A Study On The Pathogenesis Of MDSCs And The Immunosuppressive Microenviroment Induced By MDSCs Promoting The Development Of Endometriosis

Object The purpose of this experiment is to explore whether MDSCs and its relative cytokines drive a immunosuppressive micro-environment in the peripheral blood and peritoneal fluid(PF)of the patients with endometriosis(EMT).Besides,to study whether the regulatory effect of Notch pathway on MDSCs can inhibit the development of EMT or not by constructing EMT model mice.MethodClinical researches:The peripheral blood and PF of 36 patients with endometriosis and20 patients in the control group were collected for analysis.The frequency and biological function of MDSCs(including polymorphonuclear(PMN)-MDSCs and monocytic(Mo)-MDSCs)were tested by flow cytometry technology.ELISA kits were applied in enzyme-linked immunosorbent assay to determine MDSCs-related cytokines of patient’s plasma and PF supernatant,including interferon(IFN)-γand chemokine(CC motif)ligand(CCL)2,CCL3,CCL4,CCL5.Animal experiments:SPF Babl/c mice were used to construct EMT model mice,which were divided into three groups(including CM,saline-EMT and DAPT-EMT).The PF,endomertium and ectopic lesions of mice were collected for experiments.Fluorescence microplate reader was used to detect the level of reactive oxygen species(ROS)in the PF,also eutopic endomertium and ectopic lesions of mice.Fluorescence quantitative PCR was used to detect the m RNA levels of Notch pathway related molecules in eutopic endomertium and ectopic lesions of mice,including ADAM17,Notch1,Jagged1and Hes1.The frequency of MDSC,regulatory T cells(Tregs)and T-helper(TH)-17cells were analyzed by flow cytometry.ResultsClinic researches:Data showed that significant elevations of blood Mo-MDSC and CCR5~+Mo-MDSCs were detected in EMT patients compared with control group,the frequencies of CCR5~+MDSCs and CCR5~+Mo-MDSCs in PF increased notably.Also,EMT patients displayed a significant higher production of plasma CCL3(P=0.046)and PF CCL3/5(P=0.042/0.036),the levels of PF CCL5 were elevated in III and IV staged EMT patients.Otherwise,the enrichment of CCR5~+MDSCs and CCR5~+Mo-MDSCs in PF had a positive correlation with the enhanced levels of CCL5 from EMT patients.Animals experiments:Compared with the control mice,the ROS level in the EMT mice was significantly higher,while the DAPT-EMT group was significantly lower than the saline-EMT group.ADAM17,Notch1,Jagged1 and Hes1 showed a significant increase in EMT mice,while the level of those m RNA in the DAPT-EMT group was significantly decreased compared to the saline-EMT group.The frequency of MDSCs and Tregs in the ectopic lesions of EMT mice was significantly higher than that of the CM,while the frequency of MDSCs and Tregs in the DAPT-EMT group was notably lower than that in the saline-EMT group.Meanwhile,the frequency of TH17 cells decreased in EMT mice,while in the DAPT group,the frequency showed a great rise.Conclusions(1)The amount of MDSCs displayed a significant increase in peripheral blood and PF of EMT patients;(2)The levels of CCL3 and CCL5 showed notable increases in PF of EMT patients,and the increase of CCR5~+MDSCs and CCR5~+Mo-MDSCs frequencies in PF were positively correlated with the concentration of CCL5;(3)A large number of MDSCs were recruited into the patient’s abdominal cavity microenvironment,which induces the local immunosuppressive microenvironment,thereby promoting the development of EMT;(4)Inhibiting Notch pathway may weaken the immunosuppressive function of MDSCs,thereby inhibiting the further development of EMT.