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目的:探讨增塑剂邻苯二甲酸二(2-乙己基)酯(DEHP)作用于怀孕小鼠后,其仔代睾丸基因组DNA甲基化水平的改变情况。方法:妊娠KM小鼠随机分为3组:正常对照组、玉米油对照组和DEHP实验组。自妊娠12.5 d到生后第3天分别持续经口予以玉米油和DEHP[500 mg/(kg.d)],于生后第7天取仔代睾丸,应用甲基敏感扩增多态性(MSAP)技术,对仔鼠睾丸组织DNA进行EcoRⅠ/MspⅠ和EcoRⅠ/HpaⅡ两种酶切反应,经ABI 3730DNA自动测序仪电泳及检测后,运用Genescan3.1分析扩增谱带。结果:对CCGG位点,DEHP实验组小鼠仔代睾丸组织DNA甲基化程度明显增高,其甲基化水平为(34.03±3.05)%;而正常对照组和玉米油对照组小鼠仔代睾丸组织基因组DNA甲基化程度分别为(28.37±2.37)%和(28.58±2.45)%。DEHP实验组与正常对照组和玉米油对照组相比,差异具有统计学意义(P<0.05)。结论:DEHP作用孕鼠后可导致仔代睾丸基因组DNA甲基化水平改变,影响基因组表观遗传修饰改变,可能是导致生殖系统损害的重要毒理机制之一。
Objective: To investigate the changes of methylation level of genomic DNA in the testis of pregnant mice induced by plasticizer di-2-ethylhexyl phthalate (DEHP). Methods: Pregnant KM mice were randomly divided into 3 groups: normal control group, corn oil control group and DEHP experimental group. Corn oil and DEHP [500 mg / (kg · d)] were administered orally from 12.5 d after birth to 3 d after birth respectively, and the testicles were taken on the 7th day after birth. Methyl-sensitive amplification polymorphism (MSAP). The DNA of testis tissue was digested with EcoRⅠ / MspⅠ and EcoRⅠ / HpaⅡ. After electrophoresis and detection by ABI 3730 DNA sequencer, the amplified bands were analyzed by Genescan 3.1. Results: At the CCGG site, the methylation level of testis in DEHP experimental group was significantly higher (34.03 ± 3.05%) than that in normal control group and corn oil control group The degree of methylation of testis tissue DNA was (28.37 ± 2.37)% and (28.58 ± 2.45)%, respectively. Compared with the normal control group and the corn oil control group, the difference between the DEHP experimental group and the control group was statistically significant (P <0.05). CONCLUSION: DEHP induced pregnant rats may result in changes of methylation level of testis genome DNA and influence of epigenetic modification of genome, which may be one of the important toxicological mechanisms leading to reproductive system damage.