Negatively Regulation Role Of GPR30 In Inflammation Induced By MSU/Function And Mechanism Of IFN-γ In Regulating PDL1 Expression In Fibroblast

Objective:Gout is the most common inflammatory arthritis induced by monosodium urate crystal(MSU)precipitation.The incidence of gout attack was significantly higher in men than woman,indicating the important role of estrogen system in pathogenesis of gout,but the specific mechanisms underlying remained to be explored.G protein couple receptor 30(GPR30),the newly defined estrogen receptor,had been proved participated in regulating inflammation in some diseases.As different estrogen receptors paly different role in inflammation regulation,how GPR30 was involved in the inflammation induced by MSU in gout was still unknown.This study was performed to determine the role of GPR30 in inflammation induced by MSU and the potential mechanisms.Methods:We used G-1(The agonist of GPR30)to study the function of GPR30 in inflammation induced by MSU in vitro and in vivo.We further investigated expression of the TLR and NLRP3,which are the key receptor involved in pathogenesis of gout.Reactive oxygen species(ROS)and cleaved-caspase-1 expression were also detected to confirm the regulation role in NLRP3 pathway.Seahorse analysis was used to detect the metabolism profile of macrophages effected by G-1 to investigate the mechanism of GPR30 in inflammation.The relative expression of GPR30 were detected according to the inflammation state to confirm the role of GPR30 in gout.Results:Negative regulation role of G-1 in IL-1β expression and NLRP3 expression were found both in vitro and in vivo.Moreover,the negative regulation role of G-1 in ROS production and NLRP3 as well as cleaved-caspase-1 expression were also found in our study.Our data also showed that G-1 inhibited aerobic glycolysis in LPS activated macrophages,which might be responsible for IL-1β and NLRP3 expression.High expression level of GPR30 in gout patients who are inflammation remission state were also found.Conclusions:Our data suggest that GPR30 was involved in the negative inflammation regulation induced by MSU and high expression of GPR30 might contribute to part of the mechanism of inflammation remission of gout.Objective: Fibroblasts are the major cells that participated in the tissue microenvironments architecture by depositing and remodeling extracellular matrix components.Fibroblasts have been proved played key roles in setting up,supporting or suppressing the immune response and involved in diseases,like inflammation,autoimmune diseases and cancers.Different subsets of fibroblasts have been identified.Researches had proved that;different environment and different stimulation affected the fibroblasts function in immune regulation.It is still havingsome controversial of the functions of fibroblasts in T cells.In this project,we focus the functions of IFN-γ in fibroblast and its effects with T cell responses.The cell signal and the metabolism changes stimulated by IFN-γ were also discussed.Methods: IFN-γ were used to stimulate the fibroblast in vitro to detect the PDL1 expression.Normal fibroblasts and the fibroblasts stimulated with IFN-γ were used to co-culture with activated T cell to detect the proliferation and function of T cells.IFN-γ antibodies were added in the co-culture system of fibroblasts and T cells to further proved the function of IFN-γ in PDL1 expression.Psoriasis mouse model were produced to prove IFN-γ enhanced the negative regulation role of fibroblasts in inflammation.Western blot was used to detect the cell signal activation and the specific inhibitor were added to further prove the cell signal in fibroblasts.Seahorse analysis were performed to describe the metabolism changes affected by IFN-γ and the inhibitor of metabolism were used to further confirmed.Results: IFN-γ increased the PDL1 expression efficiently and IFN-γ enhanced the functions of fibroblasts in suppressing T cell proliferation.Inhibited the IFN-γ with its antibody significantly reduced the PDL1 expression and inhibited the effects of fibroblasts in T cell proliferation.Moreover,IFN-γ enhanced the function of fibroblast in psoriasis inflammation relieving.IFN-丫 activated the STAT1,ERK and Akt cell signal and the inhibitors of these cell signal reduced the PDL1 expression stimxxlated by IFN-γ.2DQ the inhibitor of glycolysis significantly reduced the PDL1 expression induced by IFN-γ.Conclusions: IFN-γ induced the high expression of PDL1 in fibroblasts and it can enhance the suppression role in T cell proliferation of fibroblast.Moreover,IFN-γ regulated the glycolysis and activated the STAT1 cell signal and participated the PDL1 expression.This project was aimed to further discuss the immune regulation roles of fibroblasts and provided new insight of inflammation and cancer therapy.