【摘要】 目的观察导入pcDNA3.1-HRE-JAB1真核表达质粒的肺癌A549细胞,在低氧环境下对健择(Gemzar)的化疗敏感性,以期找到一种提高肺癌化疗敏感性的方法。方法首先构建缺氧反应元件启动的pcDNA3.1-HRE-JAB1真核表达质粒,鉴定后将该质粒导入肺癌A549细胞。在低氧环境培养过程中加入化疗药物Gemzar,观察空白组(A549)、空载体组(A549+pcD-NA3.1-HRE)和质粒组(A549+pcDNA3.1-HRE-JAB1)肺癌A549细胞对Gemzar的敏感性。采用Real-time PCR和Westernblot分别检测各组肺癌A549细胞中JAB1的mRNA和蛋白表达水平;流式细胞术检测各组肺癌A549细胞周期分布和凋亡情况。结果通过双酶切鉴定,确定构建获得了pcDNA3.1-HRE-JAB1质粒。在有化疗药物Gemzar的情况下,质粒组分别与空白组或空载体组比较,质粒组肺癌A549细胞中JAB1mRNA和蛋白表达水平明显增加(P<0.01),细胞凋亡率明显增加(P<0.01),而细胞周期被明显阻滞于G1期(P<0.01)。结论 JAB1在低氧环境下高表达提高了药物Gemzar化疗的敏感性,这将可能成为一种理想的提高肺癌或其他实体恶性肿瘤化疗敏感性的手段。更多还原

【Abstract】 Objective To find the way of improving the lung cancer chemotherapeutic sensitivity,we investigated whether the lung cancer A549 cells introduced eukaryotic expression plasmid pcDNA3.1-HRE-JAB1 plasmid have the chemotherapeutic sensibility to Gemzar in anaerobic condition.Methods We first constructed an eukaryotic expression plasmid pcDNA3.1-HRE-JAB1 driven by hypoxia response elements promoter.Then,the plasmid was introduced into lung cancer cell line A549.After that,chemotherapeutic drug such as Gemzar was used to treat the A549 cells in anaerobic condition,and investigated that the normal control(A549),the empty vector group(A549+pcDNA3.1-HRE) and the plasmid group(A549+pcDNA3.1-HRE-JAB1)A549 cells had chemotherapeutic sensitivity.qRT-PCR and Western blot was used to assay the mRNA and protein level of JAB1.Cell cycle and apoptosis of A549 cells were also assayed according to flow cytometry.Results According to bi-enzyme digestion evaluation,obtained plasmid was pcDNA3.1-HRE-JAB1 plasmid.The results showed that JAB1 in the A549 was overexpressed after the transfection of pcDNA3.1-HRE-JAB1 compared with the normal control and the empty vector(P<0.01).The cell proliferation was arrested at G1 phase and the cell apoptosis was significantly enhanced after the transfection(P<0.01).Conclusion JAB1 overexpression significantly increase sensitivity of lung cancer cells to the chemotherapeutic drug Gemzar in anaerobic condition,which might provide an efficient strategy of improving the chemotherapeutic sensitivity of lung cancer or the other cancers.更多还原