Research On DNA Methylation Of Imprinted Genes In Sperm Of Infertile Men

Objective Male infertility is a complex multifactorial disease.The main causes are endocrine dysfunction,reproductive system anatomical defects,chromosome abnormalities and gene mutations.There is still a high proportion of infertility unknown causes,called idiopathic infertility.Spermatogenesis is a complex process of cell division and differentiation.In this process,the DNA methylation pattern of imprinted genes will be cleared in the primordial germ cells and re-established according to gender,and maintained in mature sperm,so abnormal DNA methylation level of imprinted genes may affect spermatogenesis and maturation.At present,the relationship between abnormal DNA methylation level of imprinted genes and semen quality in patients with idiopathic infertility is a hotspot in the field of reproductive research.Therefore,this study explored the relationship between DNA methylation level of imprinted genes and sperm motility and sperm DNA integrity in idiopathic infertility patients by detecting DNA methylation level of imprinted genes.Methods Semen samples of male patients with idiopathic infertility who visited the reproductive medicine center of the first affiliated hospital of Anhui Medical University from November 2017 to May 2019 were collected.The sample of the same person was divided into two parts.One part was examined by CASA for sperm concentration and motility,and by SCSA for sperm DNA integrity.In the other part,the methylation level of the Cp G site in the imprinted gene promoter region was detected by Methyl Target after bisulfite treatment.The imprinted genes selected were paternal imprinted genes KCNQ1,MEG3 and maternal imprinted genes IGF-2,KCNQ1OT1,MEST and PEG3.According to the WHO laboratory manual for the examination and processing of human semen,the patients were divided into asthenozoospermia group(A1,74)and normal group(B1,92),according to the sperm DNA fragmentation index(DFI)divided into sperm DFI330% group(A2,47)and sperm DFI(27)30%(B2,119)group.DNA methylation levels of imprinted genes were compared between the two groups.Results(1).Compared with group B1,there were 2 Cp G sites with significant hypermethylation level in IGF-2 gene,1 Cp G site with significant hypomethylation level in KCNQ1 gene and 3 Cp G sites with significant hypomethylation level in MEST gene in group A1(P<0.05).There was no difference in the overall methylation levels of KCNQ1,MEG3,igf-2,KCNQ1OT1,MEST and PEG3 genes(P>0.05).(2).Compared with group B2,323 Cp G sites of IGF-2,KCNQ1OT1,KCNQ1,MEG3,MEST and PEG3 genes were detected,and 111 sites had significant hypomethylation levels in group B2(P<0.05).There were differences in the overall methylation levels of MEG3,IGF-2,MEST and PEG3(P<0.05),while there were no differences in the overall methylation levels of KCNQ1OT1 and KCNQ1(P>0.05).Conclusions Abnormal DNA methylation of some Cp G sites of imprinted genes IGF-2,KCNQ1 and MEST were found in sperm DNA of asthenospermia patients in idiopathic infertility population.The abnormal DNA methylation level of MEG3,IGF-2,MEST and PEG3 were found in sperm DFI 330%group.