Key Laboratory of Environmental Medicine Engineering, Ministry of Education, School of Public Health, Southeast University, Nanjing, China.
DNA Cell Biol. 2012 May;31(5):856-66. doi: 10.1089/dna.2011.1318. Epub 2011 Dec 23.
The abnormal function of O(6)-methylguanine-DNA methyltransferase (MGMT) is reported to be associated with the occurrence of various tumors and malignant tumor progression. However, little evidence is available to describe its role in esophageal carcinogenesis. To address this issue, we constructed a stable MGMT-silenced esophageal cancer cell line by RNA interference, and exposed the cells to N-methyl-N-nitro-N-nitrosoguanidine (MNNG) to investigate the role that MGMT plays in toxicity. During this time, we also observed the malignant behavior of cells in vitro and in vivo. In addition, two-dimensional electrophoresis and mass spectrometry were used to detect and confirm the proteins that were differentially expressed in the MGMT-deficient and MGMT-proficient cells, which might be responsible for the malignant alteration of cells. Results showed that the IC(50) of MGMT-deficient and MGMT-proficient cells exposed to MNNG was 30 μM and 65 μM, respectively, and MGMT-deficient cells had more aggressive motility and invasive abilities compared with MGMT-proficient cells. Nineteen differentially expressed proteins were detected between the MGMT-deficient and MGMT-proficient cells, 14 of which were identified, including the membrane-cytoskeleton linker protein, Ezrin, which was confirmed by both mass spectrometry and western blot analysis. The correlation between MGMT, Ezrin expression, and the malignant behavior of one normal epithelial esophageal cell line and seven esophageal cancer lines is discussed. In conclusion, loss of MGMT expression leads EC109 esophageal cancer cells to have increased malignant behavior, which may correlate with its high Ezrin protein expression.
O(6)-甲基鸟嘌呤-DNA 甲基转移酶（MGMT）的异常功能与多种肿瘤的发生和恶性肿瘤的进展有关。然而，目前几乎没有证据表明它在食管癌发生中的作用。为解决这一问题，我们通过 RNA 干扰构建了稳定的 MGMT 沉默食管癌细胞系，并将细胞暴露于 N-甲基-N-亚硝基-N-亚硝基胍（MNNG）中，以研究 MGMT 在毒性中的作用。在此期间，我们还观察了细胞在体外和体内的恶性行为。此外，我们还使用二维电泳和质谱技术来检测和确认在 MGMT 缺陷和 MGMT 功能正常的细胞中差异表达的蛋白质，这些蛋白质可能是导致细胞恶性改变的原因。结果表明，MGMT 缺陷和 MGMT 功能正常的细胞暴露于 MNNG 后的 IC（50）分别为 30μM 和 65μM，与 MGMT 功能正常的细胞相比，MGMT 缺陷的细胞具有更强的运动性和侵袭能力。在 MGMT 缺陷和 MGMT 功能正常的细胞之间检测到 19 种差异表达的蛋白质，其中 14 种已被鉴定，包括膜细胞骨架连接蛋白 Ezrin，这一点通过质谱和 Western blot 分析得到了证实。讨论了 MGMT、Ezrin 表达与一种正常食管上皮细胞系和七种食管癌系恶性行为之间的相关性。结论是，MGMT 表达缺失导致 EC109 食管癌细胞恶性行为增加，这可能与其 Ezrin 蛋白高表达有关。
