Human Blood Dendritic Cells Induce Tregs Through the PGE2-Independent Expression of an Active Form of IDO2 Enzyme

Abstract 1047

Indoleamine 2,3-dioxygenase (IDO) enzymes (IDO1 and IDO2) critically regulate the rate-limiting step of tryptophan degradation along kynurenine pathway in several cellular subsets, including dendritic cells (DCs). In particular, it is known that IDO1 expression in DCs is induced by inflammatory mediators, especially PGE2, and enhances DCs tolerogenic capacity, such as the induction of T regulatory cells (Tregs). In absence of exogenous stimuli, IDO1 is driven to proteasomal degradation by SOCS3. Conversely, few and contrasting results are available about IDO2 expression and function in human DCs.

We, then, besides IDO1, investigated the expression of IDO2 in human DC subsets, obtained from blood samples of healthy volunteers. Circulating myeloid and plasmacytoid DCs were purified and analyzed for the expression of both IDO enzymes.

Our data demonstrate that myeloid DCs express higher levels of both IDO1 and IDO2 mRNA in comparison to plasmacytoid DCs. Moreover, PGE2 modulates IDO1 and IDO2 mRNA expression only in myeloid DCs and has no effect in plasmacytoid DCs.

At protein level, IDO1 is expressed only in myeloid DCs and is modulated by PGE2, whereas IDO2 is expressed in both myeloid and plasmacytoid DCs and is not modulated by PGE2. Interestingly, when gene transcription or protein translation are inhibited, IDO2 protein expression persists independently from PGE2, while IDO1 expression requires PGE2 presence and needs continuous transcription and translation. Such discrepancy might be derived by a different binding of IDO1 and IDO2 to SOCS3, which in absence of pro-inflammatory stimuli is known to bind IDO1 and to drive it to proteasomal degradation. Indeed, SOCS3 immunoprecipitation and proteasomal inhibition experiments show that SOCS3 does not bind IDO2 which is not, therefore, driven to proteasomal degradation and is stably expressed.

At the functional level, both myeloid and plasmacytoid DCs generate Tregs through an IDO-dependent mechanism, since the addition of IDO inhibitors 1-MT-D and 1-MT-L abrogate Tregs generation. More specifically, the silencing of IDO2 through its specific siRNA downregulates kynurenine production and Tregs generation, thus demonstrating that IDO2 is functionally active in DCs.

In conclusion, our data demonstrate that, besides IDO1, human DCs also express IDO2, which contributes to tolerance by degrading tryptophan into kynurenine and by inducing Tregs. However, unlike IDO1, IDO2 expression is independent from inflammatory mediators, such as PGE2, and does not require continuous synthesis. These findings indicate IDO pathway as a critical check-point in the fine-tuning of DC-mediated tolerance induction. In particular, while IDO2 expression may confer a homeostatic tolerogenic capacity to DCs upon steady-state conditions, inflammation results in the induction of additional mechanisms of immune tolerance, such as the up-regulation of IDO1.