苯对大鼠的生殖毒性和胚胎发育毒性研究
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摘要
目的:
探讨苯对大鼠的生殖毒性和胚胎发育毒性及其作用机制,为全面评价苯的生殖及胚胎发育毒性提供实验依据。
方法:
清洁级SD大鼠雄性36只,雌性48只,雌、雄性大鼠分别随机分为4组,各组分别吸入0、5、10和15mg/m~3的苯蒸气染毒。雄鼠先连续染毒7d,每天2hr,从雄鼠染毒后的第28d丌始,雌、雄鼠连续染毒7d、每天2hr。染毒完成后的第2天,各组大鼠取6只处死,雄性大鼠检测血清睾酮含量、睾丸和附睾脏器系数、精子畸形;雌性大鼠检测血清促卵泡生成激素、促黄体生成激素、雌二醇的含量及卵巢黄体数、卵巢脏器系数;睾丸进行常规病理学观察。染毒完的第二天下午18:00时,每组余下的3只雄性与6只雌性按1:2的比例合笼过夜,次日晨8:00时检查雌鼠阴栓,统计交配率。发现阴栓的当天早上8:00为妊娠第0d。各组孕鼠在孕6d一15d再次吸入苯染毒,于孕18d处死孕鼠,观察胚胎的生长发育状况、死胎、吸收胎及畸型胎、胎盘重量;在实验期间观察大鼠中毒情况,雌雄鼠每周称体重1次,孕鼠每天观察孕鼠体重变化。
结果:
1.染毒期间各组大鼠的毛发光泽性差、有竖毛现象,活动、食量减少。各组孕鼠在孕0—6d其体重增长率无差别(P>O.05);苯染毒后,第一个3d即孕7—9d,各组孕鼠的体重增长率差异无显著性,但在染毒后的第二个3d,即在孕10—12d,高剂量苯染毒组的体重增长率低于对照组(P 2.各组雄鼠的睾丸和附睾重量随染毒剂量的增加呈降低趋势,但脏器系数与对照组比较均无显著性差异(P>O.05);而且染毒组雌鼠的卵巢脏器系数与对照组比较也无显著性差异(P>0.05)。
3.对照组大鼠的精子畸形率为(1.90±0.14)%,低、中、高苯染毒组的精子畸形率分别为(2.18±0.34)%、(2.83±O.26)%、(4.20±0.21)%,4组精子畸形率比较差异均有统计学意义(F=182.35,P<0.01),精子主要畸形有:大头、小头、香蕉型头、勾型头、双头、三头、颈弯曲、颈扭转、体弯曲、尾折叠、短尾等。
4.睾丸病理组织学观察显示:对照组曲细精管形态规则,边缘完整,管壁内精原母细胞、初级精母细胞和次级精母细胞由外至内排列紧密,层次分明清晰,管腔位于中央处,腔内可见大量成熟的精细胞。染毒组可见曲细精管形态不规则,边缘极不完整,大小不一,管壁内的细胞由外至内排列不紧密,精原母细胞、初级精母细胞和次级精母细胞有些变性,甚至缺失,层次不清晰。染毒浓度越高,病理损害越明显。
5.中、高浓度苯染毒组雄鼠血清中睾酮水平低于对照组(P<0.05);中、高浓度苯染毒组母鼠血清中FSH、LH、E2分泌水平低于对照组(P0.05)。中、高浓度苯染毒组胎盘重量低于对照组(P 6.各染毒组大鼠交配受孕率差异无统计学意义(P>O.05),中、高浓度苯能增加胚胎的吸收、死亡率(P 结论:
1.苯具有生殖毒性:可导致雄性大鼠激素水平改变,精子畸形,睾丸组织病理改变;雌性大鼠激素水平改变。
2.苯具有胚胎发育毒性:在孕鼠致畸敏感期接触苯,可导致妊娠结局发生改变,使胚胎的生长发育延迟甚至死亡。
Objective:
To evaluate toxic effects of benzene on reproductive and embryonic development, The reproductive and embryonic developmenalt toxicity were assessed by exposing rats to the different levels of benzene.
