Mechanism of microRNA-22-3p of extracellular vesicles derived from bone marrow mesenchymal stem cells in inhibiting damage of ovarian granulosa cells induced by cyclophosphamide
Objective To analyze the effect of microRNA-22-3p(miR-22-3p)of extracellular vesi-cles derived from bone marrow mesenchymal stem cells on premature ovarian failure.Methods Follicular fluids were provided by premature ovarian failure patients with in vitro fertilization or the second-gen-eration in vitro fertilization treatment and normal volunteers with the same treatment for infertility due to male factors;the exosomes derived from bone marrow mesenchymal stem cells were isolated by dif-ferential centrifugation;the morphology,particle size and marker proteins of isolated exosomes were analyzed by transmission electron microscopy,NanoSight LM10 analyzer and Western blotting.Granulosa cells were treated with cyclophosphamide(CTX),exosomes,miR-22-3 mimics and corresponding controls,and were divided into CTX group,control group,exosome incubation group,PBS treat-ment group,CTX+miR-22-3p group,CTX+miR-NC group,CTX+Exosomes group,and CTX+PBS group;gene expression was detected by Western blotting and quantitative real-time polymerase chain reaction(qRT-PCR).The 3-(4,5-Dimethylthazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)reagent and flow cytometry were used to detect cell viability and apoptosis.Results The extracted exosomes had typical goblet vesicles,the exosome marker proteins C cluster of differentia-tion 63(CD63)and C cluster of differentiation 9(CD9)were expressed in the extracted exosomes,and exosome particle size was 80 to 150 nm;miR-22-3p was significantly lowly expressed in patients with premature ovarian failure and ovarian granulosa cells induced by CTX(P＜0.05),but was significantly highly expressed in ovarian granulosa cells incubated with exosomes derived from bone marrow mesenchymal stem cells(P＜0.05);the activity of granulosa cells in the CTX group was significantly lower than that in the control group,but was significantly higher in the CTX+miR-22-3p group or CTX+Exosomes group than that in the control group(P＜0.05);the apoptosis rate of granulosa cells in the CTX group was significantly higher than that in the control group,but was sig-nificantly lower in the CTX+miR-22-3p or CTX+Exosomes group than that in the control group(P＜0.05).Conclusion The miR-22-3p of extracellular vesicles derived from bone marrow mes-enchymal stem cells can inhibit ovarian granulosa cell injury induced by CTX.
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维普
