青春期前邻苯二甲酸二(2-乙基)己酯暴露对雌性大鼠生殖发育及过氧化物酶体增殖剂激活受体的影响

Effects of di-(2-ethylhexyl) phthalate exposure on reproductive development and PPARs in prepubertal female rats

目的探讨邻苯二甲酸二(2-乙基)己酯(DEHP)暴露对青春期前雌性大鼠性发育及生殖内分泌功能的影响及其可能机制。方法将40只健康3周龄雌性SD大鼠,随机分成对照组(玉米油)和3个实验组,每组10只。实验组按50、150、500mg/kg DEHP经口灌胃,连续染毒28天。观察阴道开口、乳房发育、第一次发情周期的日龄及体重,于末次染毒24小时后进行阴道涂片,确定动情间期,处死动物。Real-time PCR测定卵巢组织相关基因表达水平;ELISA法检测血清中卵泡刺激素(FSH)、黄体生成素(LH)、雌二醇(E2)、孕酮(P4)及睾酮(T)水平;通过病理学观察卵巢组织的变化;免疫组织化学法测定卵巢组织中PPARγ的表达。结果 500mg/kg组阴道开口日龄提前,150、500mg/kg组阴道开口时体重增加(P<0.05);150、500mg/kg组芳香化酶(P450Arom)mRNA表达与对照组比较,显著下降;各剂量组与对照组相比,T水平明显下降,150、500mg/kg组FSH、E2水平明显减少,而LH水平明显升高(P<0.05);150、500mg/kg组闭锁卵泡明显增多、黄体数目明显减少;150、500mg/kg组卵泡颗粒细胞和黄体颗粒细胞中PPARγ阳性光密度相对量明显高于对照组及50mg/kg组(P<0.05)。结论青春期前DEHP暴露对雌性大鼠性发育及生殖内分泌功能产生影响,其作用机制可能与激活PPARs有关。

Objective To investigate the effects of di-（2-ethylhexyl） phthalate（DEHP） exposure on pubertal development and reproductive endocrine function in prepubertal female rats and its possible mechanisms.Methods 40 female SD rats at 3-week-old were randomly divided into a control group（corn oil） and three exposure groups,which were exposed consecutively for 28 days to DEHP by oral gavage at doses of 50,150 and 500 mg/（kg·d）.The onset of puberty was determined by daily examination for vaginal opening（OV）,breast development and the first estrous cycle.By the end of the experiment,the rats were sacrificed during the diestrous stage.The gene expression interrelated with ovary function was detected by real-time PCR.Serum levels of follicle stimulating hormone（FSH）,luteinizing hormone（LH）,estradiol（E2）,progesterone（P4） and testosterone（T） were measured by ELISA.The morphological change of ovaries was observed by HE staining under optical microscope.The expression of PPARγ protein in ovary cells was measured by immuohistochemistry.Results The age of vaginal opening was advanced by DEHP exposure in 500 mg/ kg group,and the body weight was increased at the time of vaginal opening in 150 and 500mg/kg groups（P0.05）.Compared to the control group,the level of T in all DEHP groups was decreased;the levels of FSH and E2 were decreased and the level of P4 was increased in 150 and 500mg/kg groups（P0.05）.The gene expression of P450Aromin 150 and 500mg/kg groups was significantly decreased compared to the control group.A significant increase of atretic follicles and a significant reduction of corpora lutea were observed in 150 and 500mg/kg groups compared to the control group.The expression of PPARγ protein in granulosa cells of follicle and corpora lutea in 150 and 500mg/kg groups was higher than that in the control group and 50mg/kg group（P0.05）.Conclusion DEHP exposure can affect the pubertal development and reproductive endocrine function in prepubertal female rats,and its possible mechanism may be correlated with PPARγ.