Proteomic Profiling Of Extracellular Vesicle In Mouse Testis,Epididymis And Prostate

Objective:The aim of this study is to identify the proteins of testicular,epididymal and prostatic extracellular vesicles(EVs)in order to explore the effects of proteins on sperm development and maturation and maintenance of function.Methods:We employed TEI(Total Exosome Isolation)method to enrich the EVs of testis,epididymis and prostate.Markers of EVs were analysed by Immunoblotting.Morphology of EVs was observed with transmission electron microscopy(TEM).Distribution of particles size was detected by the dynamic light scattering(DLS).The testicula r,epididymal and prostatic EVs proteins were analyzed by liquid chromatography-tandem mass spectrometry(LC MS/MS).Using DAVID(The Database for Annotation,Visualization and Intergrated Discovery)6.8 online analysis tools,GO(gene ontology)functional analysis of the identified proteins was categorized based on cellular component,molecular function,biological processes.Results:1.CD63,TSG101,markers of extracellular vesicles were enriched in EVs by western blotting analysis.EVs show the cup-shaped morphology under the TEM.By the dynamic light scattering(DLS),testicular particles display the wide size distribution(13-450nm),major peak was 30-140 nm.The diameter of epididymal EVs is distributed in three segments of 13-24 nm,24-50 nm,50-250 nm.Prostatic EVs are distributed in three segments of 7-28 nm,28-68 nm and 68-220 nm.2.In this study,we identified 1148 testicular extracellular vesicle proteins,902 epididymal extracellular vesicle proteins,and 651 prostatic extracellular vesicle proteins.Among them,there are 277 common proteins in the three kinds of vesicles,667 specific proteins in testicular EVs,311 specific proteins in the epidid ymal EVs and 142 specific prostatic EVs proteins.3.GO enrichment analysis found that the common proteins mainly played roles in protein binding,RNA binding and nucleic acid binding,and participated in biological processes such as protein translation and transportation.Based on the analysis of the specific proteins,the testicular EVs proteins specifically were mapped to the spermatogenesis and cellular response to DNA damage and DNA repair,epididymal EVs proteins to transport,multi-metabolic process and immune process and prostatic EVs to the metabolic process.Conclusion:It can be seen that the testicular,epididymal and prostatic EVs play roles in the spermatogenesis and post-testicular sperm maturation and maintenance of function through multiple biological processes.