亚硝酸盐暴露小鼠不孕效应的实验性研究

Investigation on Mouse Infertility after Nitrite Exposure

对66只雌性且有阴栓的C57BL/6J小鼠(对照组)每日进行生理盐水灌胃处理,待幼仔娩出,统计不孕率.将对照组哺乳后且有阴栓的雌鼠随机平均分配,分别进行每天60mg/kg(低剂量亚硝酸盐组)和120mg/kg(高剂量亚硝酸盐组)的亚硝酸钠灌胃处理,待子鼠娩出,统计不孕率(第一轮暴露模型).按此方法,对第一轮暴露哺乳后且有阴栓的雌鼠进行第二轮亚硝酸钠暴露并统计不孕率.收集对照组和高剂量亚硝酸盐两轮暴露组雌鼠的卵巢,采用HE TUNEL试剂盒观察卵泡内颗粒细胞凋亡情况及诱导型一氧化氮合酶(iNOS)表达情况,利用Western blot蛋白半定量分析法分析Caspase 8、c-fos蛋白在卵巢内的相对表达量.结果显示:亚硝酸盐暴露可致小鼠不孕且具有剂量依赖性(P<0.05,n=66);亚硝酸盐暴露可诱导机体产生氧化应激反应、可使卵泡内颗粒细胞iNOS表达增加、颗粒细胞凋亡增加(P<0.01,n=20).以上结果说明,亚硝酸盐有致不孕效应,可能与NO信号通路和氧化应激诱导的颗粒细胞凋亡和排卵抑制有关.

C57BL/6J female mice with vaginal plug（control group） were treated by physiological saline daily, and the infertility rate was detected after newborn mouse delivery. The lactating female mice with vaginal plug in control group were divided into two groups. The low dose nitrite group were given 60 mg/kg sodium nitrite and the high dose nitrite group were given 120 mg/kg sodium nitrite daily, and the infertility rates of the fist round were detected after newborn mouse delivery. The lactating female mice with vaginal plug of the first round were given the same treatment and the infertility rates were also calculated by this method. The mouse＇ ovaries of the control group and the high dose nitrite group were collected. HE TUNEL kit was used to observe the apoptosis of granulosa cells and the expression of inducible nitric oxide synthase （iNOS）. The relative expressions of Caspase 8 and c fos protein in ovary were analyzed by blot Western protein semi quantitative method. Results display： Nitrite exposure can cause mouse infertility with dose-dependent （P〈0.05, n=66）. Nitrite exposure can induce oxidative stress, increase the expression of iNOS in granulosa cells and promote the granulosa cells apoptosis （P〈0.01, n=20）. These results indicate nitrite can cause infertility, which may be caused by NO signaling pathway and oxidative stress.