The Toxic Effects Of 3,4-dichloroaniline On The Reproductive System Of Male Rats

Objective:Dichloroanilines,widely used as the intermediates in the synthesis and degradation of herbicides,pesticides and azo dyes,have many ways to enter and accumulate in the environment,posing a hazard to human health.In this study, we detected the effects of 3,4-dichloroaniline(DCA) on the reproductive system of male rats in six aspects.We provided some basic information on reproductive toxicity mechanism of Anilines.It is very helpful to protect our living environments as well as human health.Methods:40 adult male rats were randomly divided into 5 groups:the oil control group and 4 exposed groups in which the rats were orally given different dosage of 3,4-dichloroaniline(39mg/kg,81mg/kg,170mg/kg and 357mg/kg).After exposed for 35 days,all the animals were sacrificed and weighed.The influences to the reproductive system were detected using the methods as follows:1 To observe the Morphology of testes and epididymis using paraffin section & HE dying.To detect the deformation of sertoli cells,germ cells and sperms by using TEM.2 To purify the spermatocytes in the testis by using portfolio enzymatic digestion-percoll division and then to detect their DNA damages with Comet Assay.3 To assess The epididymal sperm density and the viability of spermatozoa by trypan blue exclusion assay.4 To detect the acrosomal enzyme activity of the sperms by using the method of Spectrophotometry.5 To detect the ratio of acrosome reaction of sperms by using coomassie brilliant blue staining.6 To detect the DNA damage of sperms by using COMET assay.Results:All calculations and statistical analyses were generated in SPSS for windows version 11.0.The data were expressed in means±standard deviation (SD).We found that 357mg/kg exposure of 3,4-DCA could decrease the body weights significantly(P＜0.01),so did the other three groups but not ina significant way. The spleens of the rats were influenced dramatically in each dosage of DCA(P＜0.01). Except the 357mg/kg group,the thymus of the other three groups performed a decreased weights according to the different exposure of DCA,but not in an apparent changing compared with the control.The coeffecience of testis in 357mg/kg group was increased significantly,while the others didn't indicate any influence by DCA.2 With the increasing exposure of DCA,the structure of the seminiferous was damaged badly,performing an increased layers of outer folds,a decreased amount of spermatocytes and sperms,a shrinkage of cytoplasm and an elongated seminiferous tubule lumen diameter.The results of electron micrographs of testes sections showed vacuolization or cristae indistinction in mitochondria and expansion of endoplasmic reticulum after treated with 170mg/kg DCA.Spermatocytes in 170mg/kg DCA group showed collecting or abnormal distributing heterochromatins.Sperms in 170mg/kg DCA group showed a lacking mitochondrial sheath in middle piece and an abnormal outer dense fibers.3 The Comet Assay resulted in that the comet tails, the Olive values and the ratio of tail/head were all increased after the exposure of DCA and in a dose-dependent manner(P＜0.01).4 With the increasing dose of DCA exposure,the density,the viability of the sperms were inhibited but not significantly(P＜0.05),while the deformation was enhanced(P＜0.05).5 Except 39mg/kg DCA group,the acrosomal enzyme activity of the other three dosage groups were inhibited compared with the control group(P＜0.05).6 The ratio of acrosomal reaction in all the four exposure groups were decreased significantly(P＜0.01).7 Comet Assay results of the sperms showed that the DNA of the sperms were damaged dramatically in 170mg/kg and 357mg/kg DCA groups(P＜0.01),but not in a significant way in other two groups compared with the control.Conclusion:3,4-dichloroaniline can cause the morphology of the testis and epididymis,endanger the normal function of sertoli cells as well as the spermatocytes.It also can decrease the quality of the sperms and cause the damage of 'sperms' DNA,inhibit the acrosomal enzyme activity and lower the ratio of acrosome reaction.Therefore,we can make a conclusion that 3,4-dichloroaniline has a toxic effects to the reproductive system.We speculate that the mitochondria in the sertoli cells might be the target points of DCA,and the pathways might well have something to do with ROS.Anyway,lots of future work should be done in the field of the detailed signal transmission and the toxic mechanism of DCA on the reproductive system.