N-甲基-N'-硝基-N-亚硝基胍对哈萨克族食管正常上皮细胞甲基转移酶表达及活性的影响  

Effects of MNNG on methyl transferase activity and expression for Kazakh normal esophageal epithelial cells

作　　者：陈丹[1] 黄思语 陈艳[1,2] 

机构地区：[1]新疆医科大学公共卫生学院,新疆乌鲁木齐830011 [2]新疆食管癌研究所

出　　处：《毒理学杂志》2014年第6期424-427,432,共5页Journal of Toxicology

基　　金：国家自然科学基金(81460502)

摘　　要：目的体外实验检测N-甲基-N'-硝基-N-亚硝基胍(MNNG)对哈萨克族食管正常上皮细胞DNA甲基转移酶的表达及活性改变,以此探讨亚硝胺类化合物致食管上皮组织发生恶变的表遗传机制。方法以0、0.75、1.50和3.00μg/ml MNNG处理哈萨克族正常食管上皮细胞24 h,采用高效液相色谱法(HPLC)和DNA甲基转移酶活性检测试剂盒检测细胞基因组DNA整体甲基化水平和DNA甲基转移酶活性的改变;并采用RT-PCR和Western Blot法检测DNA甲基转移酶mRNA及蛋白表达。结果 1.50和3.00μg/ml MNNG组与对照组比较细胞基因组DNA整体甲基化水平分别降低8.07%和17.58%,且各染毒组之间比较差异有统计学意义(F=60.342,P<0.01);与对照组比较各染毒组DNMT1、DNMT3a、DNMT3b酶活性均升高,且与MNNG浓度呈正相关(r=0.967,P<0.01;r=0.897,P<0.01;r=0.716,P<0.01);与对照组比较各染毒组DNMT1、DNMT3a、DNMT3b基因mRNA及蛋白表达水平均升高,且与MNNG浓度呈正相关(r=0.814,P<0.05;r=0.732,P<0.01;r=0.578,P<0.05;r=0.944,P<0.01;r=0.900,P<0.01;r=0.887,P<0.01)。结论 MNNG可致哈族正常食管上皮细胞基因组DNA整体甲基化水平下降,提高DNMT1、DNMT3a、DNMT3b表达及活性。Objective In vitro to test the changes of MNNG on DNA methyhransferase （DNMTs） expression and activity for Kazakh normal esophageal epithelial ceils ,to investigate the nitrosamines compounds-induced maligrant transsformation of Kazakh normal esophageal epithelial tissue by epigenetic mechanisms. Methods Kazakh normal esophageal epithelial cells were treated with MNNG at 0,0.75, 1.5 and 3 μg/ml for 24 h. Highperformance liquid chromatography（HPLC） and DNMTs activity assay kit were applied to detect the change of global DNA methylation level and DNMTs activity. The mRAN and protein expressions of DNMTs were determined by RT-PCR and Western Blot. Results Comparing the control group with the MNNG exposed group of 1.50 and 3.00 μg/ml for 24 h, cell genome DNA methylation level depression rate were 8.07% and 17.58% respectively. The activities of DNMTs in MNNG groups were significantly higher than in control group （P 〈0. 01 ）,and was related positively with MNNG concentration （P 〈0. 01 ）. The mRAN and protein expressions of DNMTs in MNNG groups were significantly higher than in control group （P 〈 O. 05 ）, and was related positively with MNNG concentration （P 〈 0.01）. Conclusion MNNG significantly depresses the global DNA methylation level and improves methyl transferase activity and expression of Kazakh normal esophageal epithelial cells.

关 键 词：正常食管上皮细胞 甲基硝基亚硝基胍 哈萨克族 DNA甲基转移酶 

分 类 号：R735.1[医药卫生—肿瘤]