| Male infertility is one of the common andrology diseases,and its incidence has been increasing year by year in recent years.The treatment of Western medicine has some limitations,while Chinese medicine has unique advantages.As a Chinese patent medicine,Yishen Xingyang Capsule(YSXY)was mainly used to treat impotence and premature ejaculation in the early stage.Now it is more and more widely used to treat oligoasthenospermia.Therefore,it is of great significance to study and reveal the mechanism of YSXY in treating oligoasthenospermia.However,the overall pharmacodynamic mechanism has not been fully understood,so effective research ideas and methods are urgently needed to study the pharmacodynamic substance basis and mechanism of action,so as to guide the safe and rational clinical use of the medicine.This paper is divided into three parts:literature review,network pharmacology study and experimental study.The literature review systematically summarized the research progress of oligoasthenospermia by Traditional Chinese Medicine(TCM)in recent years,and discussed the understanding of TCM on oligoasthenospermia,TCM syndrome differentiation and treatment,and the research progress of Chinese herbal compound.The etiology of oligoasthenospermia,the regulation mechanism of spermatogenic function of testis and the treatment methods were discussed from the point of view of modern medicine.The discussion of the above two contents has paved the way for the smooth development of this subject.Objective:This study aims to predict the key targets,biological pathways and other related mechanisms of YSXY in the treatment of oligoasthenospermia through network pharmacology,so as to lay a foundation for the study of pharmacodynamic mechanism in animal experiments.Animal experiments were designed to observe the effects of YSXY on sperm quality,spermatogenic cell apoptosis,proliferation and differentiation,as well as sperm energy metabolism in oligoasthenospermia model,and to elaborate its mechanism of action,so as to provide experimental exploration for YSXY formulation and mechanism of action.Methods:1.Network pharmacology predictionBy collecting and screening chemical components of YSXY in multiple databases of TCM ingredients and targets,the PPI network of YSXY action targets was constructed.Meanwhile,the target of oligoasthenospermia was searched and screened from the disease gene database.The disease gene was mapped to PPI network to obtain the key target of YSXY in the treatment of oligoasthenospermia.The TCM-component-target-module-pathway network of the source of YSXY was constructed to explore the potential mechanism of action of the medicine through enrichment analysis.2.Animal experimentCyclophosphamide-induced mice models of oligoasthenospermia were established.Semen analysis,electron microscope,flow cytometry and Western Blot method were used to test sperm quality,sperm ATP content,cell cycle,ultrastructure and protein expressions of Bcl-2,Bax,PI3K,Akt,SCF,C-Kit,ASK1 and JNK in the normal group,the model group,the YSXY high,medium and low dose groups,the clomiphen citrate control group,the Viagra control group,and the Huiren Shenbao control group.Rat models induced by Tripterygium glycosides were established.Semen analysis,ELISA,flow cytometry,RT-PCR method were used to test sperm quality,hormone level,mitochondrial membrane potential,testicular TGF-?1/Smads signal transduction pathway protein,VDAC2,VDAC3,CHK1 and CHK2 proteins expressions in the normal group,the model group,the YSXY high,medium and low dose groups,the clomiphen citrate control group,the Viagra control group,and the Huiren Shenbao control group.Results:1.Network pharmacologyA total of 418 chemical components,2447 targets and 134 pathogenic genes of oligoasthenospermia in YSXY were obtained.Among them,17 targets were the key targets of YSXY in the treatment of oligoasthenospermia,and they were involved in 8 key modules;According to the analysis of TCM-component-target-module-pathway network,57 components involved in the regulation of key targets and key modules,mainly involved in the apoptosis,proliferation and differentiation of spermatogenic cells and sperm energy metabolism..2.Animal experimentsCompared with the normal control group,models with oligoasthenospermia were successfully established.Experimental pharmacodynamics of mice showed that,YSXY could improve the sperm quality and sperm ATP content of oligoasthenospermia mice(P<0.01);Compared with the model group,the number of spermatogenesis cells and supporting cell mitochondria in the YSXY group increased significantly,with slight swelling.The normal form of Golgi apparatus could be seen and the ultrastructure of testis was improved significantly.Mechanism of action showed that YSXY could regulate the spermatogenic cell cycle and the protein expressions of Bcl-2,Bax,PI3K,Akt,SCF,C-Kit,ASK1 and JNK in testicular tissues of oligoasthenospermia mice(P<0.05).Experimental pharmacodynamics of rats showed that YSXY could improve sperm quality,sex hormone level and sperm mitochondrial membrane potential in oligoasthenospermia rats(P<0.05).Mechanism of action showed that YSXY could regulate the protein expressions of TGF-?1/Smads signal transduction pathway,VDAC2,VDAC3,CHK1 and CHK2 in testis of oligoasthenospermia rats(P<0.05).The biological pathway prediction related to network pharmacology was verified by animal experiments.Conclusion:1.YSXY can significantly improve the sperm quality and testicular ultrastructural damage in the model of oligoasthenospermia.2.YSXY can inhibit the apoptosis of spermatogenic cells in oligoasthenospermia model,and the specific mechanism may be related to the improvement of the protein expressions of Bcl-2,Bax,PI3K,Akt,SCF,C-Kit,ASK1 and JNK.3.YSXY can improve the spermatogenic function of testis and promote the proliferation of spermatogenic cells,the specific mechanism may be related to the improvement of the protein expressions of TGF-?1/Smads signal transduction pathway,CHK1 and CHK2.4.YSXY can enhance the sperm energy metabolism of oligoasthenospermia model,and the specific mechanism may be related to the regulation of mitochondrial membrane potential and the improvement of VDAC2 and VDAC3 protein expressions.In conclusion,this study confirmed the therapeutic effect of YSXY in tonifying kidney essence and promoting reproduction,and revealed the relevant mechanism of YSXY in the treatment of oligoasthenospermia by inhibiting the apoptosis of spermatogenic cells,promoting the proliferation of spermatogenic cells,and enhancing sperm energy metabolism. |
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