Methods:
The SD rats (clean grade) including both male 36 and female 48, were randomly divided into four groups: placebo group, benzene vapor (BV) 5 mg/m~3 group, BV 10 mg/m~3 group and BV 15 mg/m~3 group. Firstly, the male rats continuous exposed to BV for 7 daily 2hr. Secondly, both male and female rats continuous exposed to BV for 7 daily 2hr when the males exposured to the first 28d. Thirdly, six rats from each group were sacrificed to examine for the different index at the 2d after all exposured to BV, which included not only the serum testosterone levels, the epididymal viscera coefficient, the testicular viscera coefficient and sperm abnormalities of males, but also the serum follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol levels (EL), ovarian corpus luteum (CL) and viscera coefficient of females. At the same time, the pathological observation on the testis had been also carried out. Finally, all the remaining female rats were co-caged overnight with the remaining male ones at 18:00 pm on the 2d after all exposured to BV, and the vaginal plugs of female rats and the mating rates were examined respectively at 8:00 on the next day. Afterwards, the fecundability and abortion rates of the female rats were observed during 5d, which the time we found the vagina! plugs was regarded as the "zero" day for the gestation. Next, the pregnant rats from each group were treated by BV again from the 6d to 15d. In addition, throughout the experiment period, not only all of the rat and the pregnant ones weights were investigated once weekly and every day respectively, but also the BV poisoning of rat were observed. After a short time (pregnant at 18d), the pregnant rats were sacrificed to investigate the growth and development status of embryos, fetal deaths, fetus resorptions, featus anomalies and placental weights.
Results:
1. The rats that exposed to BV had less action, less food intake, and bristling hair with poor gloss. There were no significant difference in growth rates for pregnant rats among 0 and 6d (P>0.05). After the pregnant rats from each group exposed to BV, their growth rates among 7 and 9d (pregnant 0-3d) showed no significant difference (P <0.05). However, growth rates of pregnant rats from BV 15 mg/m~3 group were lower than control group among 10 - 12d (P<0.05); in addition, growth rates from BV 10 mg/m~3 and BV 15 mg/m~3 group showed significant difference (P <0.05) bewteen 13 -15d and l6-18d.
2. The male rat' epididymal and testicular weights from each group decreased with the increase of BV dose, but their epididymal and testicular viscera coefficient were lower than control group (P<0.05). Futhermore, The female rat’ovarian viscera coefficients from each group were lower than control group (P<0.05).
3. Deformity rates of 1.90 %±0.14 %, 2.18 %±0.34 %, 2.83 %±0.26 %, 4.20 %±0.21 % had been obtained from the rats sperm of control, BV 5 mg/m~3, BV 10 mg/m~3 and BV 15 mg/m~3 group, respectively. Furthermore, deformity rates among the four group were statistically significant (F=182.35, P<0.01), which mainly include Big Head, Small head, banana head, hook-type head, double, three, neck bending, neck torsion, body bending, folding tail, short tail and so on.
4. Histopathologic studies showed that the morphous and edges of the seminiferous tubule from control rat testes remained regular and intact. Moreover, in the inner wall of seminiferous, the tubule spermatogoniums, primary and secondary spermatocyte were densely clustered together from external to internal directions with clear hiberarchy, and a lot of mature sperm cells could also be seen in the lumen round the middle of seminiferous. However, the tubule from exposed rat testes differed from this pattern. Such as, irregular morphous, quite incomplete edges, different sizes, porous arrangement of cells in the inner wall, degeneration and even loss of spermatogoniums, primary and secondary spermatocyte. In brief, the higher the dose was, the more obvious the pathological damage was.
5. The levels of testosterone in serum from exposed rat testes were lower than that from control ones (P<0.05). Though the levels of FSH、LH、E2 in serum from BV 5 mg/m~3 group were no significant difference from control ones (P>0.05), that from BV 10 mg/m~3 and BV 15 mg/m~3 group were lower than that from control ones (F<0.05).Likewise, the placental weights of rats from BV 10 mg/m~3 and BV 15 mg/m~3 group were lower than that from control ones(P<0.05).
6. The occurrence probability of fetal deaths and fetus resorptions from BV 10 mg/m and BV 15 mg/m group could be added (P<0.05) though the pregnancy rates of rats from each group were no significant difference (P>0.05). The weights, body and tail length of the fetuses in the rats from BV 10 mg/m~3 and BV 15 mg/m~3 group were lower than that from control ones (P<0.05). Therefore, Benzene can inhibit the growth of fetal rats in vivo with a dose - response relationship.
