Wang Xiaohua, Miao Jingcheng, Xie Yufeng, Sheng Weihua, Shan Yunbo, Yang Jicheng
No. 1 Affiliated Hospital of Soochow University, Suzhou 215001, China.
Sheng Wu Gong Cheng Xue Bao. 2009 Jul;25(7):1035-41.
We constructed the recombinant adenovirus vector expressing IL-24 and E1A (Ad-IL-24-E1A) and investigated the inhibition of Ad-IL-24-E1A on SMMC-7721 hepatocellular carcinoma in vitro. We amplified IL-24 gene by PCR using pAdTrack-IL-24 as template. The IL-24 gene was cloned into pAdTrack-IRES at the Bgl II and Sal I site to form pAdTrack-IL-24-IRES. E1A digested from pAdTrack-E1A was cloned into the pAdTrack-IL-24-IRES at the Xho I and EcoR V site to form the pAdTrack-IL-24-IRES-E1A. We co-transformed both pAdTrack-IL-24-IRES-E1A and pAdeasy-1 digested by Pme I and packaged to obtain Ad-IL-24-E1A. Ad-IL-24-E1A at 50 MOI infected SMMC-7721 cells. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay determined cell proliferation. Flow cytometry detected Cell apoptosis. The apoptotic rate of SMMC-7721 cells was 52% 48 h after infection with Ad-IL-24-E1A. The result showed that the growth of SMMC-7721 cells was significantly inhibited by Ad-IL-24-E1A at the MOI of 50.
我们构建了表达IL-24和E1A的重组腺病毒载体（Ad-IL-24-E1A），并在体外研究了Ad-IL-24-E1A对SMMC-7721肝癌细胞的抑制作用。我们以pAdTrack-IL-24为模板通过PCR扩增IL-24基因。将IL-24基因在Bgl II和Sal I位点克隆到pAdTrack-IRES中，形成pAdTrack-IL-24-IRES。从pAdTrack-E1A中酶切得到的E1A在Xho I和EcoR V位点克隆到pAdTrack-IL-24-IRES中，形成pAdTrack-IL-24-IRES-E1A。我们将经Pme I酶切的pAdTrack-IL-24-IRES-E1A和pAdeasy-1共转化并包装，以获得Ad-IL-24-E1A。以50 MOI的Ad-IL-24-E1A感染SMMC-7721细胞。采用3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）法检测细胞增殖。通过流式细胞术检测细胞凋亡。Ad-IL-24-E1A感染SMMC-7721细胞48小时后，细胞凋亡率为52%。结果表明，在50 MOI时，Ad-IL-24-E1A对SMMC-7721细胞的生长有显著抑制作用。
