Study On The Mechanism Of Hyperoside In The Treatment Of Oligoasthenozoospermia

Background: Idiopathic Oligoasthenozoospermia refers to an unexplained reduction in sperm count and quality.In clinical treatment,idiopathic Oligoasthenozoospermia often has no definite effective treatment because of unknown etiology.At present,the exact mechanism of idiopathic Oligoasthenozoospermia is still unclear,which seriously affects China ’s fertility policy.With the advancement of sequencing technology and bioinformatics,omics research methods play an important role in the study of disease occurrence.Hyperoside is a flavonoid derived from Cuscuta sinensis,and many flavonoids have been used to improve reproductive function.In this study,a transcriptomic approach was used to elucidate underlying mechanism of idiopathic Oligoasthenozoospermia and the therapeutic effects of hyperoside on it as well as the mechanisms.Methods:(1)The gene expression profile dataset of GSE145467,containing 10 testicular tissue samples from patients with obstructive spermatogenic dysfunction and 10 testicular tissue samples from patients with non-obstructive spermatogenic dysfunction,was obtained from the public database GEO.The differentially expressed genes(DEGs)were screened by R language software.(2)Function enrichment analysis of genes and protein-protein interaction(PPI)network were performed on the obtained DEGs.(3)The model of Oligoasthenozoospermia was established by using chemotherapeutic drug cyclophosphamide(CTX),and the transcriptome of testicular tissue was sequenced before gene function enrichment analysis.(4)Hyperoside was used to treat CTX-induced Oligoasthenozoospermia model mice,and the therapeutic effect was evaluated from multiple perspectives and aspects,such as reproductive organ weight,sperm quality,serum hormone level,testicular pathological histomorphology.Meanwhile,combined with the results of DEGs gene function enrichment analysis,the accurate target and mechanism of hyperoside in the treatment of Oligoasthenozoospermia model mice were explored,which provided a theoretical basis for its clinical application.Results:(1)3518 DEGs were screened from the GSE145467 dataset,consisting of 2259 down-regulated genes and 1259 up-regulated genes.Gene ontology(GO)analysis revealed that the main biological processes enriched by down-regulated DEGs were mainly sperm differentiation,spermatogenesis and meiosis.The up-regulated DEGs were mainly enriched in biological processes such as the development of the visual system and the Metabolic processes for sulfur compounds.Enrichment of Kyoto Encyclopedia of Genes and Genomes(KEGG)pathways revealed that reduced DEGs were mainly enriched in ubiquitination-mediated proteolysis,tyrosine metabolism,pyruvate metabolism,progesterone-mediated oocyte maturation,homologous recombination and other pathways.The up-regulated DEGs were mainly enriched in pathways such as type I diabetes,toxoplasmosis,and steroid synthesis.(2)CTX was used to establish Oligoasthenozoospermia model mice.Final body weight,testis weight,testis index,and sperm parameters were significantly lower in the CTX group than in the control group.A total of 180 differential genes were screened from the transcriptome sequencing of testicular tissue,including 10up-regulated genes and 170 down-regulated genes.GO analysis revealed that the reduced DEGs are mainly enriched in meiotic cell cycle,homologous chromosome segregation,and spermatogenesis during meiosis.KEGG analysis revealed that the main pathways enriched by reduced DEGs included cell cycle,p53 signaling pathway,DNA replication,progesterone-mediated oocyte maturation,and homologous recombination.(3)The biological processes of GO analysis co-enriched by non-obstructive Oligoasthenozoospermia patients and Oligoasthenozoospermia model mice are mainly related to meiosis I(meiosis cell cycle,meiosis I,meiosis I cell cycle process,meiosis nuclear division,meiosis cell cycle process,nuclear division,organelle fission)and the late stage of sperm maturation(germ cell development,sperm development,sperm differentiation).(4)Through constructing a model of Oligoasthenozoospermia,the results showed that hyperoside could improve testis weight,testis index and sperm parameters(sperm concentration,sperm viability and sperm motility)(p<0.05).The results of H&E staining showed that there were more spermatogenic cells in the CTX+H group than in the CTX group,and the arrangement of germ cells at all levels was more compact and orderly.In terms of reproductive hormones in serum and testicular tissue: testosterone levels were significantly restored(p<0.05),FSH and LH levels did not change significantly,while testosterone production key proteins STAR,CYP17A1 and PRKACB were significantly up-regulated(p<0.05);meanwhile,RT-PCR and immunohistochemistry results suggested that key genes related to meiosis and sperm maturation The m RNA levels of Spata2,Hormad,Rad21,Atm,GM773 and Dmc1 were significantly up-regulated,and the protein levels of Rad21,Atm and Dmc1 were significantly up-regulated.Conclusion: The combined analysis of patients with Oligoasthenozoospermia and disease models showed that meiotic arrest of germ cells and spermatogenesis disorders played an important role in the development of Oligoasthenozoospermia.Hyperoside can improve CTX-induced Oligoasthenozoospermia by regulating testosterone production,meiosis and sperm maturation.This study helps to elucidate the mechanism of Oligoasthenozoospermia and provides a theoretical basis for the treatment of Oligoasthenozoospermia.