP)法检测E1A基因对人表皮生长因子受体2(Her-2)及核因子κB(NF-κB)表达的影响。结果照射后,7721-E1A细胞存活率明显低于7721细胞[至8 Gy时,(7.91±4.44)%vs.(32.50±9.41)%)](P<0.01)。7721-E1A的凋亡率明显高于7721细胞[至8 Gy时,(88.38±15.06)%vs.(42.67±9.49)%](P<0.01)。7721-E1A的细胞周期中S期显著低于7721细胞[(25.16±7.45)%vs.(40.88±11.44)%](P<0.05),而G2期明显高于7721细胞[(40.69±6.74)%vs.(15.63±5.02)%](P<0.01)。转染EIA基因后能抑制Her-2及NF-κB的表达。结论 EIA基因能够明显提高人肝癌细胞在体外对放射线的敏感性。其作用机制可能与E1A抑制Her-2及NF-κB的表达,促进凋亡并导致S期抑制、G2期阻滞有关。
Objective To investigate the radiosensitization effect of adenovirus 5 type 1A (E1A) gene on human hepatoma cells and its mechanism. Methods E1A gene was transfected into human hepatocellular carcinoma cell line (7721 strain) by polyethylenimine (PEI) vector. 7721 cells, cells transfected with empty vector (7721-vect) and cells transfected with EIA gene (7721-E1A) were exposed to 6 MV-X radiation at 0, 2, 4, 6 and 8 Gy, respectively, Cell viability was measured to observe the effect of EIA gene on the radiosensitivity of 7721 cells. The effect of E1A gene on apoptosis and cell cycle distribution was analyzed by flow cytometry. The effect of E1A gene on the proliferation of human epidermal cells was analyzed by immunohistochemical staining (SP) On the expression of Her-2 and NF-κB. Results After irradiation, the viability of 7721-E1A cells was significantly lower than that of 7721 cells [(7.91 ± 4.44)% vs (32.50 ± 9.41)% at 8 Gy] (P <0.01). The apoptosis rate of 7721-E1A was significantly higher than that of 7721 cells [(88.38 ± 15.06)% vs (42.67 ± 9.49)%] at 8 Gy (P <0.01). The cell cycle of 7721-E1A was significantly lower in S phase than in 7721 cells [(25.16 ± 7.45)% vs (40.88 ± 11.44)%] (P <0.05) % vs. (15.63 ± 5.02)%] (P <0.01). Transfection of EIA gene can inhibit the expression of Her-2 and NF-κB. Conclusion EIA gene can significantly enhance the sensitivity of human hepatoma cells to radiation in vitro. Its mechanism may be related to the inhibition of E1A Her-2 and NF-κB expression, promote apoptosis and lead to S phase inhibition, G2 phase block.