正己烷静式染毒致SD大鼠生殖腺损伤的实验研究
发布时间：2018-11-19 21:08
【摘要】：目的建立正己烷(n-hexane)静式染毒的SD大鼠实验模型，并对实验模型的稳定性、可重复性及其改良应用进行探讨。在确定正己烷导致SD大鼠睾丸和卵巢损伤的基础上，展开关于损伤过程可能毒性机制的初步观察研究，为流行病学和职业病有关正己烷的毒理研究和制定防护措施提供参考依据。方法SD大鼠48只，随机分为4组(每组12只，雌雄各半)，1个对照组和3个染毒组(1d、3d和7d组)。置于特制染毒箱中，采用静式染毒法，染毒时间为8h／d共7d，正己烷初始浓度设为35.2g／m~3。各染毒组大鼠于染毒结束后30min内取动脉血，气相色谱-质谱仪检测全血正己烷浓度；取睾丸或卵巢称量并计算脏器系数。睾丸和卵巢组织，一侧制组织匀浆测定谷胱甘肽过氧化物酶(GSH-Px)、谷胱甘肽(GSH)、超氧化物歧化酶(SOD)活性和丙二醛(MDA)的含量，另一侧4％多聚甲醛固定后常规石蜡切片观察病理变化。免疫组织化学两步法，检测正己烷静式染毒致损害的SD大鼠睾丸组织和卵巢组织中Fas、FasL、bax、bcl-2等凋亡相关蛋白的的表达。结果染毒1d、3d和7d组大鼠，经检出血液正己烷平均浓度分别为：636.92μg／L、913.49μg／L和1090.69μg／L，均明显高于正常，表明已达到染毒要求。显微镜观察发现，实验SD大鼠睾丸和卵巢结构均出现明显病理变化。染毒实验组的SD大鼠，随染毒时间延长，睾丸和卵巢显微结构的破坏逐渐明显和加重。睾丸生精小管中支持细胞形态模糊不清，各级生精细胞排列疏松，部分小管中生精细胞严重脱落，有的管腔出现多核巨细胞。SD大鼠卵巢中生长卵泡数量减少，一些生长卵泡中卵母细胞的核膜及核仁不清晰，透明带模糊甚至已溶解，黄体中淤血致血细胞浸润。染毒鼠的体重和生殖腺重量均有所降低，染毒7d组最为明显(p＜0.05)，器官体重比也呈现明显下降趋势，3d组和7d组具有统计学意义(p＜0.05)。随染毒时间延长，睾丸或卵巢组织中SOD、GSH、GSH-Px均有降低，MDA含量升高，尤以7d组最为显著(p＜0.01)，显示正己烷染毒所致的不同程度的脂质过氧化损伤。睾丸中Fas、FasL、bax蛋白在7d组均有增强，位于生精小管和生精细胞；卵巢中各凋亡相关蛋白，随着染毒时间的延长表达强度有所增加，主要表现在黄体细胞和颗粒细胞。结论1．本研究采用“正己烷急性吸入静式染毒法”，建立的正己烷吸入致SD大鼠睾丸和卵巢损伤的实验模型，具有操作简单、效果稳定和可重复性强等优点，可为正己烷和其他毒物致代谢器官损害的实验研究提供方法学依据。2．正己烷静式染毒导致损伤的SD大鼠，体重下降，睾丸或卵巢的湿重和脏器系数降低、睾丸和卵巢的结构出现病理变化，证实正己烷毒性作用导致睾丸和卵巢损伤。染毒时间越长，睾丸和卵巢的结构组织损伤越严重。3．SD大鼠经正己烷染毒后，所测出睾丸和卵巢匀浆中的SOD、GSH-Px和GSH和MDA含量变化表明，，随着染毒时间的延长，睾丸或卵巢中SOD、GSH-Px活力均呈明显下降，而MDA含量呈明显升高，反映正己烷的脂质过氧化毒性作用已导致睾丸和卵巢清除自由基能力明显下降，脂质过氧化产物大量堆积。脂质过氧化损伤是正己烷导致生殖腺损伤的机制之一。4．本研究检测与细胞凋亡相关的Fas、FasL、bax和bcl-2等的免疫组化表达特征，结果表明，正己烷中毒对睾丸和卵巢的影响，最终导致生殖细胞凋亡的过程。由于雄性和雌性个体的生殖细胞发育过程具有不同的特点，正己烷中毒对睾丸和卵巢的干扰和影响表现出不同的状况。但预示正己烷的毒性作用将促使和导致生殖细胞凋亡，必然会影响卵巢生殖功能。
[Abstract]:Objective To establish an experimental model of n-hexane-based SD rat model, and to study the stability, repeatability and application of n-hexane. Based on the determination of the testicular and ovarian damage of SD rats caused by n-hexane, a preliminary study on the possible toxicity mechanism of the injury process was conducted to provide a reference for the toxicological study and protective measures of n-hexane related to epidemiology and occupational disease. Methods Forty-eight SD rats were randomly divided into 4 groups (12 in each group, half of male and female), 1 control group and 3 groups of exposure (1d, 3d and 7d). The initial concentration of n-hexane was 35. 2g/ m ~ 3, and the initial concentration of n-hexane was 35. 2g/ m ~ 3. Blood and gas chromatography-mass spectrometer (GC-MS) were used to measure the concentration of n-hexane in the whole blood for 30min after the end of the exposure, and the testis or the ovary was weighed and the coefficient of the organ was calculated. The contents of glutathione peroxidase (GSH-Px), glutathione (GSH), superoxide dismutase (SOD) and malondialdehyde (MDA) were measured on one side of the testis and the ovary tissue, and the pathological changes were observed in the normal paraffin section after the fixation of 4% paraformaldehyde on the other side. The expression of Fas, FasL, bax, bcl-2 and other apoptosis-related proteins in the testis and ovarian tissues of SD rats was detected by two-step immunohistochemical method. Results The average concentration of n-hexane in the rats was 636. 92. mu. g/ L, 913. 49. m u.g/ L and 1090. 69. m u.g/ L, respectively. The results showed that both the testis and the ovary structure of SD rats had obvious pathological changes. The SD rats of the experimental group were prolonged with the time of exposure, and the destruction of the microstructure of the testis and the ovary became more and more serious. In the testis spermatogenic small tube, the cell morphology is blurred, and the spermatogenic cells at all levels are arranged to be loose, and the spermatogenic cells in some small tubes are seriously detached, so that the tube cavity has a plurality of giant cells. The number of follicles in the ovary of SD rats was decreased, and the nuclear membrane and the nucleolus of the oocytes in some of the growing follicles were not clear, the clear band was blurry and even dissolved, and the blood cells in the corpus luteum were infiltrates. The weight of the rats and the weight of the gonads decreased, and the group was the most obvious (p <0.05), and the weight ratio of the organs