The Study Of Japanese Encephalitis Virus Induced Differentiation Of Myeloid-derived Suppressor Cells And CD3 Positive Macrophages

Japanese encephalitis virus(JEV)is one of the most important members of flavivirus family.It is a typical zoonotic pathogen,which has caused huge social and economic losses worldwide.In China,due to the vaccination,this disease has been effectively controlled among human.However,the breeding industry still suffers from incidence and economic loss.JEV's immune escape when entering the organism enabled the invasion of the central nervous system(CNS),leading to the inflammatory storm in CNS.The further study on the regulation of immune response by JEV will be pivotal to the treatment and prevention methods of JE.However,there is no detailed study about how JEV regulates the immune response and immune process.Our study on the infiltrated cells in the brain tissue of JEV infected mice showed that,the infiltrated cells were mainly M-MDSCs,which further differentiated into CD3~+macrophages.Myeloid derived suppressor cells(MDSCs)are the main regulatory cells in the immune system which is discovered in recent years and play a key role in many diseases as a group of heterogeneous cells.Therefore,the in-depth study of MDSCs in JEV infection model will be not only beneficial to reveal the internal mechanism of JEV and other flaviviruses in regulating the immune system,but also helpful in understanding MDSCs.Firstly,JEV P3 was injected into C57BL/6 mice via tail vein.Flow cytometry was used to analyze the infiltrating cell phenotypes in spleen and brain tissue after infection.The results indicated that JEV P3 infection induced increasing amount of M-MDSCs in spleen and the infiltrating cells in brain tissue are mainly composed of M-MDSC with partially differentiated CD3~+macrophages.The T cell proliferation inhibition experiment of M-MDSCs showed that JEV P3 induced M-MDSCs in spleen and brain could inhibit T cell proliferation through both ARG1 and i NOS.Secondly,we used splenectomy model and MDSCs elimination model by ATRA sustained-release method to explore the source of MDSCs and its impact on the pathogenicity of JEV.The results indicated that M-MDSCs induced by JEV P3 mainly derived from bone marrow and infiltrated into CNS through blood.At the same time,the MDSCs elimination significantly increased the survival rate of infected mice.Subsequently,the transcriptome analysis of MDSCs induced by JEV P3 demonstrated that the ZBP1-IRF7 signaling pathway is highly activated.The recombinant mutation virus infection,lentivirus overexpression experiments and in vitro experiment of IRF7 deficient mice indicated that NS1'and NS5 of JEV could induced M-MDSCs production which was mainly mediated by ZBP1-IRF7 signaling pathway.Finally,our analysis of the mediators inducing MDSCs infiltrated CNS and the differentiation conditions of CD3~+macrophages indicated that MDSCs could infiltrate into CNS through the chemokine CCL2/N-CCL2 derived from astrocytes and neurons,and differentiate into CD3~+macrophages under the guidance of M-CSF,IL-6 and IFN-?in the brain microenvironment.The transcriptome analysis of CD3~+macrophages showed that ZBP1-IRF7 signaling pathway also played an important role in the differentiation of CD3~+macrophages.In conclusion,this study investigated the generation of M-MDSCs induced by NS1'and NS5 proteins of JEV P3 through ZBP1-IRF7 signaling pathway,mediators inducing M-MDSCs infiltrated CNS and the differentiation conditions of M-MDSCs into CD3~+macrophages.These studies will contribute to the development and application of drugs to the treatment of Japanese encephalitis by targeting the immune system,and provide reference for other diseases in which MDSCs play a major role.