Topographical and Biological Evidence Revealed FTY720-Mediated Anergy-Polarization of Mouse Bone Marrow-Derived Dendritic Cells In Vitro

Abnormal inflammations are central therapeutic targets in numerous infectious and autoimmune diseases. Dendritic cells (DCs) are involved in these inflammations, serving as both antigen presenters and proinflammatory cytokine providers. As an immuno-suppressor applied to the therapies of multiple sclerosis and allograft transplantation, fingolimod (FTY720) was shown to affect DC migration and its crosstalk with T cells. We posit FTY720 can induce an anergy-polarized phenotype switch on DCs in vitro, especially upon endotoxic activation. A lipopolysaccharide (LPS)-induced mouse bone marrow-derived dendritic cell (BMDC) activation model was employed to test FTY720-induced phenotypic changes on immature and mature DCs. Specifically, methods for morphology, nanostructure, cytokine production, phagocytosis, endocytosis and specific antigen presentation studies were used. FTY720 induced significant alterations of surface markers, as well as decline of shape indices, cell volume, surface roughness in LPS-activated mature BMDCs. These phenotypic, morphological and topographical changes were accompanied by FTY720-mediated down-regulation of proinflammatory cytokines, including IL-6, TNF-α, IL-12 and MCP-1. Together with suppressed nitric oxide (NO) production and CCR7 transcription in FTY720-treated BMDCs with or without LPS activation, an inhibitory mechanism of NO and cytokine reciprocal activation was suggested. This implication was supported by the impaired phagocytotic, endocytotic and specific antigen presentation abilities observed in the FTY720-treated BMDCs. In conclusion, we demonstrated FTY720 can induce anergy-polarization in both immature and LPS-activated mature BMDCs. A possible mechanism is FTY720-mediated reciprocal suppression on the intrinsic activation pathway and cytokine production with endpoint exhibitions on phagocytosis, endocytosis, antigen presentation as well as cellular morphology and topography

NFAT primes the human RORC locus for RORγt expression in CD4 + T cells

International audienceT helper 17 (Th17) cells have crucial functions in mucosal immunity and the pathogenesis of several chronic inflammatory diseases. The lineage-specific transcription factor, RORγt, encoded by the RORC gene modulates Th17 polarization and function, as well as thymocyte development. Here we define several regulatory elements at the human RORC locus in thy-mocytes and peripheral CD4 + T lymphocytes, with CRISPR/Cas9-guided deletion of these genomic segments supporting their role in RORγt expression. Mechanistically, T cell receptor stimulation induces cyclosporine A-sensitive histone modifications and P300/CBP acetylase recruitment at these elements in activated CD4 + T cells. Meanwhile, NFAT proteins bind to these regulatory elements and activate RORγt transcription in cooperation with NF-kB. Our data thus demonstrate that NFAT specifically regulate RORγt expression by binding to the RORC locus and promoting its permissive conformation