Both IL10– and IL12–Secreting Mature Dendritic Cells Induce Tolerance to Experimental Autoimmune Uveoretinitis (EAU) in C57BL/6 Mice

Abstract

Purpose: : Dendritic cells (DCs) determine the direction of an immune response: disease or tolerance. Previous studies in B10RIII mice have shown that immature bone marrow derived DC (BMDC) induce tolerance to EAU, whereas LPS–stimulated mature DC fail to demonstrate this tolerogenic effect. However, recent work has shown that the timing of TLR4 engagement by LPS is critical in determining the DC cytokine profile, and that IL10–secreting phenotypically mature early–LPS–activated DC induce tolerance to EAU. The aim of this study was to investigate how differentially LPS–activated DC induce EAU tolerance in C57BL/6 mouse.

Methods: : Freshly purified cultured C57BL/6 mouse BMDC were treated with LPS at 0 hr (early DC) or 20 hrs (late DC). CD4⁺CD25^+high T regulatory cells (Tregs) were isolated from the draining cervical lymph nodes three days after inoculation with peptide–loaded early DC. IL10/IL12 production was analysed by multiplex cytokine bead assay (CBA). In vivo cell trafficking studies were done using cells from GFP–Tg mice and detected using flow cytometry.

Results: : Adoptive transfer of both IL10– (early) and IL12– (late) producing DC induced tolerance to IRBP–induced EAU. In addition, early DC administration significantly expanded Tregs in the draining cervical lymph nodes, and adoptive transfer of early DC–induced Tregs also suppressed EAU. Tracking studies of intraperitoneally inoculated peptide–loaded early DC–induced GFP Tregs revealed that Tregs concentrated in the inguinal lymph node and the spleen, but not in the eye. In addition, Tregs derived from inguinal lymph node from early DC treated mice expanded specifically to IRBP in vitro.

Conclusions: : We propose that since both IL10– and IL12– secreting mature DC suppress EAU in the C57BL/6 mouse, DC Cytokine production alone does not determine tolerogenic function. Our data suggest that early DC induce EAU tolerance via a mechanism that involves the expansion of CD4⁺CD25⁺ Tregs at the draining cervical lymph node and that these Tregs home to secondary lymphoid tissue where they are available to act in regional draining sites of inflammation.

Keywords: autoimmune disease • uveitis-clinical/animal model • immune tolerance/privilege