Cong Hui, Yao Ru-Yong, Sun Zhen-Qing, Qiu Wen-Sheng, Yao Ya-Sai, Feng Tong-Tong, Xin Chao, Liang Jun, Yue L U
Department of Oncology, Affiliated Hospital of Qingdao University, Qingdao, Shandong 266003, P.R. China; Tumor Molecular and Translational Medicine Lab, Affiliated Hospital of Qingdao University, Qingdao, Shandong 266003, P.R. China.
Central Laboratory, Affiliated Hospital of Qingdao University, Qingdao, Shandong 266003, P.R. China.
Oncol Lett. 2016 Jan;11(1):842-848. doi: 10.3892/ol.2015.3937. Epub 2015 Nov 18.
The vimentin gene is a hallmark of epithelial-to-mesenchymal transition and has been observed to be overexpressed in various types of tumor cell line and tissue. Previous studies have reported correlations between vimentin DNA methylation levels and subsequent vimentin expression levels in solid tumors, including breast and colorectal cancer; however, to the best of our knowledge, such a correlation has not been reported for gastric cancer (GC) using Lauren classification. Therefore, the present study aimed to quantify DNA methylation levels of the vimentin gene using quantitative (q) methylation-specific polymerase chain reaction (PCR) in intestinal-type GC cell lines (MKN-28, AGS and MKN-1), diffuse-type GC cell lines (SGC-7901, SNU-5 and KATO III), the GES-1 immortalized human non-neoplastic gastric epithelial cell line, as well as in tumor and paratumor normal tissue samples. Furthermore, the present study analyzed the messenger RNA expression of the vimentin gene in these cell lines and tissues by reverse transcription-qPCR. A comparison of the clinicopathological features was conducted between patients, grouped according to the Lauren classification. The present study identified that the vimentin promoter region was hypermethylated in all GC cell lines and tumor tissue samples when compared with immortalized normal gastric epithelial cells and paratumor normal tissues. In addition, vimentin promoter methylation levels were observed to be higher in intestinal-type cell lines when compared with those of diffuse-type lines and tissues. Correspondingly, vimentin expression levels were lower in intestinal-type gastric cell lines compared with those of diffuse-type cell lines and tissues, and were lowest in the non-neoplastic gastric cell line and paratumor normal tissues. Patients with diffuse-type GC were on average younger (P=0.023), and exhibited higher tumor (P=0.020), node (P=0.032) and TNM classification of malignant tumor stage (P=0.039) than those with intestinal-type GC. Following treatment of AGS cells (which demonstrated the highest methylation level of the vimentin gene) with 5-aza-2'-deoxycytidine, vimentin expression was restored significantly. Thus, the present study revealed that vimentin promoter methylation levels are inversely correlated with vimentin expression levels in GC (according to Lauren classification). High levels of methylation in the vimentin gene promoter region may be involved in carcinogenesis and the development of GC, and may provide a novel molecular classification for GC.
波形蛋白基因是上皮-间质转化的标志，已观察到其在各种类型的肿瘤细胞系和组织中过表达。先前的研究报道了实体瘤（包括乳腺癌和结直肠癌）中波形蛋白DNA甲基化水平与随后的波形蛋白表达水平之间的相关性；然而，据我们所知，尚未有使用劳伦分类法对胃癌（GC）进行此类相关性报道。因此，本研究旨在使用定量（q）甲基化特异性聚合酶链反应（PCR）来量化波形蛋白基因在肠型GC细胞系（MKN-28、AGS和MKN-1）、弥漫型GC细胞系（SGC-7901、SNU-5和KATO III）、GES-1永生化人非肿瘤性胃上皮细胞系以及肿瘤和瘤旁正常组织样本中的DNA甲基化水平。此外，本研究通过逆转录-qPCR分析了这些细胞系和组织中波形蛋白基因的信使RNA表达。根据劳伦分类法对患者进行分组，比较了他们的临床病理特征。本研究发现，与永生化正常胃上皮细胞和瘤旁正常组织相比，所有GC细胞系和肿瘤组织样本中的波形蛋白启动子区域均发生了高甲基化。此外，观察到肠型细胞系中的波形蛋白启动子甲基化水平高于弥漫型细胞系和组织。相应地，肠型胃细胞系中的波形蛋白表达水平低于弥漫型细胞系和组织，且在非肿瘤性胃细胞系和瘤旁正常组织中最低。弥漫型GC患者的平均年龄比肠型GC患者年轻（P = 0.023），且肿瘤（P = 0.020）、淋巴结（P = 0.032）和恶性肿瘤TNM分期（P = 0.039）更高。用5-氮杂-2'-脱氧胞苷处理波形蛋白基因甲基化水平最高的AGS细胞后，波形蛋白表达显著恢复。因此，本研究表明，GC中波形蛋白启动子甲基化水平与波形蛋白表达水平呈负相关（根据劳伦分类法）。波形蛋白基因启动子区域的高甲基化水平可能参与了GC的致癌过程和发展，并可能为GC提供一种新的分子分类方法。
