Effect Of Acupuncture On Methylation DNA For Mouse Ovary Tissue With Poor Ovary Response

Objective:This study established a model with poor ovarian response(POR)in vitro fertilizationembryo transfer(IVF-ET)technique to observe the effects of acupuncture on ovarian function and ovarian response in mice,and methylation capture sequencing technique was used to explore the effect of acupuncture on DNA methylation status and differential methylation genes in ovarian tissue.Methods:After selecting the normal estrous cycle mice,30 female C57BL/6N mice aged 7-8weeks with normal estrous cycle were divided into three groups: blank group(n=10),model group(n=10)and acupuncture group(n=10).After intragastric administration of Tripterygium wilfordii polyglycoside tablet suspension or 0.9% normal saline for 14 days,acupuncture intervention was performed for 10 days.Then PMSG and HCG were injected intraperitoneally to induce ovulation.Finally,blood,fallopian tube and ovary were taken and the whole genomic methylation of ovarian tissue was sequenced.Results:1.Number of retrieved oocytes: Compared with the blank group,the number of retrieved oocytes in the model group decreased significantly,and there was no significant difference with the acupuncture group;compared with model group,the number of retrieved oocytes in the acupuncture group increased,and the difference was statistically significant(P<0.05).2.Ovarian wet weight and ovarian index: compared with the blank group,the ovarian wet weight and ovarian index decreased in the model group(P<0.05),and there was no significant difference in the ovarian wet weight and ovarian index in the acupuncture group(P>0.05);compared with the model group,the wet weight and ovarian index of mice in the acupuncture group showed an upward trend,but there was no statistical difference(P>0.05).3.Sex hormone and AMH: Compared with the blank group,AMH and E2 in the model group and the acupuncture group both decreased significantly,while the FSH increased significantly,and the difference was statistically significant(P<0.05).Compared with the model group,the AMH and E2 in the acupuncture group had an upward trend and FSH had a downward trend,but there was no significant difference between the model group(P>0.05).4.Methylation sequencing:(1)Overall methylation level: compared with the blank group,the overall methylation level of the model group was not significantly different from that of the blank group,while compared with the model group,the overall methylation level of the acupuncture group was mainly down-regulated.(2)GO enrichment analysis: The differentially methylated genes screened by the model group and the blank group are mainly enriched in the functions of FSH hormone secretion,negative regulation of androgen receptor signaling pathway,and activation of endothelial cells.The differentially methylated genes screened by the model group and acupuncture group were mainly enriched in activin binding,kinase inhibitory activity,negative regulation of neurite regeneration.(3)KEGG enrichment analysis: The differentially methylated genes screened by the model group and the blank group are mainly enriched in endocrine resistance,vitamin B6 metabolism,tyrosine metabolism,stem cell diversity regulation,relaxation signal and other pathways.The differentially methylated genes screened by the model group and acupuncture group were mainly enriched in ovarian steroid hormone production,vitamin metabolism and absorption,thiamine metabolism,hippo signal and other pathways.(4)Gene selecting: After further screening,through differential methylation region analysis differentially methylated gene TGFBR3 and PI3K/ Akt signal pathway was obtained.Conclusions:1.Acupuncture can improve the ovarian function of mice with poor ovarian response,mainly by increasing the number of retrieved oocytes and improving the wet weight and ovarian index of mice.2.The therapeutic effect of acupuncture on mice with poor ovarian response may be achieved through down-regulation of the methylation level of differential genes,which is closely related to down-regulation of the methylation level of TGFBR3 gene and PI3K/ Akt signaling pathway.