The Mechanisms Of Chemerin Contribute To Insulin Resistance Of Granulosa Cells In Polycystic Ovarian Syndrome

Background and Objective: The ovarian granulosa cells from polycystic ovary syndrome(PCOS)patients were liable to suffer from insulin resistance(IR)accounting for dysfunction in steroidogenesis and anovulation.Chemerin is an adipokine that serve an important role in inflammation,cell differentiation and glycolipid metabolism.However,the pathogenesis of IR of ovarian granulosa cells and the role of chemerin in PCOS remain unclear.In this study,we aim to investigate the relationship and mechanism of chemerin in the ovary with IR of human granulosa-lutein cells(h GLs)in PCOS.Methods: The expression of chemerin and its receptor CMKLR1 on granulosa cells was confirmed by PCR,western blot and immunofluorescence.The follicular fluid and serum of non-PCOS group,PCOS with IR group and PCOS without IR group were collected and their granulose cells were obtained for m RNA extraction.The concentration of chemerin in follicular fluid and serum were detected by immunoassay kit,and the abundance of chemerin m RNA in uncultured granulosa cells were measured by Real-time PCR.In vitro,cultured granulosa cells were treated with chemerin and si RNA which can knockdown the expression of chemokine-like receptor 1(CMKLR1).The effect of chemerin-CMKLR1 on IR in granulosa cells was detected by western blot,serosal sepharose chromatography and 18F-FDG marked glucose uptake assay.Besides,cultured granulosa cells were treated insulin to detect the effect of insulin on the expression of chemerin.Furthermore,Real-time PCR and western blot were used to detect the expression of autophagy-related genes and related proteins.The correlation between autophagy-related genes and clinical as well metabolic parameters of patients was analyzed to explore the role of autophagy in mediating chemerin-induced IR.Results:(1)The concentration of chemerin in follicular fluid and serum and the abundance of chemerin m RNA were significantly increased in granulosa cells of PCOS with IR;(2)Granulosa cells in PCOS patients owned a declined sensitivity to insulin stimulation.Increased ser307 phosphorylation of insulin receptor substrate 1(Irs1),diminished phosphorylation of Akt and weakend ability of glucose uptake were showed in granulosa cells of PCOS with IR;(3)Chemerin attenuated glucose transporter-4(GLUT4)translocation and diminished glucose uptake ability through down-regulating Akt phosphorylation via increasing ser307 phosphorylation of Irs1 in cultured h GLs;(4)The effects of chemerin on IR of cultured h GLs could be abolished by si RNA-mediated knockdown of chemokine-like receptor 1(CMKLR1);(5)Insulin promoted the expression of chemerin in a dose-dependent manner in cultured granulosa cells;(6)The abundance of autophagy related genes ATG5,ATG7,BECN1 m RNA and the ratio of autophagy marker protein light chain 3B II/I(LC3 II/I)were significantly increased whereas the abundance of the autophagy substrate SQSTM1/p62 was decreased in ovarian granulosa cells from PCOS patients;(7)There was a positive correlation between autophagy-related genes in granulosa cells and the level of serum basal testosterone,fasting insulin,the levels of chemerin in follicular fluid and granulosa cells,suggesting that autophagy may be related to chemerin-induced IR of h GLs in PCOS patients;Conclusion: Chemerin induced IR of human granulosa cells in PCOS,suggesting the potential contribution of chemerin to the pathogenesis of PCOS.