Conclusions:
1. Benzene, responsible for reproductive toxicity of rat, can induce change in some index, such as hormone levels, sperm morphology, testis tissue of male rat, and hormone levels of female rat.
2. Benzene, also responsible for embryonic development toxicity, can result in adverse pregnancy outcomes, growth and development delay of fetus and even death.
探讨苯对大鼠的生殖毒性和胚胎发育毒性及其作用机制,为全面评价苯的生殖及胚胎发育毒性提供实验依据。
方法:
清洁级SD大鼠雄性36只,雌性48只,雌、雄性大鼠分别随机分为4组,各组分别吸入0、5、10和15mg/m~3的苯蒸气染毒。雄鼠先连续染毒7d,每天2hr,从雄鼠染毒后的第28d丌始,雌、雄鼠连续染毒7d、每天2hr。染毒完成后的第2天,各组大鼠取6只处死,雄性大鼠检测血清睾酮含量、睾丸和附睾脏器系数、精子畸形;雌性大鼠检测血清促卵泡生成激素、促黄体生成激素、雌二醇的含量及卵巢黄体数、卵巢脏器系数;睾丸进行常规病理学观察。染毒完的第二天下午18:00时,每组余下的3只雄性与6只雌性按1:2的比例合笼过夜,次日晨8:00时检查雌鼠阴栓,统计交配率。发现阴栓的当天早上8:00为妊娠第0d。各组孕鼠在孕6d一15d再次吸入苯染毒,于孕18d处死孕鼠,观察胚胎的生长发育状况、死胎、吸收胎及畸型胎、胎盘重量;在实验期间观察大鼠中毒情况,雌雄鼠每周称体重1次,孕鼠每天观察孕鼠体重变化。
结果:
1.染毒期间各组大鼠的毛发光泽性差、有竖毛现象,活动、食量减少。各组孕鼠在孕0—6d其体重增长率无差别(P>O.05);苯染毒后,第一个3d即孕7—9d,各组孕鼠的体重增长率差异无显著性,但在染毒后的第二个3d,即在孕10—12d,高剂量苯染毒组的体重增长率低于对照组(P
3.对照组大鼠的精子畸形率为(1.90±0.14)%,低、中、高苯染毒组的精子畸形率分别为(2.18±0.34)%、(2.83±O.26)%、(4.20±0.21)%,4组精子畸形率比较差异均有统计学意义(F=182.35,P<0.01),精子主要畸形有:大头、小头、香蕉型头、勾型头、双头、三头、颈弯曲、颈扭转、体弯曲、尾折叠、短尾等。
4.睾丸病理组织学观察显示:对照组曲细精管形态规则,边缘完整,管壁内精原母细胞、初级精母细胞和次级精母细胞由外至内排列紧密,层次分明清晰,管腔位于中央处,腔内可见大量成熟的精细胞。染毒组可见曲细精管形态不规则,边缘极不完整,大小不一,管壁内的细胞由外至内排列不紧密,精原母细胞、初级精母细胞和次级精母细胞有些变性,甚至缺失,层次不清晰。染毒浓度越高,病理损害越明显。
5.中、高浓度苯染毒组雄鼠血清中睾酮水平低于对照组(P<0.05);中、高浓度苯染毒组母鼠血清中FSH、LH、E2分泌水平低于对照组(P
1.苯具有生殖毒性:可导致雄性大鼠激素水平改变,精子畸形,睾丸组织病理改变;雌性大鼠激素水平改变。
2.苯具有胚胎发育毒性:在孕鼠致畸敏感期接触苯,可导致妊娠结局发生改变,使胚胎的生长发育延迟甚至死亡。
Objective:
To evaluate toxic effects of benzene on reproductive and embryonic development, The reproductive and embryonic developmenalt toxicity were assessed by exposing rats to the different levels of benzene.
