Photoperiod Regulates The Epididymal Structure And Expression Of GPX5 In The Epididymis Of Cricetulus Barabensis Through Androgen

Photoperiod plays an important role in regulating the activity of seasonal breeding animals.Seasonal changes in the photoperiod can activate or inhibit the reproductive system.Sperm produced by the testis need to pass through the epididymis to become mature sperm.The function of the epididymis is highly dependent on androgens,so the content of androgens also affects the function of the epididymis.The androgen that plays a major role in the epididymis is dihydrotestosterone(DHT),which is the product of testosterone(T)catalyzed by 5α-reductase.T and DHT need to bind to androgen receptors to work(AR).Glutathione peroxidase 5(GPX5)is an antioxidant enzyme that is almost exclusively found in the epididymis and has a strong protective effect on sperm in the epididymis.However,the detection of androgen content in epididymis under different photoperiod conditions is very poor,and the relationship between GPX5 and photoperiod is also rarely reported.The Cricetulus barabensis is a small rodent whose breeding activity is typically seasonal.In this study,C.barabensis were selected as animal models to detect the effects of different photoperiod conditions on the structure of the epididymis,the content of androgens,and the expression of GPX5 in the epididymis,so as to explore the relationship between photoperiod,androgen and GPX5.18 adult male C.barabensis were randomly divided into three groups: Long photoperiod group(14L / 10 D,4:00 to 18:00 light,LP);Medium photoperiod group(12L / 12 D,6:00 to18:00 light,MP);Short photoperiod group(10L / 14 D,8:00-18 :00 light,SP).Animals in all groups were kept at 22 ° C for 28 days of photoperiod treatment.Body mass of the hamsters was weighed at the end of treatment.After the hamster was killed by carbon dioxide anesthesia,both sides of the epididymis were quickly removed,the caput and cauda epididymis were separated and stored separately for subsequent experiments:(1)The contents of T and DHT were detected by ELISA kits;(2)The luminal area and epithelial thickness of the caput epididymis and cauda epididymis were observed by paraffin section and HE staining;(3)The expression localization of S5AR1,S5AR2,AR,and GPX5 in caput epididymis and cauda epididymis were observed by immunohistochemistry(IHC);(4)The m RNA expressions of Srd5a1,Srd5a2,AR,and GPX5 in the caput epididymis and cauda epididymis were detected by quantitative real-time PCR;(5)The protein levels of S5AR1,S5AR2,AR,and GPX5 in the caput epididymis and cauda epididymis were detected by Western Blot.The results showed:(1)After different photoperiod treatments,there was no significant difference in body mass among the groups,but the epididymal weight of the SP group was significantly lower than that of the LP group.(2)The results of ELISA showed that in the caput epididymis,the contents of T and DHT in LP group were the highest,which were significantly higher than those in MP and SP groups;in the cauda epididymis,the contents of T and DHT in LP and SP groups were significantly higher than those in MP group.(3)In the caput epididymis,there were significant differences in the epithelial thickness and lumen area of the epididymal duct in different groups.The epithelial thickness of the LP group was significantly higher than that of the MP and SP groups,but the epididymal lumen area of the LP group was the smallest,which was significantly smaller than that of the MP and SP groups.In the cauda epididymis,the epididymal duct epithelial thickness and lumen area of different groups were also significantly different.The epididymal duct epithelial thickness of SP group and LP group was larger than that of MP group,while the epididymal duct lumen area of MP group was significantly larger than that of LP group.(4)The positive staining of S5AR1 and S5AR2 was observed in the cytoplasm of epididymal epithelial cells of the three groups,and the strong staining of AR was observed in the cytoplasm and nucleus of the epididymal epithelial cells,and the strong staining of GPX5 was observed in the cytoplasm and sperm cells of the epididymal epithelial cells.(5)The results of quantitative PCR showed that in the caput epididymis,there were significant differences in the relative expression of Srd5a1 m RNA among the three groups.The relative expression of Srd5a1 m RNA in the LP group was significantly higher than that in the SP group,but there was no significant difference in the relative expression of Srd5a2 m RNA.The relative expression of AR m RNA in LP and MP groups was significantly higher than that in SP group,and the relative expression of GPX5 m RNA was significantly different,and the relative expression of GPX5 m RNA in LP group was significantly higher than that in MP and SP group.In the cauda epididymis,the relative expression of Srd5a1 m RNA was significantly different among the three groups.The relative expression of Srd5a1 m RNA in LP group was significantly higher than that in SP group,but the relative expression of Srd5a2 m RNA and AR m RNA was not significantly different.The relative expression of GPX5 m RNA was significantly different,and the relative expression of GPX5 m RNA in LP and SP groups was significantly higher than that in MP group.(6)Western Blot results showed that in the caput epididymis,there were significant differences in the level of S5AR1 protein among the three groups,the level of S5AR1 protein in LP group was significantly higher than that in SP group,the level of S5AR2 protein was not significantly different among the three groups,and the level of AR protein was significantly different.The AR protein level of LP group was significantly higher than that of SP group,but the GPX5 level was not significantly different among the three groups.In the cauda epididymis,the S5AR1 protein level of the three groups was also significantly different,the S5AR1 protein level of the LP group was significantly higher than that of the MP and SP groups,the S5AR2 and AR protein level was not significantly different among the three groups,the GPX5 protein level was significantly different,and the GPX5 protein level of the MP group was significantly lower than that of the LP and SP groups.In summary:(1)The increase of androgen content in the caput epididymis of C.barabensis under long photoperiod promoted the growth of epididymal head epithelial cells;the significant increase in the lumen area of the cauda epididymis during the medium photoperiod is beneficial to the storage of a large number of sperm to ensure the subsequent reproduction process.(2)In the epididymis of C.barabensis,the expression of 5α-reductase 1 is more dominant,and the long photoperiod mainly promotes the conversion of testosterone into dihydrotestosterone through the action of 5α-reductase 1.(3)The photoperiod-regulated androgen content in the epididymis is consistent with GPX5 expression levels,suggesting that photoperiod may regulate GPX5 levels in the epididymis of C.barabensis through the androgen/androgen receptor pathway.The results lay the foundation for elucidating the mechanism of the antioxidant system in the epididymis regulated by photoperiod,and it is also of great significance for understanding the mechanism of the regulation of the epididymis by seasonal reproduction in mammals.