The Study On The Correlation Between Protein LGALS3BP From Seminal Plasma And Male Infertility Associated With Varicocele

ObjectiveIn the male infertility problem,Varicocele(VC)is one of the causes of male infertility,but the specific mechanism of VC infertility is still unclear.Therefore,a better understanding of the pathophysiology of VC-related male infertility is essential to elucidate the harmful effects of VC on spermatogenesis and testicular function.It is particularly important to explore new biomarkers for the diagnosis and treatment of male infertility and the prediction of pregnancy outcomes.Previous studies suggest that high concentrations of protein in the seminal plasma are good research objects.Due to the complex composition of semen,exosomes in body fluids are characterized by the enrichment of specific diagnostic biomarkers,which will help researchers to discover relatively low expression biomarkers that are not easily to detect,making exosomes and the substance contained in it can play a role in the detection,treatment and prognosis assessment of diseases.As an emerging biological marker,exosomes and their encapsulated substances have shown the important role in the process of disease diagnosis in many studies.Based on the pre-study of spermatic cord varicose vein infertility patients’ seminal plasma exosome protein expression profile and existing literature basis,this study will screen differentially expressed proteins based on the results of the previous research,and use bioinformatics to analyze biological function,screen key proteins,At the same time,on the premise of successful extraction of seminal plasma exosomes,expand the sample size,detect the expression level of seminal plasma exosome LGALS3BP protein,explore LGALS3BP protein as a biological marker,in VC-related male,The clinical significance of infertility.It provides a theoretical basis for further exploring the biomarkers of VC-related male infertility and the mechanism of VC infertility,so as to promote the progress of male infertility research.Methods1.According to the inclusion/exclusion criteria,the clinical samples were included,and the quantitative proteomics based on TMT labeling combined with tandem mass spectrometry was used to compare the seminal plasma exosomes of infertile patients with varicocele and normal physical examination.Proteomics research.Set the screening conditions to screen out the union histone and the intersection histone.The "clusterprofile" R package was used to perform GO enrichment analysis on the union group protein and the intersection group protein.2.Use ultra-high-speed centrifugation to extract seminal plasma exosomes,and use three methods:WB,NTA,and TEM to identify the exosome surface marker protein,exosome particle size,and exosome morphology under electron microscope.Exosomes.Western Blot,ELISA,immunohistochemistry and other methods were used to detect the protein expression in normal male testis tissue,mouse GC1,GC2,TM4 cells,VC rat model and clinical samples to verify the protein expression and cell localization.3.Finally,the statistical method Spearman correlation coefficient(r),ROC curve analysis and other methods are used to verify the clinical significance of LGALS3BP protein as a biological marker for VC-related male infertility.Results1.The results of comparative proteomics research showed that a total of 1457 proteins were identified in the spermatic vein varicocele exosome protein profile,and 430 histones were combined and 114 histones were intersected.The results of bioinformatics analysis show that most of the proteins with statistical differences are derived from exosomes,and their biological processes and molecular functions are related to spermatogenesis and development.2.Screen the differential protein LGALS3BP as the candidate key protein according to the previous research results and existing literature reports.3.Successful identification of seminal plasma exosomes and verification of the reliability of differential protein profiles.The expression level of LGALS3BP in normal testis tissues in men is moderate,and is located in the cytoplasm and cell membrane.The expression level of this protein in mouse testicular support cells is significantly higher than that of mouse spermatogonia and mouse spermatocytes.LGALS3BP protein is lowly expressed in seminal plasma exosomes and rat testis tissues of VC infertility group.4.The expression level of seminal plasma exosome LGALS3BP protein was positively correlated with total sperm concentration(Concentration),forward motility sperm rate(Progressive motility,PR)and total sperm motility(PR+NP).The ROC curve analysis results show that the seminal plasma exosomal protein LGALS3BP protein has higher diagnostic efficacy than the seminal plasma protein.ConlusionsThe expression level of LGALS3BP protein is positively correlated with the degree of spermatogenic impairment,sperm motility,and sperm concentration,and has the potential clinical value for the diagnosis of varicocele combined with male infertility.