Effect of moxibustion on expression profile of miRNAs in Tripterygium glycoside-induced decreased ovarian reserve

OBJECTIVE:To explore the possible regulatory mechanism of microRNA(miRNA)in moxibustion treatment for decreased ovarian reserve(DOR).METHODS:The DOR model was constructed by intragastrical Tripterygium glycoside suspension administration,and moxibustion therapy was simultaneously given.The morphological ovarian changes were observed by hematoxylin and eosin staining.The miRNA expression profile was detected by RNA sequencing,and bioinformatics analysis was performed.Cytoscape software 3.6.1 was used to establish a regulatory network and differentially expressed miRNAs were verified by reverse transcription quantitative polymerase chain reaction(RT-qPCR).RESULTS:Decreased number of mature follicles,increased atresia follicles,and abnormal granulosa cell morphology were observed in the model group compared with the control group.The moxibustion group demonstrated increased mature follicles,decreased atretic follicles,and significantly decreased abnormal morphology of granulosa cells compared with the model group.Additionally,RNA sequencing results manifested significantly up-regulated miRNA expressions(miR-92b-3p,miR-26-5p_R+1_1ss10TC,miR-206-3p,miR-9993b-3p_1ss6GA,miR-7857-3p_R-1,miR-219a-2-3p_1ss10GC,miR-3968-p5_1ss10AT,and PC-5p-6478_1795)and down-regulated miR-664-2-5p_R+1 in the model group,compared with the control group,and the moxibustion group reversed abnormal disorder levels of these miRNAs.Moreover,these differentially expressed miRNAs were mainly involved in the phosphatidylinositol-3-kinase/protein kinase B signaling pathway and nuclear factor erythropoietin-2-related factor 2/heme oxygenase 1 signaling pathway.Finally,network and RT-qPCR verification revealed miR-9993b-3p_1ss6GA as the most critical miRNA.CONCLUSION:This experiment proved the effectiveness of moxibustion in improving the ovarian reserve of rats by regulating miRNA expression,especially miR-9993b-3p_1ss6GA.