Silencing of glutathione peroxidase 3 through DNA hypermethylation is associated with lymph node metastasis in gastric carcinomas.
Silencing of glutathione peroxidase 3 through DNA hypermethylation is associated with lymph node metastasis in gastric carcinomas.
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DOI：
10.1371/journal.pone.0046214
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
El-Rifai W
中科院分区：
文献类型：
--
作者：
Peng DF;Hu TL;Schneider BG;Chen Z;Xu ZK;El-Rifai W
胃癌仍然是世界上癌症相关死亡的第二大原因。H.幽门螺杆菌感染是胃癌的主要危险因素，它产生高水平的活性氧（ROS）。谷胱甘肽过氧化物酶3（GPX 3）是血浆中的GPX成员，是ROS的主要清除剂，催化还原型谷胱甘肽还原过氧化氢和脂质过氧化物。为了研究GPX 3的表达及其基因调控，我们采用实时荧光定量RT-PCR方法检测了9种胃癌细胞株、108例胃癌组织和45例癌旁正常胃粘膜组织中GPX 3基因的表达。GPX 3在9种胃癌细胞系中有8种表达下调或沉默，胃癌组织中GPX 3表达下调或沉默的比例为83%（90/108），与癌旁正常组织相比差异有统计学意义（P<0.0001）。GPX 3基因启动子甲基化检测结果显示，6/9的胃癌细胞系和60%的胃癌组织中存在GPX 3基因启动子甲基化（焦磷酸测序法测定甲基化水平≥10%）（P = 0.007）。 胃癌组织中GPX 3基因拷贝数明显减少（P<0.001）。用5-氮杂-2 ′脱氧胞苷处理SNU 1和MKN 28细胞，GPX 3基因表达恢复，GPX 3启动子显著去甲基化。GPX 3基因表达下调和启动子区甲基化与胃癌淋巴结转移显著相关（P = 0.018和P = 0.029）。 我们还观察到在大约三分之一的肿瘤邻近正常胃组织样本中GPX 3的下调、DNA拷贝数丢失和启动子高甲基化，这表明在肿瘤样本附近区域存在场缺陷。GPX 3在AGS细胞中的重建降低了细胞迁移的能力，如通过划痕伤口愈合测定所测量的。总之，GPX 3在胃癌中的功能障碍是由遗传和表观遗传改变介导的，表明调节ROS的机制受损，并可能参与胃肿瘤的发生和转移。
Gastric cancer remains the second leading cause of cancer-related death in the world. H. pylori infection, a major risk factor for gastric cancer, generates high levels of reactive oxygen species (ROS). Glutathione peroxidase 3 (GPX3), a plasma GPX member and a major scavenger of ROS, catalyzes the reduction of hydrogen peroxide and lipid peroxides by reduced glutathione. To study the expression and gene regulation of GPX3, we examined GPX3 gene expression in 9 gastric cancer cell lines, 108 primary gastric cancer samples and 45 normal gastric mucosa adjacent to cancers using quantitative real-time RT-PCR. Downregulation or silencing of GPX3 was detected in 8 of 9 cancer cell lines, 83% (90/108) gastric cancers samples, as compared to non-tumor adjacent normal gastric samples (P<0.0001). Examination of GPX3 promoter demonstrated DNA hypermethylation (≥10% methylation level determined by Bisulfite Pyrosequencing) in 6 of 9 cancer cell lines and 60% of gastric cancer samples (P = 0.007). We also detected a significant loss of DNA copy number of GPX3 in gastric cancers (P<0.001). Treatment of SNU1 and MKN28 cells with 5-Aza-2′ Deoxycytidine restored the GPX3 gene expression with a significant demethylation of GPX3 promoter. The downregulation of GPX3 expression and GPX3 promoter hypermethylation were significantly associated with gastric cancer lymph node metastasis (P = 0.018 and P = 0.029, respectively). We also observed downregulation, DNA copy number losses, and promoter hypermethylation of GPX3 in approximately one-third of tumor-adjacent normal gastric tissue samples, suggesting the presence of a field defect in areas near tumor samples. Reconstitution of GPX3 in AGS cells reduced the capacity of cell migration, as measured by scratch wound healing assay. Taken together, the dysfunction of GPX3 in gastric cancer is mediated by genetic and epigenetic alterations, suggesting impairment of mechanisms that regulate ROS and its possible involvement in gastric tumorigenesis and metastasis.
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