【作者】 刘茹；

【导师】 王心如； 王守林；

【作者基本信息】 南京医科大学 ， 劳动卫生与环境卫生， 2004， 硕士

【摘要】 第一部分 氰戊菊酯、辛硫磷对大鼠卵巢功能影响的整体实验研究 研究氰戊菊酯(Fen)、辛硫磷(Pho)对大鼠卵巢内分泌干扰作用及抗氧化系统功能的影响。应用Fen、Pho对雌性SD大鼠进行经口染毒，剂量分别为1.91、9.55、31.80mg／kg和5.88、29.4、98mg／kg，对照组给予等量色拉油。阴道脱落细胞涂片法观察动情周期变化。连续染毒30d后处死，分离卵巢、子宫、脑、肝、脾、肾、肾上腺等脏器，计算脏器系数；分光光度法测定大鼠卵巢组织和血清中的超氧化物歧化酶(SOD)、谷胱甘肽硫转移酶(GST)的活性以及丙二醛(MDA)、谷胱甘肽(GSH)的含量；放射免疫法测定血清中雌二醇 南京医科大学硕士学位论文 (E:)和孕酮(P;)的水平;提取卵巢组织的DNA，进行琼脂糖凝胶电泳，观察DNA完整性;免疫组化SP法观察bc人2蛋白在卵巢组织中的表达;运用光镜和电镜观察卵巢组织和细胞超微结构的变化。 结果显示:Fen各染毒组大鼠的动情前期和动情后期缩短，与对照组相比，在1.91mg瓜g和31.80mg/kg组具有显著性差异;动情期和动情间期则延长，31.80mg/kg组的动情间期持续时间明显比对照组短;各组动情周期异常率均高于对照组，在31 .80m眺g组差异具有显著性。各染毒组大鼠的卵巢、子宫重量均降低，在31.80m眺g组具有显著性差异;卵巢SOD、GST和GSH水平均增高，其中各剂量组GST活力均显著增强，MDA含量降低;血清中SOD、GST水平降低，MDA含量升高;血清中EZ含量显著高于对照组，P4含量则逐渐下降，在9.55mg/kg和31 .80m眺g组与对照组相比差异具有显著性;卵巢bcl一2蛋白表达增强，在9.55m留kg组差异具有显著性。Pho各染毒组大鼠的各动情周期未见明显变化，但动情周期异常率均高于对照组，在98mg瓜g组差异具有显著性;体重增长均低于对照组，其中98mg/kg组与对照组相比差异具有显著性;各染毒组大鼠的卵巢、子宫重量均显著降低;卵巢SOD、GST和GSH水平均增高，MDA含量降低，血清中SOD、GST水平显著降低，MDA含量升高;各剂量组E:含量降低，P;含量升高，但与对照组相比差异均无显著性;卵巢bcl一2蛋白表达增强，在5.88mg瓜g组显著具有显著性。Fen、Pho各剂量组的大鼠卵巢组织DNA完整、无凋亡样断裂。光镜下观察，Fen、Pho各组闭锁卵泡增多，但与对照组相比差异均无显著性。电南京医科大学硕万}学位论文镜下观察:在Fen 31 .80mg/kg组可见卵巢黄体细胞部分内质网肿胀;线粒体呈空泡状，峪消失;核仁边聚。Pho 29.4m留kg组卵巢黄体细胞胞浆内脂滴增大、质不均，峪消失。以上结果提示:增多;在31.核膜不完整;线粒体扩张，出现空泡，基80mg/kg剂量下，Fen对大鼠的动情周期和激素分泌有明显的干扰作用，而Pho的作用不明显。在1.91、9.55、31.somg/kg剂量和5.88、29.4和98mg/kg剂量下，Fen、pho均能使卵巢的抗氧化能力代偿性增强，Pho的作用更为明显，但尚未对卵巢的抗氧化系统功能造成损伤。关健词:氰戊菊醋;辛硫磷;大鼠;卵巢;动情周期;bcl-2第二部分氛戊菊醋、辛硫磷对大鼠卵粱颖拉细胞线拉体功能的影响 研究Fen、Pho对卵巢颗粒细胞线粒体功能的影响，为探讨其内分泌干扰作用机制提供依据。原代培养大鼠卵巢颗粒细胞(rGCS)，分别用Feno、5、25、1 25协mol/L或PhoO、5、25、1 25林mol/L处理细胞24h。MTT试验检测线粒体酶活力;二乙酞二氯氢化荧光素(DCFH一DA)负载，流式细胞术检测细胞内ROS水平;碘布七丙唆(PI)和罗丹明123(Rh123)双标记，流式细胞术检测细胞活力和线粒体膜电位(MMP);生物发光法检测细胞内ATP水平。 结果显示:Fen引起细胞内ROS水平升高，25和1 25月mol几与对照组相比差异具有显著性;PI的荧光强度下降，细胞活力降低，25 南京医科人学硕士学位论文似mol几和125尽mol几组与对照组相比差异具有显著性;细胞损伤率逐渐升高，与对照组比较，25、125拼mol/L组差异具有显著性意义;反映线粒体酶活性的MTT OD值显著升高，5、25、1 25尽mol几组与对照组相比，差异均具有显著性;反映线粒体膜电位的Rh123荧光强度比对照组明显升高;细胞ATP水平呈明显的下降趋势，各剂量组与对照组相比差异均具有显著性。Pho染毒后，细胞内ROS产生增加，其中5林mol几组明显高于对照组;PI荧光强度升高，在125月mol几组与对照组相比差异具有显著性。细胞损伤率逐渐升高，与对照组相比，125林mol几组差异具有显著性。MTT OD值升高，但各剂量组与对照组相比，差异均未见显著性。Rh123荧光强度上升，但与对照组相比，各剂量组差异均未见显著性。与对照组相比，各剂量组的细胞ATP水平随染毒剂量降低，但差异均未见显著性。 以上结果提示:Fen、 Pho能引起大鼠卵巢颗粒细胞的损伤，表现为细胞损伤率增加，ATP水平下降。但Fen并非通过影响线粒体功能而损伤细胞，Fen使细胞活力增强，促进线粒体功能，使线粒体膜电位增加，线粒体酶活性增强;Pho对线粒体功能作用不明显。更多还原

