Asthenozoospermia Related Protein GPI In Sperm Motility

Asthenozoospermia, as one of the most common findings in infertile males, is still a severe reproductive problem. About 10% to 20% of married couples suffer from reproductive difficulties, almost 50% of the infertility was caused due to the low male reproductive capacity. Male infertility was directly related to low sperm motility (asthenozoospermia), decreased sperm density (oligozoospermia) and abnormal sperm morphology(teratospermia). Among them, asthenospermia was the most important factor of male sub-fertility and infertility. Proteomics technology is one of the most valuable ways of doing life science research. It is efficient in analysis of proteins with large-scale application of biochemistry and molecular biology methods. Recent studies have already provided some evidence that proteomics research in the identification of several proteins involved in asthenozoospermia using the method of 2-D gel electrophoresis is available. The subject of this study with the application of new proteomics technology which TMT label and LC-MS/MS involved was to compare the sperm protein of idiopathic patients with asthenospermia and normal donors. We also tried to find some specific proteins that regulate the sperm motility. First, sperm of asthenospermia patients and normal sperm specimens were collected. Then the whole sperm protein was extracted. The protein was labeled, isolated and identified. Finally,we identified the expression of 2574 different sperm protein by comparing 5 asthenozoospermic samples to that of 5 semen donor controls using LC-MS/MS(Orbitrap).A series of experiments of verification and function were conducted under the guidance of the proteomic data base.There were 131 proteins with an increased or with a decreased amount range>1.5,240 proteins with an increased or with a decreased amount range>1.3.Then with the classical route of identification of unknown protein, Experiments of western blotting and enzyme-linked immunosorbent assay (ELISA) were done to verify the different expression of protein GPI (the protein is a key enzyme in the glycolytic pathway) in asthenospermic sperm and normal ones.The result confirmed the concentration of protein GPI in asthenozoospermic sample(30.2±35.6pg/ml) is significantly lower than(P<0.001) that in the normozoospermic samples(86.9±49.5pg/ml);western blotting showed the same result. Also we selected GPI as a target for clinic therapy and conducted drug trails which showed the positive effect on sperm motility. Supplemented F-6-P(fructose-6-phosphate, reaction product which GPI-catalyzed) was added in sperm of different motility types, then the motility was observed to reflect the effect of F-6-P. Results show that the motility were significantly increased in moderately weak sperm (P<0.01) and normal sperm (P<0.05) compared to the control group after the addition of F6P.Meanwhile ATP concentration was significantly higher(P<0.05) in sperm which drug effected compared to control groups in drug trials.We have identified much more proteins in asthenozoospermic sperm samples compared with the recent data.We not only provide systematic and detailed research route which better proteomics labeling, separation, identification techs involved, but also find some ideas and therapeutic targets for clinical diagnosis and treatment of infertility caused by asthenospermia.