Methods:
The SD rats (clean grade) including both male 36 and female 48, were randomly divided into four groups: placebo group, benzene vapor (BV) 5 mg/m~3 group, BV 10 mg/m~3 group and BV 15 mg/m~3 group. Firstly, the male rats continuous exposed to BV for 7 daily 2hr. Secondly, both male and female rats continuous exposed to BV for 7 daily 2hr when the males exposured to the first 28d. Thirdly, six rats from each group were sacrificed to examine for the different index at the 2d after all exposured to BV, which included not only the serum testosterone levels, the epididymal viscera coefficient, the testicular viscera coefficient and sperm abnormalities of males, but also the serum follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol levels (EL), ovarian corpus luteum (CL) and viscera coefficient of females. At the same time, the pathological observation on the testis had been also carried out. Finally, all the remaining female rats were co-caged overnight with the remaining male ones at 18:00 pm on the 2d after all exposured to BV, and the vaginal plugs of female rats and the mating rates were examined respectively at 8:00 on the next day. Afterwards, the fecundability and abortion rates of the female rats were observed during 5d, which the time we found the vagina! plugs was regarded as the "zero" day for the gestation. Next, the pregnant rats from each group were treated by BV again from the 6d to 15d. In addition, throughout the experiment period, not only all of the rat and the pregnant ones weights were investigated once weekly and every day respectively, but also the BV poisoning of rat were observed. After a short time (pregnant at 18d), the pregnant rats were sacrificed to investigate the growth and development status of embryos, fetal deaths, fetus resorptions, featus anomalies and placental weights.
Results:
1. The rats that exposed to BV had less action, less food intake, and bristling hair with poor gloss. There were no significant difference in growth rates for pregnant rats among 0 and 6d (P>0.05). After the pregnant rats from each group exposed to BV, their growth rates among 7 and 9d (pregnant 0-3d) showed no significant difference (P <0.05). However, growth rates of pregnant rats from BV 15 mg/m~3 group were lower than control group among 10 - 12d (P<0.05); in addition, growth rates from BV 10 mg/m~3 and BV 15 mg/m~3 group showed significant difference (P <0.05) bewteen 13 -15d and l6-18d.
2. The male rat' epididymal and testicular weights from each group decreased with the increase of BV dose, but their epididymal and testicular viscera coefficient were lower than control group (P<0.05). Futhermore, The female rat’ovarian viscera coefficients from each group were lower than control group (P<0.05).
3. Deformity rates of 1.90 %±0.14 %, 2.18 %±0.34 %, 2.83 %±0.26 %, 4.20 %±0.21 % had been obtained from the rats sperm of control, BV 5 mg/m~3, BV 10 mg/m~3 and BV 15 mg/m~3 group, respectively. Furthermore, deformity rates among the four group were statistically significant (F=182.35, P<0.01), which mainly include Big Head, Small head, banana head, hook-type head, double, three, neck bending, neck torsion, body bending, folding tail, short tail and so on.
4. Histopathologic studies showed that the morphous and edges of the seminiferous tubule from control rat testes remained regular and intact. Moreover, in the inner wall of seminiferous, the tubule spermatogoniums, primary and secondary spermatocyte were densely clustered together from external to internal directions with clear hiberarchy, and a lot of mature sperm cells could also be seen in the lumen round the middle of seminiferous. However, the tubule from exposed rat testes differed from this pattern. Such as, irregular morphous, quite incomplete edges, different sizes, porous arrangement of cells in the inner wall, degeneration and even loss of spermatogoniums, primary and secondary spermatocyte. In brief, the higher the dose was, the more obvious the pathological damage was.
5. The levels of testosterone in serum from exposed rat testes were lower than that from control ones (P<0.05). Though the levels of FSH、LH、E2 in serum from BV 5 mg/m~3 group were no significant difference from control ones (P>0.05), that from BV 10 mg/m~3 and BV 15 mg/m~3 group were lower than that from control ones (F<0.05).Likewise, the placental weights of rats from BV 10 mg/m~3 and BV 15 mg/m~3 group were lower than that from control ones(P<0.05).
6. The occurrence probability of fetal deaths and fetus resorptions from BV 10 mg/m and BV 15 mg/m group could be added (P<0.05) though the pregnancy rates of rats from each group were no significant difference (P>0.05). The weights, body and tail length of the fetuses in the rats from BV 10 mg/m~3 and BV 15 mg/m~3 group were lower than that from control ones (P<0.05). Therefore, Benzene can inhibit the growth of fetal rats in vivo with a dose - response relationship.
Conclusions:
1. Benzene, responsible for reproductive toxicity of rat, can induce change in some index, such as hormone levels, sperm morphology, testis tissue of male rat, and hormone levels of female rat.
2. Benzene, also responsible for embryonic development toxicity, can result in adverse pregnancy outcomes, growth and development delay of fetus and even death.
引文
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