Dendritic cells expressing ectopic IL-35Ig induce IDO-related tolerance in vivo.

Dendritic cells (DCs) are specialized APCs playing a central role in activating, but also in regulating, T-cell responses, their dual role depending on both their default presentation profile and the superimposed environmental stimuli, which may reprogram or subvert DC function. In previous studies, we demonstrated that this functional plasticity can be exploited to trigger and amplify a tolerogenic state (‘infectious tolerance’) via suppressive molecules (i.e., CTLA4-Ig, GITR-Ig, and TGF-beta) that will convert otherwise immunogenic DCs into tolerogenic cells. Here we focused on the possible modulation of immune responses by ectopic expression of IL-35 in DCs, a potent suppressive cytokine produced by T regulatory cells. We transfected immunogenic CD8– DCs with a single-chain IL-35-Ig gene construct and we found that, after P815AB antigen peptide pulsing, even a minority fraction of transfected cells inhibited the immunogenic presentation of the peptide in a skin test assay, triggering the onset of a long-lasting, specific tolerance subverting the otherwise immunogenic priming to the peptide. This suppressive effect was attributable to changes occurring in vivo in total splenocytes and in their CD11c+ DC fraction after DC/IL-35-Ig/P815AB injection, including induction of tolerogenic indoleamine 2,3-dioxygenase-1 (IDO1) and production of mediators sustaining the mechanism of infectious tolerance (via the IDO1 metabolite kynurenine, TGF-β, and IL-10). On the basis of these findings, we attempted a tolerogenic vaccination protocol to prevent autoimmune diabetes. Interestingly, on treating prediabetic NOD mice with DCs overexpressing IL-35 and presenting the IGRP peptide (one of the most relevant type 1 diabetes self-antigens), we observed delayed and less severe onset of hyperglycemia. In conclusion, DCs overexpressing ectopic IL-35Ig exert an important suppression in vivo, which, on combination with specific antigen, might represent a powerful, selective means of negative vaccination in autoimmune diseases.