1 Central Laboratory, First Affiliated Hospital of Fujian Medical University , Fuzhou, China .
2 Key Lab of Radiation Biology, Fujian Universities , Fuzhou, China .
Cancer Biother Radiopharm. 2018 May;33(4):146-154. doi: 10.1089/cbr.2017.2346.
There is a high incidence of nasopharyngeal carcinoma (NPC), malignant head and neck tumors, in southern China. Radioresistance is the main cause affecting the efficacy of NPC treatments. The POLG gene particularly plays an important role in radiation-induced damage repair. In this study, the authors established RNAi CNE-1 and CNE-2 knockdown in two NPC cell lines to observe whether this gene affects the radiosensitivity of NPC cells.
Four short hairpin RNA (shRNA) expression plasmids targeting POLG gene were constructed and transfected into the NPC cell lines CNE-1 and CNE-2. Screening was performed to evaluate the stable expression of cloned cells, which were named CNE-1/POLG-shRNA1, CNE-1/POLG-shRNA2, CNE-2/POLG-shRNA1, and CNE-2/POLG-shRNA2. The negative controls CNE-1/Neg-shRNA and CNE-2/Neg-shRNA were additionally used. The MTT method, flow cytometry, clone formation analysis, cell migration, and other experimental methods were employed to verify changes in the radiosensitivity of the NPC cells.
Fluorescent quantitative PCR and Western blot confirmed the downregulation of the PLOG gene through diminished PLOG messenger RNA and protein levels. Consequently, the authors report the stable knockdown of the POLG gene in an NPC model. Dose-dependent radiation exposure of POLG inhibited NPC cell growth and increased apoptosis compared with control cells (p < 0.01), as demonstrated through colony formation assay and flow cytometry. Functional assays indicated that knockdown of the POLG in CNE-1 and CNE-2 cells remarkably reduced cell viability and proliferation. Specifically, POLG knockdown led to G1 phase arrest and apoptosis.
Overall, the authors conclude that POLG downregulation alters the radiosensitivity of NPC cells, indicating that the gene is likely involved in conferring the radiation response of the cells. In addition, findings in this study suggest a novel role for POLG as a potential predictive marker for NPC radiotherapy efficiency. POLG gene can be used as a potential clinical target to effectively improve the radiosensitivity of NPC.
中国南方鼻咽癌（NPC）和头颈部恶性肿瘤的发病率较高。放射抵抗是影响 NPC 治疗效果的主要原因。POLG 基因在辐射诱导损伤修复中尤其发挥着重要作用。本研究通过构建 NPC 细胞系 CNE-1 和 CNE-2 的 POLG 基因 RNAi 敲低，观察该基因是否影响 NPC 细胞的放射敏感性。
构建针对 POLG 基因的 4 个短发夹 RNA（shRNA）表达质粒并转染 NPC 细胞系 CNE-1 和 CNE-2。筛选克隆细胞以评估稳定表达，分别命名为 CNE-1/POLG-shRNA1、CNE-1/POLG-shRNA2、CNE-2/POLG-shRNA1 和 CNE-2/POLG-shRNA2。另外还使用阴性对照 CNE-1/Neg-shRNA 和 CNE-2/Neg-shRNA。采用 MTT 法、流式细胞术、克隆形成分析、细胞迁移等实验方法验证 NPC 细胞放射敏感性的变化。
荧光定量 PCR 和 Western blot 证实 POLG 基因通过降低 POLG 信使 RNA 和蛋白水平而下调。因此，作者报告了 NPC 模型中 POLG 基因的稳定敲低。与对照组细胞相比，POLG 抑制 NPC 细胞生长并增加细胞凋亡（p<0.01），通过集落形成实验和流式细胞术证实。功能测定表明，CNE-1 和 CNE-2 细胞中 POLG 的敲低显著降低了细胞活力和增殖。具体而言，POLG 敲低导致 G1 期阻滞和细胞凋亡。
总体而言，作者得出结论，POLG 下调改变 NPC 细胞的放射敏感性，表明该基因可能参与赋予细胞的放射反应。此外，本研究的结果表明 POLG 作为 NPC 放疗效率的潜在预测标志物具有新的作用。POLG 基因可作为提高 NPC 放射敏感性的潜在临床靶点。