decreased significantly, and the 3d and 7d groups were of statistical significance (p <0.05). The contents of SOD, GSH and GSH-Px in the testis or the ovary were decreased with the time of exposure, and the content of MDA increased, especially in the 7dgroup (p <0.01). The expression of Fas, FasL and bax in the testis was enhanced in the 7dgroup, and the expression of Fas, FasL and bax in the ovary was increased, and the expression of Fas, FasL and bax in the ovary increased with the time of exposure, mainly in the yellow cells and the granulosa cells. Conclusion 1. The experimental model of the rat's testis and the ovary injury induced by n-hexane inhalation by using the 鈥渘-hexane acute inhalation and static exposure method鈥
本文编号：2343425
[Abstract]:Objective To establish an experimental model of n-hexane-based SD rat model, and to study the stability, repeatability and application of n-hexane. Based on the determination of the testicular and ovarian damage of SD rats caused by n-hexane, a preliminary study on the possible toxicity mechanism of the injury process was conducted to provide a reference for the toxicological study and protective measures of n-hexane related to epidemiology and occupational disease. Methods Forty-eight SD rats were randomly divided into 4 groups (12 in each group, half of male and female), 1 control group and 3 groups of exposure (1d, 3d and 7d). The initial concentration of n-hexane was 35. 2g/ m ~ 3, and the initial concentration of n-hexane was 35. 2g/ m ~ 3. Blood and gas chromatography-mass spectrometer (GC-MS) were used to measure the concentration of n-hexane in the whole blood for 30min after the end of the exposure, and the testis or the ovary was weighed and the coefficient of the organ was calculated. The contents of glutathione peroxidase (GSH-Px), glutathione (GSH), superoxide dismutase (SOD) and malondialdehyde (MDA) were measured on one side of the testis and the ovary tissue, and the pathological changes were observed in the normal paraffin section after the fixation of 4% paraformaldehyde on the other side. The expression of Fas, FasL, bax, bcl-2 and other apoptosis-related proteins in the testis and ovarian tissues of SD rats was detected by two-step immunohistochemical method. Results The average concentration of n-hexane in the rats was 636. 92. mu. g/ L, 913. 49. m u.g/ L and 1090. 69. m u.g/ L, respectively. The results showed that both the testis and the ovary structure of SD rats had obvious pathological changes. The SD rats of the experimental group were prolonged with the time of exposure, and the destruction of the microstructure of the testis and the ovary became more and more serious. In the testis spermatogenic small tube, the cell morphology is blurred, and the spermatogenic cells at all levels are arranged to be loose, and the spermatogenic cells in some small tubes are seriously detached, so that the tube cavity has a plurality of giant cells. The number of follicles in the ovary of SD rats was decreased, and the nuclear membrane and the nucleolus of the oocytes in some of the growing follicles were not clear, the clear band was blurry and even dissolved, and the blood cells in the corpus luteum were infiltrates. The weight of the rats and the weight of the gonads decreased, and the group was the most obvious (p <0.05), and the weight ratio of the organs decreased significantly, and the 3d and 7d groups were of statistical significance (p <0.05). The contents of SOD, GSH and GSH-Px in the testis or the ovary were decreased with the time of exposure, and the content of MDA increased, especially in the 7dgroup (p <0.01). The expression of Fas, FasL and bax in the testis was enhanced in the 7dgroup, and the expression of Fas, FasL and bax in the ovary was increased, and the expression of Fas, FasL and bax in the ovary increased with the time of exposure, mainly in the yellow cells and the granulosa cells. Conclusion 1. The experimental model of the rat's testis and the ovary injury induced by n-hexane inhalation by using the 鈥渘-hexane acute inhalation and static exposure method鈥
本文编号：2343425
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