EIF4A3-induced Exosomal CircLRRC8a Alleviates Granulosa Cells Senescence Via MiR-125a-3p/NFE2L1 Axis

BackgroundPremature ovarian failure(POF)is an important cause of female infertility and seriously impacts the physical and psychological health of patients.Mesenchymal stem cell-derived exosomes(MSCs-Exos)play a crucial role in the treatment of reproductive disorders,especially POF.Bone marrow mesenchymal stem cells(BMSCs)are easily obtained,and the role of exosomes derived from BMSCs in premature ovarian failure has become the focus of research.However,the biological functions and therapeutic mechanisms of MSCs-Exos circular RNA(circ RNA)in POF remain to be determined.We found that circ LRRC8 A was down-regulated in POF vivo models,peripheral blood of POF patients and ovarian granulosa cells(GCs)by bioinformatics analysis and functional assays and served as a key molecule for BMSCs-Exos-mediated ovarian function repair.Mechanistic studies showed that circ LRRC8 A endogenously adsorbed mi R-125a-3p and down-regulated NFE2L1 expression,and eukaryotic initiation factor 4A-III(EIF4A3),a pre-m RNA splicing factor,promoted circ LRRC8 A cyclization and expression by directly binding to LRRC8 A m RNA transcripts.Notably,EIF4A3 silencing reduced circ LRRC8 A expression and attenuated the therapeutic effect of BMSCs-Exos on oxidatively damaged GCs.By elucidating the role and mechanism of circ LRRC8A-rich Exos-mediated mi R-125a-3p/NFE2L1 axis regulation to inhibit senescence and repair ovarian function in GCs,this study demonstrates a novel therapeutic pathway for cellular senescence protection against oxidative damage and paves the way for establishing cell-free therapeutic approaches for POF.In addition,circ LRRC8 A is expected to be a circulating biomarker for POF diagnosis and prognosis.MethodsIn vivo and in vitro models of POF were established and identified.BMSCs-Exos were extracted,isolated and identified.Bioinformatics methods were used to screen and identify differential circ RNAs,mi RNAs associated with POF.q RT-PCR was used to detect circ RNAs,mi RNAs expression.Reactive oxygen species(ROS)staining assays and SA-β-gal staining assays were used to assess the levels of oxidative stress and senescence in GCs.The role of circ LRRC8A/mi R-125a-3p/NFE2L1 axis in POF was analyzed by rescue assay,nucleoplasmic isolation assay,dual luciferase reporter gene assay,RNA pull-down and Western blot assay.The regulatory role of EIF4A3 on circ LRRC8 A biosynthesis was examined by RIP and other assays.Results1.BMSCs-Exos enhances the antioxidant damage capacity of GCs in vitro and repairs ovarian function in POF rats.2.circ LRRC8 A expression was downregulated in serum and GCs of POF patients as well as in vivo and in vitro POF model.3.BMSCs-Exos repair ovarian function and resist oxidative damage by GCs in POF rats through the release of circ LRRC8 A.4.Circ LRRC8 A upregulates NFE2L1 and inhibits senescence of GCs through sponge adsorption of mi R-125a-3p.5.EIF4A3 promotes circ LRRC8 A cyclization and biogenesis.ConclusionCirc LRRC8 A expression was downregulated in serum and GCs of POF patients as well as in vivo and in vitro models of POF.BMSCs-Exos could resist oxidative damage in GCs and repair ovarian function in POF rats by releasing circ LRRC8 A,inhibiting mi R-125a-3p,upregulating NFE2L1,and EIF4A3 could promote cyclization and biogenesis of circ LRRC8 A,and Exos-circ LRRC8 A is expected to be a new target for POF diagnosis and treatment.