Rorγt+ innate lymphocytes and γδ T cells initiate psoriasiform plaque formation in mice

Abstract

Psoriasis is a common, relapsing inflammatory skin disease characterized by erythematous scaly plaques. Histological manifestations of psoriasis include keratinocyte dysregulation and hyperproliferation, elongated rete ridges, and inflammatory infiltrates consisting of T cells, macrophages, dendritic cells, and neutrophils. Despite the availability of new effective drugs to treat psoriasis, the underlying mechanisms of pathogenesis are still poorly understood. Recent studies have shown that Aldara cream, used to treat benign skin abnormalities, triggers psoriasis-like disease in humans and mice and have implicated Th17 cells in disease initiation. Using this as a model, we found a predominant role for the Th17 signature cytokines IL-17A, IL-17F, and IL-22 in psoriasiform plaque formation in mice. Using gene-targeted mice, we observed that loss of Il17a, Il17f, or Il22 strongly reduced disease the severity of psoriasis. However, we found that Th17 cells were not the primary source of these pathogenic cytokines. Rather, IL-17A, IL-17F, and IL-22 were produced by a skin-invading population of γδ T cells and RORγt(+) innate lymphocytes. Furthermore, our findings establish that RORγt(+) innate lymphocytes and γδ T cells are necessary and sufficient for psoriatic plaque formation in an experimental disease model that closely resembles human psoriatic plaque formation.

Figure 1. IL-17A, IL-17F, and IL-22 are important for psoriatic plaque formation.

(A and B) WT mice were treated with Aldara and anti–IL-12/23p40 mAb or isotype control on day 2. (A) Kinetics of skin inflammation as percent increase in thickness over 6 days (n = 4). (B) Skin sections were stained with H&E and anti-MPO. Scale bars: 100 μm. (C) Dot plots display the secretion of IL-17A, IL-17F, and IL-22 among CD45⁺ cells from the skin of wild-type mice on day 5 of Aldara treatment. (D and E) WT, Il17a^–/–, Il17f^–/–, and Il22^–/– mice were treated with Aldara or control cream for 5 days. Scatter plot shows percent increase in skin thickness (n = 4) (D). Skin sections of Aldara-treated mice taken on day 6 (E) were stained with H&E. Scale bar: 100 μm. (F) IL-17AF heterodimer concentration was measured in the supernatant of LN or skin cells cultured for 24 hours using IL-17AF FlowCytomix Simplex kit (eBioscience). Each experiment was performed independently at least 3 times. *P < 0.05, **P < 0.01, ^#P < 0.001.

Figure 2. RORγt⁺ γδ T cells and innate lymphocytes are the main producers of IL-17A, IL-17F, and IL-22 in psoriasiform plaques.

(A) Intracellular cytokine staining in the skin of Rorc-Cre × EYFP mice after 5 days of Aldara treatment, gated on CD45⁺ cells (n = 3), with (B) scatter plots showing percent distribution (n = 3). (C) Cytokine staining of V4γ⁺ versus Vγ5⁺ for IL-17F and IL-22, pre-gated on TCRγδ⁺ cells. (D and E) Kinetics of Aldara-induced skin inflammation in (D) WT versus Tcrb^–/– and Tcrd^–/– mice (n = 4) and (E) WT versus Il15ra^–/– mice (n = 3), shown as the percent increase in skin thickness. Each experiment was performed independently at least 3 times.*P < 0.05, ^#P < 0.001.

Figure 3. RORγt⁺ innate lymphocytes are essential for psoriasiform plaque formation.

(A) Kinetics of Aldara-induced skin inflammation in WT versus Tcrd^–/– versus Rag1^–/– mice. (B) Cytokine staining in ILCs in the skin of Aldara- versus control-treated mice, pre-gated on Lin^–CD45⁺ cells (n = 3). (C) Fate map analysis and IL-22 staining in the Aldara-treated skin of Rorc-Cre X EYFP and Rorc-Cre × EYFP × Rag1^–/– mice gated on CD45⁺ cells. Kinetics of Aldara-induced skin inflammation in (D) WT versus Rag1^–/– versus Rag2^–/–Il2rg^–/– mice and (E) WT versus Rorc^–/– mice showing percent change in skin thickness (n = 3). (F) Staining for IL-17F and IL-22 in CD45⁺ cells from the skin of control and Aldara-treated WT versus Rorc^–/– mice (n = 3). Each experiment was performed independently at least 3 times. ^#P < 0.001.