The Regulation Of MiR-223-3p/NLRP3 Signaling Pathway By Morinda Citrifolia Inhibits The Inflammatory Response Of PCOS Rats

Objective:In this experiment,We use the ultra-performance liquid chromatography tandem mass spectrometry(UPLC-MS/MS)to identify the chemical composition of Morinda citrifolia,and though animal experiment verify the mi R-223-3p/NLRP3 signaling pathway inhibit the inflammatory response on rats with polycystic ovary syndrome(PCOS),in order to prove the potential effect and mechanism of Morinda citrifolia in the PCOS.Methods: 1.UPLC-MS/MS analyze the effective chemical composition of Morinda citrifolia.2.Animal experiments: SD female rats were randomly divided into three groups: normal control group(n=5),PCOS model group(n=5)and Morinda citrifolia gavage group(n=5).HE staining and electron microscopy observe the morphology of ovarian histology,ELISA method detect the expression levels of T,LH,FSH,INS in serum and IL-1β and IL-18 in follicular fluid,RT-q PCR detect the m RNA expression of mi R-223-3p,Caspase-1 and NLRP3 in ovarian tissue,and Western blot detect the protein expression of Caspase-1 and NLRP3 in ovarian tissue.Results:1.UPLC-MS/MS results: There are 641 chemical composition,including 387 positive ion composition and 254 negative ion composition.Combined with literature,luteolin,Apigenin and emodin involve the occurrence and development of PCOS.2.Animal experiment results:(1)Morphological changes in rat ovarian tissues: The ovaries of normal control group rat can be seen all levels of developing follicles,granule cells are multilayered and normal corpus luteum;In the PCOS model group,the number of ovary granule cell layers and the corpus luteum decreased,follicular cystic dilation,increased atresia follicles and vanished oocytes or radial crowns.The proportion of follicles at all levels is normal in the Morinda citrifolia gavage group,and the number of corpus luteum increased.(2)ELISA test results:(1)Compared with the normal control group,the serum T in the PCOS model group is elevated(P<0.01),and increased LH and INS(P<0.05),FSH decreased(P<0.05),increased IL-18(P<0.05),increased IL-1β(P<0.01).(2)Compared with the normal control group,in the Morinda citrifolia gavage group,serum T is elevated(P<0.05),LH,FSH,INS are not differentiated(P>0.05),elevated IL-1β(P<0.01),and undifferentiated IL-18(P>0.05).(3)Compared with the PCOS model group,T,LH,IL-18,IL-1β decreased(P<0.05),INS decreased(P<0.01),and FSH increased(P<0.05)in the Morinda citrifolia gavage group.The results showed that Morinda citrifolia could reduce the levels of T,LH,INS and inflammatory factors IL-18,IL-1βand increase the level of FSH in PCOS rats.(4)RT-q PCR results:(1)Compared with the normal control group,the mi R-223-3p m RNA of rat ovarian tissue in the PCOS model group is reduced(P<0.01),and NLRP3 m RNA is increased(P<0.01),and Caspase-1 m RNA is significantly increased(P<0.0001).(2)Compared with the normal control group,in the Morinda citrifolia gavage group,Caspase-1 m RNA is elevated(P<0.05),mi R-223-3p and NLRP3 is insignificant(P>0.05).(3)Compared with the PCOS model group,mi R-223-3p m RNA is elevated(P<0.05),and NLRP3 m RNA is decreased(P<0.05),and Caspase-1 m RNA is significantly reduced(P<0.0001)in the Morinda citrifolia gavage group.The results showed that Morinda citrifolia is able to reduce the NLRP3 and Caspase-1 m RNA expression and increase the expression level of mi R-223-3p in PCOS rats.(5)Western blot results:(1)Compared with the normal control group,the expression of NLRP3 protein in the ovarian tissue in the PCOS model group is increased(P<0.01),and the protein expression of Caspase-1 is significantly increased(P<0.001).(2)Compared with the normal control group,the protein expression of Caspase-1 is elevated(P<0.05))and the protein expression of NLRP3 is not differentiated(P>0.05)in the Morinda citrifolia gavage group.(3)Compared with the PCOS model group,the protein expression of NLRP3 in the ovarian tissue in the Morinda citrifolia gavage group is reduced(P<0.01)and the protein expression of Caspase-1 is significantly reduced(P<0.001).The results showed that Morinda citrifolia is able to reduce the protein expression of NLRP3 and Caspase-1 in PCOS rats.Conclusion:1.Morinda citrifolia may involve the occurrence and development of PCOS through a variety of active element.2.Animal experiments verify the Morinda citrifolia may regulate the mi R-223-3p/NLRP3 signaling pathway to inhibat the chronic low-grade inflammatory response of PCOS rats,and to improve the occurrence of PCOS.