Objectives To study whether the abnormal global DNA methylation levels or disturbed promoter methylation pattern of DNA mismatched repair genes increases the incidence of neural tube defects ( NTDs ). Methods A hospital-based case-control study was conducted in the Lvliang area of Shanxi Province, China. NTD-affected cases and controls matched in maternal age and gestational age were recruited from county hospitals and maternal and child health hospitals. All subjects were diagnosed by B-mode ultrasound and confirmed by autopsy. The methylation Quantification Ultra Kits ( Epigentek ) were used to determine the global DNA methylation levels in brain and skin tissues. The Methylation-specific multiplex ligation-dependent probe amplification ( MS-MLPA ) kits were used to quantify the methylation levels of promoter regions of 7 DNA mismatched repair genes ( MLH1, MSH2,MSH6,MSH3,MLH3,PMS2 and MGMT). Results 65 NTD-affected fetuses and 48 normal controls were collected. ① In brain tissue, global DNA methylation levels were significantly decreased in NTD compared with control group ( 5. 3% vs 6. 5% ,P < 0.001 ). DNA hypomethylation caused a significant 4.98-fold increased risk for NTDs ( CI: 1.42 - 17.53 ). No statistical difference was seen for skin tissue in two groups ( 13. 3% vs 13. 1% , P >0. 05 ). ② Compared with control group, NTDs group had significantly lower methylation levels in the promoter regions of MSH6{ MSH6-301 :2. 5% vs 3. 7% , P < 0. 05 )and PMS2{ PMS2-328:5. 7% vs 6. 7% ; PMS2-142-.2. 0% vs 2. 7% , P <0. 05 ). Conclusions Global DNA hypomethylation in fetal brain tissueis associated with NTD-affected pregnancy. Disturbed methylation patterns in promoter regions of two mismatched repair genes, MSH6 and PMS2 are associated with NTD-affected pregnancy.%目的 研究神经管畸形(NTDs)胚胎脑组织和皮肤组织DNA总体甲基化以及错配修复基因启动子区甲基化水平.方法 在山西省吕梁地区以医院为基础进行病例-对照研究.从县级医院及妇幼保健院收集经B超诊断为NTDs的胎儿标本,同时收集非病理性引产、无畸形和发育迟缓的胎儿作为正常对照组.运用甲基化定量试剂盒测定脑组织和皮肤组织的DNA总体甲基化水平,甲基化特异性多连接依赖性探针扩增试剂盒检测7个错配修复基因(MLH1,MSH2,MSH6,MSH3,MLH3,PMS2和MGMT)启动子区甲基化水平.结果 共有65例NTDs胚胎,48例对照胚胎进入研究.①NTDs组的脑组织DNA总体甲基化水平显著低于对照组(5.3% vs 6.5%,P<0.001),DNA总体低甲基化显著增加了NTDs发生风险(OR=4.98,95%CI:1.42~17.53).皮肤组织DNA总体甲基化水平在两组间差异无统计学意义(13.3% vs 13.1%,P>0.05);②NTDs和对照组的MSH6启动子(MSH6-301:2.5% vs 3.7%,P<0.05)和PMS2启动子(PMS2-328:5.7% vs 6.7%;PMS2-142:2.0% vs 2.7%,P<0.05)的甲基化水平存在显著差异.结论 DNA总体低甲基化、错配修复基因启动子区的异常甲基化修饰与NTDs发生有关.
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