环境内分泌干扰物联合作用对大鼠生殖及子代发育的影响及代谢组学研究

Environmental endocrine disruptors（EEDs）mainly be divided into three category: (1) nature estrogen, such as estrone, estladiol(E2) and estriol(E3); (2) phytoestrogen, such as isoflavone and coumestrol and so on; (3) synthesis estrogen-like compounds, such as pesticide, plasticiser and many other industry products. Synthesis estrogen-like compounds, exist in the environment with the features of wide source, variety and low concentration, and they can enter body through multiple pass and influence health after many years. Many researchers have paid more attention on this problem and got a great achievement. But they usually do the research with 1 kinds EEDs, less report is about the joint effect of EEDs, and even less report is about the joint effect's and metabonomics research. In this study, 3 different category EEDs, such as estradiol（E2）, genstein and di(2-ethylhexyl)phthalate (DEHP) and their mixture will be used to study whether they exist joint effects and what's the biomarkers and the mechanism. Estradiol has the strongest estrogen activity, genstein as a main active component of isoflavone and a selective estrogen receptor modulator largely exist in soybean food, and DEHP usually be used in plastics and easily transference into food through food packing. In the cell culture research, differsnt concentration and different proportin estradiol, genstein , DEHP and their mixture will add to the medium of human mammary cancer cell(MCF-7), and we can get the data of estrogen activity and effective dosage in each other to evaluate the extent of their function. Through 3 stage animal experiment ( 3 kinds EEDs or mixture given to the adult rats by intragastric administration, and continute to the pregnant rats and first filial generation delactation rats) , we will know the joint effect of this 3 EEDs on reproduction, embryon and offspring development. Further through metabonomics analysis and gene or protein expression detection, we will find some biomarkers of 3 EEDs, the change of metabolite by EEDs mixture and reveal the possible mechanism. .In a word, this research will help people understand the joint effects of 3 difference source EEDs, their influence on the reproductive system of adult rats, pregnant rats and the development of fetal rats and first filial generation delactation rats and further reveal their influence on biomarker and possible mechanism.

环境内分泌干扰物（EDCs）的存在特点是来源广、种类多、浓度低，对人体健康产生的影响具有潜伏性。目前在EDCs研究领域进行多种混合物相互作用的研究较少，而从体内、体外两方面同时开展3种不同类别环境内分泌干扰物（DEHP、雌二醇与大豆异黄酮）的相互作用及代谢组学研究的文献还未见报道。本课题选择目前环境污染范围广、雌激素活性强、人体暴露量大的3种不同来源、具有不同生理学特性的环境内分泌干扰物DEHP、雌二醇与大豆异黄酮主要活性成分genstein的混合物进行联合作用研究。利用细胞培养技术了解其雌激素活性相互作用的情况及作用剂量；同时通过多个动物实验系统研究它们的生殖毒性作用；利用代谢组学方法对代谢产物进行定性定量分析，确定这3种环境内分泌干扰物在机体内的生物标志物及相互作用的效果。该研究项目将使我国在环境内分泌干扰物的研究方面有新的突破，同时为综合评价环境内分泌干扰物对人体健康的影响提供依据。

目的：了解3 种不同的环境内分泌干扰物（DEHP、雌二醇与大豆异黄酮）单独及其联合作用对不同性别大鼠的影响。.方法：选用刚断乳的SD大鼠（雌、雄各半），按照体重进行随机分为8组，给予一定种类的受试物，喂养4周。在实验结束前3天，收集大鼠24小时尿液，冷冻保存，用于代谢组学分析。处死大鼠，取脏器组织进行称重，分离血清测定雌二醇、睾酮等激素及DEHP的水平，子宫及睾丸进行病理及免疫组化检测。.结果：研究发现（1）给定剂量的3种受试物对大鼠体重的增加没有显著影响。（2）DEHP对大鼠肝脏的脏体比有显著影响（P<0.05），SIF和E2可减轻DEHP对雄性大鼠肝脏的影响。（3）给予DEHP的大鼠血清中DEHP浓度显著高于其他各组，而SIF和E2能够降低血清中DEHP的浓度。（4）3种受试物对雌性和雄性各组大鼠血清中的雌激素E2和雄激素睾酮浓度没有显著性影响，但剂量组的E2水平均低于对照组。（5）各组大鼠血清中T3浓度没有显著差异；单纯给予SIF和DEHP可使雌性大鼠血清中TSH浓度增加，E2对其浓度没有影响；3种受试物联合作用使雄性大鼠组TSH显著高于对照组及2种受试物的联合作用的各组。（6）3种受试物联合可使雌性大鼠子宫的雌激素受体蛋白表达增加、子宫内膜增生。在给定剂量下对雄性大鼠睾丸没有影响。.结论：SIF和雌二醇可降低体内DEHP在水平，3种受试物联合作用对雌性大鼠具有较大影响。