Natural agonists for aryl hydrocarbon receptor in culture medium are essential for optimal differentiation of Th17 T cells

Abstract

Th17 cell differentiation is dependent on interleukin (IL)-6 and transforming growth factor (TGF)-␤ , and it is modulated by activation of the aryl hydrocarbon receptor (AhR). In this study, we show that differentiation of Th17 cells, but not Th1 or induced regulatory T (iT reg) cells, is increased by endogenous AhR agonists present in culture medium. Th17 development from wild-type mice is suboptimal in the presence of the AhR antagonist CH-223191, similar to the situation in AhR-defi cient mice, which show attenuated IL-17 production and no IL-22 production. The presence of natural AhR agonists in culture medium is also revealed by the induction of CYP1A1, a downstream target of AhR activation. However, the most commonly used medium, RPMI, supports very low levels of Th17 polarization, whereas Iscove ' s modifi ed Dulbecco ' s medium, a medium richer in aromatic amino acids, which give rise to AhR agonists, consistently results in higher Th17 expansion in both mouse and human cells. The relative paucity of AhR agonists in RPMI medium, coupled with the presence of factors conducive to IL-2 activation and enhanced Stat5 phosphorylation, conspire against optimal Th17 differentiation. Our data emphasize that AhR activation plays an essential part in the development of Th17 cells and provide a rational explanation for the poor in vitro polarization of Th17 cells that is reported in the majority of publications for both mouse and human cells.

Activation of the aryl hydrocarbon receptor reveals distinct requirements for IL‐22 and IL‐17 production by human T helper cells

“…Interestingly, when the AHR antagonist was added to the cultures in the absence of TCDD, we observed a significantly reduced production of IL-22 and a trend toward an increased production of IL-17A. These results not only show that AHR is involved in TCDD-induced regulation of IL-22 and IL-17A but also, consistent with previous observations [28], suggest that natural AHR ligands present in the cells and/or in the external milieu exert an important biological activity capable of regulating the production of IL-22 and IL-17A.…”

“…Interestingly, when the AHR antagonist was added to the cultures in the absence of TCDD, we observed a significantly reduced production of IL-22 and a trend toward an increased production of IL-17A. These results not only show that AHR is involved in TCDD-induced regulation of IL-22 and IL-17A but also, consistent with previous observations [28], suggest that natural AHR ligands present in the cells and/or in the external milieu exert an important biological activity capable of regulating the production of IL-22 and IL-17A.IL-23 and IL-1 are not required for TCDD-induced increase in IL-22 production IL-23 is reported to favor Th17 cell terminal differentiation and coexpression of 17]. To determine whether IL-23 was required for the production of IL-22, we activated purified CD4 1 T cells by CD3 and CD28 cross-linking in the presence of a neutralizing anti-IL-23 mAb.…”

“…3 and natural AHR ligands [31]. They are at variance with the enhanced IL-22 production and concomitant enhanced IL-17A production observed in the presence of FICZ and TCDD in the mouse [27][28][29]. TCDD was, in addition, shown to induce Treg in the mouse [25,26], whereas in our in vitro system we could not trace the emergence of Treg as far as TCDD did not upregulate the expression of FOXP3, nor the production of IL-10 and TGF-b.…”

“…AHR binds to, and is activated by a range of structurally divergent chemicals including natural dietary, endogenous ligands, and synthetic environmental agents among which dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)) is the most extensively studied pure agonist [23,24]. In the mouse, AHR is reported to regulate Th17 and Treg differentiation in ligand-specific fashion [25][26][27] and natural AHR agonists favor Th17 differentiation in vitro and enhanced IL-22 production [27][28][29][30]. Human lymphocytes may, however, behave differently and in a recent report AHR agonists appear to favor IL-22 but not IL-17 production [31].…”

Protein function analysis: rapid, cell-based siRNA-mediated ablation of endogenous expression with simultaneous ectopic replacement

“…The AHR exhibits diverse cellular activities, the most studied of which concerns its ability to regulate a number of genes involved in xenobiotic metabolism . More recently, the AHR has been shown to regulate certain aspects of inflammatory signaling such as T cell differentiation, acute phase response and IL6 expression in tumor cells (Veldhoen et al 2009;Patel et al 2009;Hollingshead et al 2008). Some of these effects are seen to occur at different levels across cell types.…”