CXCR3 signaling in glial cells ameliorates experimental autoimmune encephalomyelitis by restraining the generation of a pro-Th17 cytokine milieu and reducing CNS-infiltrating Th17 cells

Abstract

Background: Experimental autoimmune encephalomyelitis (EAE) is a mouse model of multiple sclerosis (MS). It has been shown that Th17 cells are critical for EAE pathogenesis. Mice lacking CXCR3 develop aggravated EAE compared with wild-type (WT) mice. This study investigated the effect of CXCR3 on Th17 expansion during EAE and further addressed the underlying mechanism. Methods: Both active EAE and adoptive-transfer EAE experiments were employed for studying EAE pathogenesis in WT and CXCR3-/- mice. Demyelination and leukocyte infiltration in the spinal cord of mice were analyzed by luxol fast blue staining and flow cytometry analysis, respectively. Glial cells expressing CXCR3 in the spinal cord were analyzed by immunofluorescence staining. Cytokine and chemokine levels in the spinal cord were analyzed using quantitative real-time PCR and enzyme-linked immunosorbent assay (ELISA). The glial cell line U87MG was employed for studying the CXCR3 signaling-mediated mechanism regulating Th17 expansion. Results: CXCR3-/- mice exhibited more severe EAE and had significantly increased central nervous system (CNS)-infiltrating Th17 cells compared with WT mice. Adoptive-transfer experiments showed that CXCR3-/- recipient mice that received Th17 cells polarized from splenocytes of myelin oligodendrocyte glycoprotein (MOG)-immunized CXCR3-/- mice or MOG-immunized WT mice always developed more severe EAE and had significantly increased CNS-infiltrating Th17 cells compared with WT recipient mice that received Th17 cells from the same origin. Furthermore, during EAE, the number of activated glial cells was increased in the CNS of MOG-immunized CXCR3-/- mice, and CXCR3-deficient glial cells expressed increased levels of cytokine genes required for Th17 expansion and recruitment. Finally, we found that extracellular signal-regulated kinase (ERK) activation elicited by CXCR3 signaling in U87MG cells attenuated the activation of NF-ΚB, a key transcription factor critical for the induction of IL-23 and CCL20, which are required for Th17 cell expansion and recruitment, respectively. Conclusions: This study demonstrates a previously unrecognized role of CXCR3 signaling in glial cells in negatively regulating Th17 cell expansion during EAE. Our results demonstrate that, in addition to its well-known role in the recruitment of immune cells, CXCR3 in CNS glial cells plays a critical role in restraining the pro-Th17 cytokine/chemokine milieu during EAE, thereby diminishing Th17 cell expansion in the CNS and suppressing disease development.