Expression Of Testicular Nerve Growth Factor And Vascular Endothelial Growth Factor In Streptozotocin-induced Diabetic Mice

Objectives: We aimed to observe Expression of testicular Nerve growth factor and Vascular endothelial growth factor in Streptozotocin-induced diabetic mice,further to investigate the correlation between testicular damage and testicular VEGF and NGF levels in diabetic mice.The study provided theoretical basis and new therapeutic direction for the treatment of diseases such as diabetes induced testicular damage and male reproductive system.Methods: Four weeks old male C57BL/6 mice were housed with free access to water and laboratory chow.Mice were maintained in a 12 h light/12 h dark cycle at constant room temperature(22~26?),humidity(50~80%).Mice were divided into two groups: group A(n=25)and group B(n=25).Diabetes were induced by a single intraperitoneal injection of streptozotocin(200mg/kg).One week after streptozotocin treatment,induction of diabetes in the experimental group was confirmed by blood glucose levels over 11.1mmol/L.Twenty mice from group A were successfully inducted of diabetes.Eighteen mice from above group were randomly divided into DM groups(n=18),eighteen mice from group B were randomly divided into control group(n=18).The testes tissues were extracted for histological examination from six mice of each group randomly at 8,12 and 20 weeks after that.Testes tissue samples were fixed in 10% formalin in phosphate buffer for 24 h.The tissues were sectioned into sequential 5?m sections using a microtome.Immunohistochemical staining of testis tissues was performed using the streptavidin/biotin method.The immunohistochemistry procedure for VEGF(Catalog number BA0407 Boster Immunoleader,Wuhan,China)and NGF(Catalog number BA0611-1 Boster Immunoleader,Wuhan,China)was performed.Distilled water was used in place of the primary antibody as negative controls.Immunohistochemical evaluation was performed based on the intensity of VEGF and NGF immunoreactivity in the testes.NGF and VEGF level were studied by immunohistochemistry measuring integrated optical density(IOD).Results are presented as mean±S.E.M.Statistical analysis was performed on SPSS 17.0 software.Difference in parameters of two groups was analyzed by using the Student's t-test(normality)or nonparametric test(one or two annormality).Correlations among groups were calculated using Pearson correlation analysis.A P value less than 0.05 was deemed significant.Results:Histological findings of each group:Seminiferous Tubules and interstitium in testicular tissue were found to be normal in the control animals.In testis tissue,NGF immunolabelling was localized in the nuclei of Sertoli cells,in the cytoplasm of pachytene spermatocytes,in round spermatids,and in elongating spermatids resembling round vesicular like structures.It is evident that Leydig cells show a markedly positive staining for VEGF.The blood vessels in the interstitial compartment did not stain for VEGF.Within the tubules,Sertoli cells showed a positive staining.Diabetic condition appeared to have impaired spermatogenetic process.As evidenced in the representative photographs which show VEGF and NGF expression,VEGF and NGF reactivity appears to be more prominent in control group compared to diabetic group.IOD of NGF and VEGF immunoreactivity was higher in control group compared to diabetic group.Expression fo NGF and VEGF in diabetes mice testis tissue decreased as time goes by(P<0.01).In correlation analysis,a very strong positive correlation was found between VEGF and NGF levels(r = 0.778,P<0.01)Conclusions:The research results show that seminiferous Tubules and interstitium in testicular tissue were found to be normal in the control animals.Germinal cells were found to be degenerated and disorganized and also reduced in number in diabetic rats.Diabetic mice the expression level of VEGF and NGF plays were reduced in the testicles,at the same time accompanied by increased apoptosis and testicular tissue damage.Speculated that diabetes is one of the causes for testicular tumor and testicle reproductive dysfunction,provided new train of thought and theoretical basis of pathogenesis.