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目的探讨围产期高脂喂食对雄性子代大鼠肠系膜动脉二甲基精氨酸二甲胺水解酶2(DDAH2)启动子甲基化水平的影响及在动脉血压增高中的作用。方法 SD大鼠分为正常脂肪喂食组(NFD)和高脂喂食组(HFD),围产期分别给予标准饲料(含4.43%脂肪)和高脂饲料(含30%脂肪)喂养,分娩后各组随机选择9只雄性子鼠以标准饲料喂养,18周龄时采用无创性尾套法测量子鼠动脉血压水平,检测肠系膜动脉内皮依赖性舒张功能,采用硝酸还原酶法检测肠系膜动脉一氧化氮含量,酶联免疫吸附试验法检测非对称性二甲基精氨酸(ADMA)含量及内皮型一氧化氮合酶(eNOS)活性,实时荧光定量聚合酶链反应(PCR)检测DDAH2mRNA表达水平,Western-blot方法检测DDAH2蛋白表达水平,巢式甲基化PCR法检测DDAH2启动子甲基化水平。结果与NFD组相比,HFD组子代大鼠收缩压升高[(143.3±2.6)比(121.7±3.8)mm Hg,P<0.05];肠系膜动脉血管环的舒张百分比降低(均P<0.05),敏感性降低(pD2:NFD 6.44±0.18比HFD 7.56±0.26,P<0.05);肠系膜动脉一氧化氮含量降低[(0.42±0.01)比(0.53±0.01)μmol/g,P<0.01],eNOS活性降低[(0.57±0.01)比(0.73±0.01)kU/g pro,P<0.01],ADMA含量增加[(0.32±0.07)比(0.29±0.01)μmol/g pro,P<0.05];肠系膜动脉DDAH2mRNA降低(0.12±0.01比0.15±0.01,P<0.01),DDAH2蛋白降低(0.24±0.01比0.28±0.01,P<0.01),启动子甲基化水平升高(1.20±0.03比0.98±0.04,P<0.01)。结论围产期高脂喂食可以导致雄性子代大鼠肠系膜动脉DDAH2启动子甲基化水平增高,引起DDAH2/ADMA/eNOS/一氧化氮通路障碍,是其成年后动脉血压升高的原因之一。
Objective To investigate the effect of perinatal high-fat diet on the methylation of dimethylarginine dimethylamine hydrolase 2 (DDAH2) promoter in mesenteric artery of male offspring rats and its role in arterial hypertension. Methods SD rats were divided into normal fat feeding group (HFD) and normal fat feeding group (HFD). The perinatal period was fed with standard diet (containing 4.43% fat) and high fat diet (containing 30% fat) Nine male rats were randomly selected to receive standard diet. At 18 weeks of age, the arterial blood pressure was measured by noninvasive tail cuff method. The endothelium-dependent vasodilatation of mesenteric artery was detected. Nitric acid reductase (ADMA) and endothelial nitric oxide synthase (eNOS) activity were detected by enzyme-linked immunosorbent assay (ELISA). The expression of DDAH2 mRNA was detected by real-time fluorescence quantitative polymerase chain reaction (PCR) The expression of DDAH2 protein was detected by Western-blot and the methylation level of DDAH2 promoter was detected by nested methylation PCR. Results Compared with NFD group, the systolic blood pressure of offspring in HFD group was significantly higher than that of NFD group [(143.3 ± 2.6) vs (121.7 ± 3.8) mm Hg, P <0.05] ), The sensitivity decreased (pD2: NFD 6.44 ± 0.18 vs HFD 7.56 ± 0.26, P <0.05); the content of nitric oxide in the mesenteric artery decreased (0.42 ± 0.01 vs 0.53 ± 0.01 μmol / g, P 0.01) (0.72 ± 0.07), (0.29 ± 0.01) μmol / g pro, P <0.05], and decreased the activity of eNOS [(0.57 ± 0.01) vs 0.73 ± 0.01 kU / g pro (0.12 ± 0.01 vs 0.15 ± 0.01, P <0.01), DDAH2 protein decreased (0.24 ± 0.01 vs 0.28 ± 0.01, P <0.01), and the level of promoter methylation was higher in the mesenteric artery (1.20 ± 0.03 vs 0.98 ± 0.04, P <0.01). Conclusions High-fat diet during perinatal period may lead to increased methylation level of DDAH2 promoter in mesenteric artery of male offspring and cause DDAH2 / ADMA / eNOS / NO pathway dysfunction, which is one of the causes of arterial blood pressure increase after adulthood .