Departments of aOccupational Therapy bGeneral Surgery cInternal Medicine, Division of Infectious Diseases, E-Da Hospital, I-Shou University dDepartment of Kinesiology, Health and Leisure Studies, National University of Kaohsiung eSchool of Pharmacy, Kaohsiung Medical University, Kaohsiung, Taiwan.
Anticancer Drugs. 2014 Apr;25(4):456-61. doi: 10.1097/CAD.0000000000000082.
In this study, we collected 44 hepatitis B virus surface antigen positivity HBsAg (+) tumor and nontumor hepatocellular tissues from hepatocellular carcinoma (HCC) patients during hepatectomy, and quantified the APOBEC3G (A3G) mRNA by using a real-time PCR. Our results showed higher expression of A3G mRNA in the nontumor tissues than in the tumor tissues of the HBsAg (+) HCC patients. To further investigate this phenomenon, we constructed a pLV-A3G vector and transfected it into the human HCC cell line, Hep 3B. The results of an immunofluorescence analysis showed the overexpression of A3G in the cytoplasm. We then evaluated A3G cytotoxicity by using a cell viability assay (MTS assay), the results of which showed that Hep 3B cell viability was 88 and 58% after the transfection of pLV and pLV-A3G, respectively, indicating the growth inhibitory effects of A3G on Hep 3B cells. To further evaluate the tumor suppressive effects of A3G, we used a plastic pipette tip to scratch Hep 3B cells grown on a culture dish (to 70-80% confluence) after transfection with pLV-A3G. Our data indicated a ratio of wound closure of 100% in the control cells and in the pLV-expressing cells, compared with 43% in the pLV-A3G-overexpressing cells, 72 h after the wound scratch, as observed using phase-contrast microscopy. These results indicated that A3G inhibits wound healing in Hep 3B cells. Overall, our results suggest that A3G inhibits the growth of human hepatoma cells.
在这项研究中，我们从肝癌（HCC）患者的肝切除术中收集了 44 例乙型肝炎病毒表面抗原阳性（HBsAg（+））肿瘤和非肿瘤肝细胞组织，并使用实时 PCR 定量了 APOBEC3G（A3G）mRNA。我们的结果显示，HBsAg（+）HCC 患者的非肿瘤组织中 A3G mRNA 的表达高于肿瘤组织。为了进一步研究这种现象，我们构建了 pLV-A3G 载体并将其转染到人肝癌细胞系 Hep 3B 中。免疫荧光分析的结果显示 A3G 在细胞质中的过表达。然后，我们通过细胞活力测定（MTS 测定）评估了 A3G 的细胞毒性，结果显示 pLV 和 pLV-A3G 转染后 Hep 3B 细胞活力分别为 88%和 58%，表明 A3G 对 Hep 3B 细胞的生长抑制作用。为了进一步评估 A3G 的肿瘤抑制作用，我们使用塑料移液管尖端划痕 Hep 3B 细胞培养皿（至 70-80%汇合）转染 pLV-A3G 后。我们的数据表明，在划痕后 72 小时，与对照细胞和 pLV 表达细胞相比，pLV-A3G 过表达细胞的伤口闭合率为 100%，而 pLV-A3G 过表达细胞的伤口闭合率为 43%，相差 72%。相差显微镜观察。这些结果表明 A3G 抑制 Hep 3B 细胞的伤口愈合。总体而言，我们的结果表明 A3G 抑制人肝癌细胞的生长。
