| Zinc is one of the essential trace elements in the body.In the male reproductive system,zinc ions are particularly important for the development and function of testes and sperm.Changes in zinc homeostasis affect fertility.Zinc levels in diabetic patients have been imbalanced and lead to male infertility through multiple levels of action.Strict control of zinc homeostasis is essential for maintaining body health.The zinc ion balance is mainly regulated by two functionally opposite families of zinc transporters,including the Zips family and the ZnTs family.The purpose of this study is to explore the expression of zinc transporter in male reproductive gland of diabetes mellitus,and to provide some basic experimental data for revealing the mechanism of diabetes on male fertility.Methods:Twenty SD male rats were randomly divided into control group and diabetic group?DM?with 10 rats each.The control group was fed with normal diet and the DM group was fed with high fat and sugar diet.Rats in the DM group were given a 30 mg·kg-1streptozotocin intraperitoneally after 8 weeks.Rats in the control group were intraperitoneally injected with the same amount of sodium citrate buffer.Fasting blood sugar above 11.7mmol/L was regarded as the success of modeling.Four weeks after the establishment of the model,rats in each group were killed.Testicles,epididymis,seminal vesicles and prostate tissues were taken out completely.Some of them were fixed in 10%formalin solution.The remaining tissues were immediately put into liquid nitrogen,and then stored at-80°C.The pathological changes of the above tissues were observed by HE staining,and the localization of ZnT1,ZnT8,Zip1 and Zip2 in the above tissues was determined by immunohistochemistry.The expression of ZnT1,ZnT8,Zip1 and Zip2 in the above tissues was detected by Western Blot and RT-PCR.Results:?1?After 4 weeks of successful modeling,the final body weight,fasting blood glucose,TG,TC,HDL-C,and LDL-C values in the diabetic group were[?534.96±26.67?g,?16.48±1.47?mmol/L,respectively.?1.65±0.25?mmol/L,?2.82±0.28?mmol/L,?0.31±0.05?mmol/L,?0.43±0.06?mmol/L,and the final weight of the control group,fasting blood glucose,TG,The values of TC,HDL-C and LDL-C were[?440.10±57.14?g,?6.72±1.16?mmol/L,?0.73±0.24?mmol/L,?1.49±0.38?mmol/L,?0.58±,respectively?.0.06)mmol/L,?0.29±0.04?mmol/L,the difference was statistically significant?P<0.05?;In the diabetic group,sperm concentration[?36.44±4.88?×106/ml]and sperm motility[?41.37±3.61?%]were significantly lower than the normal group[?45.36±7.90?×106/ml,?52.46±7.38?%],the difference was statistically significant?P<0.05?.?2?HE staining:The different types of cells in the testicular seminiferous tubules of the control rat were arranged in an orderly manner,the structure was relatively complete,the layers were distinct,and a large number of sperm could be seen;In the epididymis duct,a large number of sperm can be clearly seen and arranged irregularly.At the same time,the nucleus and cytoplasm of the cells can be clearly observed;The seminal vesicle glandular cavity contains eosinophilic secretions,and there are many folds in the glandular cavity;The prostate usually grows stationary,showing thin epithelium,smooth inner wall,deep and complete cytoplasm staining.In the DM group,the number of cell layers in the seminiferous tubules of the testis was reduced,some interstitial cell structures were also destroyed,vacuoles were observed in the middle of the tissues,and the number of sperm was also greatly reduced.The number of sperm in the epididymis has decreased significantly,and many changes have taken place in tissue morphology.At the same time,vacuole-like changes were observed in epididymal epithelial cells and the nuclei of some main cells were dissolved and ruptured;The folds in seminal vesicle gland cavity were reduced,and the volume of coagulation gland was reduced accordingly;Prostate inner wall folds increased,showing epithelial hyperplasia,cytoplasmic staining was shallow and the loose structure changed.?3?Immunohistochemical staining:ZnT1 was mainly expressed in plasma membrane,interstitial cells and spermatogenic cytoplasm of rat testis.The expression in control group was significantly higher than that in DM group;The expression of ZnT8 in Leydig cells and spermatogenic cytoplasm of testis was lower in the control group than in the DM group;Zip1 and Zip2 were mainly expressed in the cytoplasm and plasma membrane of testicular spermatogenic cells,and the control group was significantly higher than the DM group.ZnT1 and Zip1 were mainly expressed in the cytoplasm of the epididymis,and the control group was lower than the DM group;The expression of ZnT8 in epididymis was mainly in the epithelial cells of epididymis duct,basal cells of epididymis epithelium and intraluminal cytoplasm,and the control group was lower than the DM group;Zip2 mainly expressed epididymal ductal cells and intraluminal cytoplasm in the epididymis,and the control group was higher than the DM group.ZnT1,ZnT8,Zip1 and Zip2 were expressed on the cytoplasm and membrane of rat seminal vesicle epithelial cells,and the expression control groups of ZnT1 and Zip1 were significantly higher than those of DM group.ZnT1,ZnT8,Zip1 and Zip2 were mainly expressed in the cytoplasm and membrane of prostate epithelial cells in the rat prostate,and the control group was slightly lower than the DM group.?4?Western blot and RT-PCR:The expression of ZnT1 protein and mRNA in the testis and seminal vesicles was significantly higher than that in the DM group?P<0.05?,and the control group in the epididymis was significantly lower than the DM group?P<0.05?.The expression of ZnT1 mRNA was significantly increased in DM group?P<0.05?.The expression of ZnT8 protein and mRNA in testis,epididymis,seminal vesicle and prostate were lower than those in DM group?P<0.05?;Zip1 protein and the expression of mRNA in the testis and prostate tissues was significantly higher than that in the DM group?P<0.05?.The expression of Zip1 protein and mRNA in the epididymis was significantly higher in the DM group than in the control group?P<0.05?.The expression of Zip2 protein was significantly higher in the epididymis than in the DM group?P<0.05?,and there was no statistical difference in the other three tissues.The expression of Zip2 mRNA was decreased in testis and epididymis of DM group,and increased in seminal vesicle and prostate.There was no statistical difference in the seminal vesicle?P>0.05?,and there was significant difference between the other two groups?P<0.05?.Conclusions:The expressions of ZnT1,ZnT8 and Zip1 and Zip2 mRNA and protein in gonads of diabetic rats are presumed to play an important role in the balance of zinc homeostasis.It is speculated that the changes of gonadotropin transporter in diabetic male rats may cause a decrease in reproductive ability. |
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