IL-23-responsive innate lymphoid cells are increased in inflammatory bowel disease

Abstract

Results of experimental and genetic studies have highlighted the role of the IL-23/IL-17 axis in the pathogenesis of inflammatory bowel disease (IBD). IL-23-driven inflammation has been primarily linked to Th17 cells; however, we have recently identified a novel population of innate lymphoid cells (ILCs) in mice that produces IL-17, IL-22, and IFN-γ in response to IL-23 and mediates innate colitis. The relevance of ILC populations in human health and disease is currently poorly understood. In this study, we have analyzed the role of IL-23-responsive ILCs in the human intestine in control and IBD patients. Our results show increased expression of the Th17-associated cytokine genes IL17A and IL17F among intestinal CD3⁻ cells in IBD. IL17A and IL17F expression is restricted to CD56⁻ ILCs, whereas IL-23 induces IL22 and IL26 in the CD56⁺ ILC compartment. Furthermore, we observed a significant and selective increase in CD127⁺CD56⁻ ILCs in the inflamed intestine in Crohn's disease (CD) patients but not in ulcerative colitis patients. These results indicate that IL-23-responsive ILCs are present in the human intestine and that intestinal inflammation in CD is associated with the selective accumulation of a phenotypically distinct ILC population characterized by inflammatory cytokine expression. ILCs may contribute to intestinal inflammation through cytokine production, lymphocyte recruitment, and organization of the inflammatory tissue and may represent a novel tissue-specific target for subtypes of IBD.

Figure 1.

Th17 signature genes are expressed in intestinal CD3⁻ cells and overexpressed in IBD. (A) Relative messenger RNA (mRNA) expression of Th17 signature cytokines in intestinal tissue homogenates from control, UC, and CD patients. (B and C) mRNA expression of Th17-related genes in CD3⁺ cells (B) and CD3⁻ cells (C) isolated from blood and intestine of control (open circles) and IBD (closed circles) patients. In some experiments, B cells have been excluded (CD3⁻CD19⁻ cells). (A–C) The horizontal bars represent the mean of each of the groups. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Figure 2.

ILCs are a source of IL-17 in IBD. (A) mRNA expression of IL22, IL17A, IL17F, and IL26 in CD3⁻ cells from control (open bars; n = 9 for IL22, IL17A, and IL17F and n = 7 for IL26) and IBD colon (closed bars; n = 4) after overnight culture in complete media with no addition (NA) and in the presence of 10 ng/ml IL-23. In some experiments, B cells have been excluded (CD3⁻CD19⁻ cells). **, P = 0.006. (B) mRNA expression of IL22, IL26, IL17A, and IL17F in Lin⁻CD45⁺CD56⁺ and Lin⁻CD45⁺CD56⁻ cells from the ileum (n = 4) and colon (n = 4) of patients with CD after overnight stimulation in complete media with no addition (NA) and in the presence of 10 ng/ml IL-23. *, P = 0.029. (A and B) Mean ± standard error of the mean is represented. (C) Intracellular staining for IL-17A and IFN-γ after PMA/ionomycin stimulation in LPMCs isolated from the ileum of a patient with CD (representative of two experiments). LPMCs were gated on the lymphocytic gate (FSC/SSC), were CD3⁻CD19⁻CD14⁻, CD16⁻, CD45⁺, and costained with CD56 and CD127.

Figure 3.

CD56⁻ ILCs accumulate in the intestine in CD. (A) Representative staining of CD127⁺CD56⁻ and CD127⁺CD56⁺ ILCs from control and CD intestine. LPMCs were gated on the lymphocytic gate in the FSC/SSC plot, the Lin⁻ and CD45⁺ population. (B and C) Percentage of CD127⁺CD56⁻ (B) and CD127⁺CD56⁺ ILCs (C) in the Lin⁻CD45⁺ population, using the gates shown in A, in the colon of control, CD, and UC patients and in the ileum of control and CD patients. (B and C) The horizontal bars represent the mean of each of the groups. *, P = 0.048; **, P = 0.004.