Chemerin/CMKLR1 Signaling In Ovarian Steroidogenesis And Luteolysis

Corpus Luteum(CL) mainly secrets progesterone which plays an essential role in maintenance reproductive function of mammals. Mechanism of luteolysis is the focus of reproductive biology and pharmacology. Chemerin is a novel cytokine associated with obesity and metabolic syndrome and a growing number of studies have proved its some regulation effects on gonadal hormone secretion. Some studies also proved that cytokines were involved in luteolysis associated with macrophages. There is no literature reported the function of Chemerin/CMKLR1 in ovarian steroidogenesis and luteolysis.Therefore, this study used C57 wild type mice, CMKLR1 gene knockout mice and in vitro luteal cells & macrophages co-culture models to detect CMKLR1 morphology localization and compare the level of progesterone and estradiol between WT/CMKLR-/- and research Chemerin/CMKLR1 pathway whether plays roles in follicle development, CL development and luteolysis. The following results were obtained:1. Expression and localization of CMKLR1 in ovary and uterus(1) Ovary: RT-q PCR results showed that Chemerin and CMKLR1 were expressed in the ovaries of different estrus cycle of mice, and the main expression stages were estrus and metestrus associated with follicle development and corpus luteum formation. IHC results showed that CMKLR1 were mainly localized in theca cells and follicular clearance. CMKLR1 significantly increased after PMSG-48 h treated. All above indicated that Chemerin/CMKLR1 signaling pathway may have certain regulation effects on follicule development and luteal function. In addition, F4/80 was also observed in dioestrus ovaries. It suggested CMKLR1 may be involved in the degradation process of physiological function of CL.(2) Uterus: IHC results showed that CMKLR1 was expressed in different estrous cycle of female uterus, especially in the longitudinal and circular muscles down by endometrium. Since uterus is the mammals PGF2 alpha source during corpus luteum degradation period. It suggested that Chemerin was involved in CL degeneration.2. CMKLR1 in ovarian steroidogenesisProgesterone secretion level of superovulation serum, follicle culture and corpus luteum culture were suppressed by Chemerin via CMKLR1 and the level recovered from the blocking of CMKLR1. Besides, estradiol level was overall within the trend of progesterone. The recovery of progesterone and estradiol level in CL culture were limited after CMKLR1 was blocked, which indicated that there may be other pathways involved in the steroidogenesis suppression.3. CMKLR1 in luteolysis(1) PGF2 alpha induced destruction of CL progesterone secretion was suppressed while CMKLR1 was blocked. Basal progesterone level of CMKLR1 KO group was obviously higher than WT. Alpha-Neta could not obviously improve the progesterone damage of CMKLR1 KO group. Estradiol level was restored after CMKLR1 blocked, which confirmed the inhibitory effects of Chemerin on estradiol secretion.(2) The lower basal level of progesterone/estradiol secretion co-culture could be relieved by blocking of Chemerin/CMKLR1 which demonstrated that blocking of Chemerin/CMKLR1 could remove the inhibition of macrophages on LCs progesterone/estradiol secretion.(3) After the inhibition of Chemerin/CMKLR1 pathway, Caspase-3, Caspase-9 declined and cytokines IL-beta and IL-6 also decreased, which indicated that macrophages may destroyed LCs via this pathway. However, the opposite regulatory direction of TNF alpha, which TNF alpha mRNA level increased after Chemerin/CMKLR1 blocked, indicating that Chemerin is a key factor triggering TNF alpha and its principle may have nothing to do with Chemerin/CMKLR1 pathway.The above results indicated that it could suppress secretion of progesterone and estradiol via Chemeing/CMKLR1 pathway during follicle development and CL development. And there may be other receptors involved in the suppression. In addition, CMKLR1 could mediate the destruction of the corpus luteum and luteolysis process by macrophages.