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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 investigate the protective effect of folic acid (FA) on genetic damage and DNA methylation induced by 1,3-butadiene (BD) in mice. Methods: Male Kunming mice were fed with the corresponding diets for 3 weeks to establish animal models of folic acid deficiency (FA-C), folic acid deficiency (FA-D) and folic acid supplementation (FA-S) The mice were divided into control group, BD low dose and high dose exposure groups (0, 13.82, 1 382.14 mg / m ~ 3). The mice in the exposure group were exposed to inhalation for 6 hours daily for 5 days, . The serum concentrations of FA and total genomic DNA were detected by enzyme-linked immunosorbent assay (ELISA). The DNMT1 and DNMT3a mRNA expressions were detected by real-time quantitative PCR. The cytokinesis Nuclear test analysis of peripheral blood lymphocyte chromosome damage. Results: Compared with FA-C group, FA concentration in FA-D group was significantly decreased and FA-S group was significantly increased after 3 weeks of feeding. Under the same conditions of FA treatment, the DNA methylation level and the expression of methyltransferase in liver tissue of mice decreased with the increase of BD dose, and the ratios of micronuclei, nucleus pontis and nuclear buds were significantly increased. Under the same conditions of BD exposure, compared with FA-C group, the DNA methylation level and DNMT1, DNMT3a mRNA expression in FA-D group showed a downward trend, and the chromosome damage such as micronucleus rate showed an upward trend; on the contrary, DNA methylation level and the mRNA expression of DNMT1 and DNMT3a in FA-S group tended to increase, and chromosome damage tended to decrease. Especially in high-dose BD exposure conditions, the effects of folic acid supplementation on these indexes were significantly different (P <0.05 or 0.01). CONCLUSION: BD can induce DNA hypomethylation and chromosomal damage in mice. Folic acid deficiency aggravates DNA hypomethylation and genetic damage induced by BD, whereas folic acid supplementation has the effect on BD-induced methylation and genetic damage Protection effect.