叶酸对1,3-丁二烯诱发的小鼠DNA低甲基化和染色体损伤的影响

Effect of folic acid on DNA demethylation and chromosomal damage in mice after their inhalation exposure to 1,3-butadiene

目的:探讨叶酸(FA)对1,3-丁二烯(BD)诱发的小鼠遗传损伤和DNA甲基化改变的保护作用。方法:雄性昆明小鼠喂养相应饲料3周建立叶酸正常组(FA-C)、叶酸缺乏组(FA-D)以及叶酸补充组(FA-S)动物模型,每组小鼠18只,再随机分为对照组、BD低剂量和高剂量染毒组(0、13.82、1 382.14 mg/m^3),染毒组小鼠每天吸入染毒6 h,每周5 d,连续染毒2周。采用酶联免疫吸附测定法(ELISA)检测小鼠血清中FA浓度和肝组织基因组DNA总体甲基化水平,荧光定量PCR检测DNA甲基转移酶DNMT1和DNMT3a mRNA的表达,胞质分裂阻滞微核试验分析外周血淋巴细胞的染色体损伤。结果:动物喂养3周后,与FA-C组相比,FA-D组血清FA浓度显著降低,FA-S组显著升高。在相同FA处理条件下,随BD染毒剂量升高,小鼠肝组织基因组DNA甲基化水平及甲基转移酶表达出现降低趋势,微核、核质桥及核芽突比率显著升高。在相同BD染毒条件下,与FA-C组比较,FA-D组的DNA甲基化水平和DNMT1、DNMT3a的mRNA表达出现下降趋势,而微核率等染色体损伤出现升高趋势;相反,FA-S组的DNA甲基化水平及DNMT1、DNMT3a的mRNA表达出现升高趋势,染色体损伤出现降低趋势,尤其在高剂量BD染毒条件下,叶酸补充对上述指标的影响均存在显著性差异(P<0.05或0.01)。结论:BD可诱导小鼠的DNA低甲基化改变及染色体损伤,叶酸缺乏可加重BD所致DNA低甲基化和遗传损伤,而补充叶酸则对BD诱导的甲基化改变和遗传损伤具有保护效应。

OBJECTIVE:To explore the effect of folic acid(FA)on genetic damage and DNA methylation whichwere induced by1,3-butadiene(BD)in mice.METHODS:Male Kunming mice were fed with corresponding diets for3weeks to establish FA-C(normal folic acid group),FA-D(folic acid deficiency group)and FA-S(folic acid supplementation group)animal models.Then,each group of FA-treated mice was randomly divided into control,and BD3low-and high-dose groups(0,13.82,1382.14mg/m)and treated with the corresponding dosages of BD by inhalation for2weeks(6h/d,5d/week).Enzyme-linked immunosorbent assay(ELISA)was applied to detect the concentrations of FA in serum and genomic DNA methylation levels in liver tissue.The expression of methyltransferase DNMT1and DNMT3a was quantified by qPCR and chromosomal damage in peripheral blood lymphocytes was detected by cytokinesis-block micronucleus test(CBMNT).RESULTS:After3weeks of FA treatment,the concentrations of serum folic acid were significantly lower in FA-D group but higher in FA-S group compared with the FA-C group.With increase of the BD doses,methylation of global DNA and expression of methyltransferase were decreased while the frequencies of micronuclei,nucleoplasmic bridge and nuclear bud were increased within the same FA treated group.On the other hand,under the same BD exposure conditions,FA-D group showed a decreased level of DNA methylation and methyltransferase expression and increased chromosomal damage while FA-S group resulted in an increased level of DNA methylation and mRNA level of methyltransferase1,3a expression and decreased micronucleus rate of chromosome compared with FA-C group,especially in the high dose BD exposure conditions(P<0.05or0.01).CONCLUSION:BD can induce global DNA demethylation and chromosomal damage in mice.Folic acid deficiency can aggravate the BDinduced abnormalities.However,folic acid supplementation has a protective effect against the BD-induced abnormalities.