【Abstract】 Part I In Vivo Study on the Effects of Fenvalerate and Phoxim onthe Function of Rat OvaryObjective To investigate the effects of fenvalerate (Fen) and Pho (Pho) on the ovarian antioxidant function.Methods Fen and Pho were administrated ig to the adult female Sprague Dawley rats at dose of 1.91, 9.55, 31.80mg/kg and 5.88, 29.4, 98 mg/kg respectively for one month. Vaginal smears of rats were performed to determine estrous cycle. Some organs such as ovary, uterus, brain, liver,spleen, kidney and adrenal were dissected and organ coefficients were calculated. The activities of superoxide dismutase (SOD) and glutathione-s-transferase (GST), and the levels of malondialdehyde (MDA) and glutathione (GSH) in the ovary and serum were measured by photometry. The estradiol (E) and progesterone (P) in the serum were determined by radioimmunoassay (RIA). Total DNA was extracted from ovary and the DNA fragmentation was valued by agarose gel electrophoresis. The expression of bcl-2 protein in the ovary was observed with SP immunohistochemical method. The morphological changes and ultrastructure of ovary were examined by the light and electronic microscopy.Results In the Fen group, the rate of estrous cylce abnormality significantly increased and the gain of body weight was obviously lower in 31.80mg/kg group than that of control. Proestrus and metestrus prolonged significantly in 1.91mg/kg and 31.80mg/kg group than that of the control and the duration of estrus and diestrus was reduced. The relative organ weights (ovary, uterus) were declined in a dose dependent manner, which was markedly reduced at the dose of 31.80mg/kg. Fen increased the levels of SOD and GSH in ovary and decreased the content of MDA. The levels of GST declined obviously. On the contrary, the SOD and GST activities in the serum of exposed rats were reduced while the content of MDA was enhanced. The synthesis of E was stimulated-6-significantly by Fen in serum at doses of 9.55 and 31.80mg/kg while the level of P was declined obviously. The expression of bcl-2 protein in ovary was up-regulated by Fen in 9.55mg/kg group. Pho increased the rate of estrous cycle abnormality significantly and reduced the gain of body weight obviously in 98mg/kg group than that of control. No any obvious change was observed in the duration of each estrous cycle in the exposed groups. Meanwhile, the ovary and uterus weights were declined in a dose dependent manner. Pho increased the levels of SOD, GST and GSH in ovary and decreased the content of MDA. On the contrary, the SOD and GST activities in the serum of exposed rats had been markedly reduced than that of the control while the contents of MDA was enhanced. No any significant effect of E and P was observed in Pho Treated group. The expression of bcl-2 protein in ovary was increased in low dose group. The morphological changes under the light and electronic microscopy examination indicated that the amount of atrestic follicle in the Fen and Pho exposed groups was increased, but there is no obvious alteration in atrestic follicle of treated group as compared with the control. The ultrastructure changes of ovary under the electronic microscopy indicated that parts of endoplasmic reticulum expanded, mitochondria vacuolized and cristae lost in the Fen group. Nucleolus showed margin assemblling. The morphological alterations in Pho group showed the increase and enlargement of the lipid dropletand the loss of-7-integrity of the nuclear membrane. Mictochondrial swelled and cristaedisappeared, and the matrix showed asymmetry.Conclusion The results suggest that Fen at the doses of 1.91, 9.55, and 31.80mg/kg could significantly disrupt the estrous cycle and secretion of hormone, while Pho at the doses of 5.88, 29.4, 98 mg/kg had not obvious effect as Fen did. Fen and Pho could induce the compensatory increase of antioxidant capacity and the action of Pho was more notable, yet they could not result in a significant damage to the antioxidant system of rat ovary.Key words fenvalerate; phoxim; rat; ovary; estrous更多还原