Title: An IL-23-STAT4 Axis Regulates Classical Dendritic Cell Expansion and Function in Experimental Autoimmune Encephalomyelitis
The signal transducer and activator of transcription 4 (STAT4) promotes protective immunity and autoimmunity downstream of pro-inflammatory cytokines including IL-12 and IL-23. In experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS), Stat4-/- mice are resistant to the development of inflammation and paralysis. Here, we examined cell-type requirements and found that in addition to T cells, STAT4 is required in dendritic cells for development of EAE. Deficiency of STAT4 in CD11c-expressing cells resulted in decreased T cell priming and inflammation in the CNS. EAE susceptibility was recovered following adoptive transfer of wild type bone marrow-derived DC to mice with STAT4-deficient DCs, but not adoptive transfer of STAT4- or IL-23R-deficient DCs. Single cell RNA-seq identified STAT4-dependent genes in DC subsets that paralleled a signature in MS patient DCs. Together, these data define a novel IL-23/DC/STAT4 pathway in DCs that could be a key to novel therapeutic targets in MS. Overall design: Mice from the two strains WT and Stat4fl/flCD11cCre were immunized for EAE. On day 15 post-immunization, mice were euthanized and mononuclear cells were recovered from the CNS of EAE mice. Cells were pooled from each group and stained with viability stain for 15 minutes,  preincubated with FC block for 10 minutes at 4ºC, followed by staining with fluorescent conjugated-antibodies for 30 minutes at 4ºC, and then washed with 1% BSA in PBS.  Cells were FACS sorted for all MHCII+ CD11c+ populations. Sorted cells suspension were partitioned into an emulsion of nanoliter-sized droplets using 10x Genomics Chromium Single Cell Controller