Effects Of Maternal Deoxynivalenol Exposure On Testicular Development And Spermatogenesis In Male Offspring

Deoxynivalenol(DON)is one of the most common mycotoxins,which poses a serious threat to the health of the animal husbandry due to its widespread and serious contamination.Recent studies have shown that DON exposure can reduce reproductive performance in male livestock,and DON exposure during lactation can cause mammary gland damage and affect milk quality in mother mice.However,it is still unknown whether DON exposure affects the reproductive performance of offspring.Therefore,21-day-old weaning mice(PND21)and 70-day-old adult mice(PND70)produced by DON exposure mother mice were used to investigate the effects of maternal DON exposure on male offspring testicular development and spermatogenesis.Immunohistochemical fluorescence,q RT-PCR,ELISA,Western blot,flow cytometry and other technologies were used to investigate the effects and potential mechanism of maternal DON exposure on testicular development and spermatogenesis in offspring male mice,and we also investigated the effects of maternal DON exposure on the transcriptomics of testis tissue using RNA-seq.The main results of this study are as follows:1.Effect of maternal deoxynivalenol exposure on testicular morphology and pathological changes in male offspring miceThe body weight,testicular weight and testicular size of mice were recorded,and it was found that maternal DON exposure significantly reduced the weight and testicular volume of PND21 and PND70 mice.HE staining showed that there were significant loss of sertoli cells and germ cells and disorganization of the germ cell layer in the seminiferous tubules of PND21 and PND70 mice after maternal DON exposure.The area of seminiferous tubules and the thickness of germinal epithelium were significantly reduced.2.Effect of maternal DON exposure on steroid hormone synthesis in male offspring miceThe ELISA method was used to detect the hormone levels in serum and testis,and the results showed that the testosterone concentrations in the testis of both PND21 and PND70 mice decreased significantly after maternal DON exposure,with a decrease of45.7% and 50.0%,respectively.And there was a significant decrease in both estrogen and testosterone levels in the serum of PND70 mice,with a decrease of 12.9% and 11.1%,respectively.Meanwhile,there was a significant decrease in both testis and serum concentration of INH-B.Further,the level changes of steroid hormone synthesis proteins in testis were detected by Western blot,and the results showed that maternal DON exposure significantly decreased the levels of steroid hormone synthesis-related proteins St AR,CYP11A1,and CYP17A1 in PND21 and PND70 mice.3.Effect of maternal DON exposure on proliferation,apoptosis and inflammation in the testis of male offspring miceThe ELISA was used to detect the levels of inflammatory factors in testicular tissues,and the results showed that maternal DON exposure significantly increased the contents of IFN-γ of the PND21 mice and IL-1β,IL-6,TNF-α of PND21 and PND70 mice.Among them,the levels of IL-1β increased 1.99 and 0.70-fold,IL-6 increased 0.85 and 0.39-fold,and TNF-α increased 2.30 and 0.43-fold,respectively.The level of IFN-γ increased0.67-fold in the testis of PND21 mice.The results of testicular immunofluorescence staining showed that maternal DON exposure significantly reduced the number of PCNA-positive cells in the seminiferous tubules of PND21 and PND70 mice.Western blot analysis showed that maternal DON exposure significantly reduced the level of proliferation-related protein PCNA,but significantly increased the level of the apoptosis-associated protein BAX.4.Effect of maternal DON exposure on the transcriptome of testicular cells of male offspring miceThe transcriptome results showed that 1098 differential expressed genes were identified between maternal DON exposure PND21 mice and control PND21 mice,including 151 up-regulated genes and 947 down-regulated genes.GO enrichment result showed that differential genes were mainly enriched in sperm development,spermatogenesis and other processes.A total of 2732 differential expressed genes were identified between maternal DON exposure PND70 mice and control PND70 mice,including 1122 up-regulated genes and 1610 down-regulated genes.GO enrichment result showed that the differential genes were mainly enriched in the entries of sperm development and tight junctions.5.Effect of maternal DON exposure on spermatogenesis and epididymal seminal quality in male offspring miceThe immunofluorescence staining of testicular tissue showed a significant decrease in the number of positive cells for DDX4 and SCP3 in both maternal DON exposure PND21 and PND70 mice.DNA ploidy analysis of the testis in the maternal DON exposure PND70 mice showed a significant increase in the number of haploid cells and a significant decrease in the number of tetraploid cells in the testis.The epididymis was collected for semen quality analysis,and the results showed that the sperm density,sperm motility and sperm viability in the epididymis were significantly reduced,and the sperm malformation rate was significantly increased.6.Effect of maternal DON exposure on the blood-testis barrier in the testis of male offspring miceThe immunofluorescence staining of testicular tissue showed that maternal DON exposure significantly reduced the number of SOX9-positive cells and the mean fluorescence intensity of ZO-1 protein in maternal DON exposure PND21 and PND70 mice.Western Blot further revealed that maternal DON exposure significantly reduced the levels of ZO-1,Occludin,and Claudin3.In summary,maternal DON exposure can induce inflammatory response in the testis of weaning mice and adult mice,promote apoptosis of testicular cells,inhibit testicular cell proliferation,reduce the synthesis and release of reproductive hormones,and affect the integrity of the blood-testis barrier,ultimately leading to a decrease in sperm density and quality.