IL-1F6, IL-1F8 and IL-1F9 and the IL-1R6(RP2) receptor antagonist IL-1F5 constitute a novel IL-1 signaling system that is poorly characterized in skin. To further characterize these cytokines in healthy and inflamed skin, we studied their expression in healthy control (NN), uninvolved psoriasis (PN) and psoriasis plaque (PP) skin using QRT-PCR and immunohistochemistry. Expression of IL-1F5, -1F6, -1F8, and -1F9 were increased 2-3 orders of magnitude in PP versus PN skin, which was supported immunohistologically. Moreover, treatment of psoriasis with etanercept led to significantly decreased IL-1F5, -1F6, -1F8 and -1F9 mRNAs, concomitant with clinical improvement. Similarly increased expression of IL-1F5, -1F6, -1F8 and -1F9 was seen in the involved skin of two mouse models of psoriasis. Suggestive of their importance in inflamed epithelia, IL-1α and TNF-α induced IL-1F5, -1F6, -1F8, and -1F9 transcript expression by normal human keratinocytes. Microarray analysis revealed that these cytokines induce the expression of anti-microbial peptides and matrix metalloproteins by reconstituted human epidermis. In particular, IL-1F8 increased mRNA expression of HBD2, HBD3 and CAMP and protein secretion of HBD2 and HBD3. Collectively, our data suggest important roles for these novel cytokines in inflammatory skin diseases and identify these peptides as potential targets for antipsoriatic therapies.

IL-17C is a functionally distinct member of the IL-17 family that binds IL-17RE/A to promote innate defense in epithelial cells and regulate Th17 cell differentiation. We demonstrate that IL-17C (not IL-17A) is the most abundant IL-17 isoform in lesional psoriasis skin (1058pg/ml vs. 8pg/ml; p<0.006) and localizes to keratinocytes (KCs), endothelial cells (ECs) and leukocytes. ECs stimulated with IL-17C produce increased TNFα and KCs stimulated with IL-17C/TNFα produce similar inflammatory gene response patterns as those elicited by IL-17A/TNFα, including increases in IL-17C, TNFα, IL-8, IL-1α/β, IL-1F5, IL-1F9, IL-6, IL-19, CCL20, S100A7/A8/A9, DEFB4, LCN2 and PI3 (p<0.05); indicating a positive pro-inflammatory feedback loop between the epidermis and ECs. Psoriasis patients treated with etanercept rapidly decrease cutaneous IL-17C levels, suggesting IL-17C/TNFα-mediated inflammatory signaling is critical for psoriasis pathogenesis. Mice genetically engineered to overexpress IL-17C in KCs develop well-demarcated areas of erythematous, flakey “involved” skin adjacent to areas of normal appearing “uninvolved” skin despite increased IL-17C expression in both areas (p<0.05). Uninvolved skin displays increased angiogenesis and elevated S100A8/A9expression (p<0.05) but no epidermal hyperplasia; whereas involved skin exhibits robust epidermal hyperplasia, increased angiogenesis and leukocyte infiltration and upregulated TNFα, IL-1α/β, IL-17A/F, IL-23p19, VEGF, IL-6 and CCL20 (p<0.05) suggesting that IL-17C, when coupled with other pro-inflammatory signals, initiates the development of psoriasiform dermatitis. This skin phenotype was significantly improved following 8 weeks of TNFα inhibition. These findings identify a role for IL-17C in skin inflammation and suggest a pathogenic function for the elevated IL-17C observed in lesional psoriasis skin.

The IL-1 family members IL-36α (IL-1F6), IL-36β (IL-1F8) and IL-36γ (IL-1F9) and the receptor antagonist IL-36Ra (IL-1F5) constitute a novel signaling system that is poorly understood. We now show that these cytokines have profound effects on the skin immune system. Treatment of human keratinocytes with IL-36 cytokines significantly increased the expression of CXCL1, CXCL8, CCL3, CCL5, and CCL20, potent chemotactic agents for activated leukocytes, and IL-36α injected intradermally resulted in chemokine expression, leukocyte infiltration and acanthosis of mouse skin. Blood monocytes, myeloid dendritic cells (DC) and monocyte-derived DC (MO-DC) expressed IL-36R and responded to IL-36. In contrast, no direct effects of IL-36 on resting or activated human CD4+ or CD8+ T cells, or blood neutrophils, could be demonstrated. Monocytes expressed IL-1A, IL-1B and IL-6 mRNA and IL-1β and IL-6 protein and mDC upregulated surface expression of CD83, CD86 and HLADR and secretion of IL-1β and IL-6 after treatment with IL-36. Furthermore, IL-36α-treated MO-DC enhanced allogeneic CD4+ T cell proliferation, demonstrating that IL-36 can stimulate the maturation and function of DC and drive T cell proliferation. These data indicate that IL-36 cytokines actively propagate skin inflammation via the activation of keratinocytes, antigen presenting cells and, indirectly, T cells.

To further elucidate molecular alterations in psoriasis, we performed a gene expression study on 58 paired lesional and uninvolved psoriatic and 64 control skin samples. Comparison of involved psoriatic (PP) and normal (NN) skin identified 1,326 differentially regulated transcripts encoding 918 unique genes (548 up- and 369 down-regulated), of which over 600 were to our knowledge unreported, including S100A7A, THRSP and ELOVL3. Strongly up-regulated genes included SERPINB4, PI3, DEFB4 and several S100 family members. Strongly down-regulated genes included Wnt-inhibitory factor-1 (WIF1), betacellulin (BTC), and CCL27. Enriched gene ontology categories included immune response, defense response, and keratinocyte differentiation. Biological processes regulating fatty acid and lipid metabolism were enriched in the down-regulated gene set. Comparison of the psoriatic transcriptome to the transcriptomes of cytokine-stimulated cultured keratinocytes (IL-17, IL-22, IL-1α, IFN-γ, TNF-α and oncostatin-M) revealed surprisingly little overlap, with the cytokine stimulated keratinocyte expression representing only 2.5%, 0.7%, 1.5%, 5.6%, 5.0% and 1.9% of the lesional psoriatic dysregulated transcriptome, respectively. This comprehensive analysis of differentially regulated transcripts in psoriasis provides additional insight into the pathogenic mechanisms involved and emphasizes the need for more complex yet tractable experimental models of psoriasis.

Ligands of the EGF family regulate autocrine keratinocyte proliferation and IL-1 family cytokines orchestrate epithelial defense responses. While members of both families are overexpressed in wound healing and psoriasis, their roles in regulating the innate immune functions of keratinocytes remain incompletely explored. Using sensitive assays, we found significant increases of HB-EGF, TGF-α and amphiregulin mRNA and protein in lesional psoriasis compared with uninvolved or control skin. In normal human keratinocyte (NHK) monolayers, EGFR ligands were ineffective in inducing DEFB4, S100A7, and CCL20 mRNAs and hBD-2 peptide. Combined with IL-1α, however, EGFR ligands provoked 250× more DEFB4 and CCL20 and a 9-fold rise in S100A7 mRNA relative to the EGFR ligand alone. This synergy was also reflected in secreted hBD-2 protein, both from NHK and reconstituted human epidermis. Keratinocyte differentiation was critical for these responses, as postconfluent NHK yielded mRNA and protein levels an order of magnitude greater than subconfluent cells. Differentiation also influenced signal transduction, with subconfluent cells utilizing NF-κB and postconfluent cells utilizing EGFR, MEK1/2, and p38. We propose that EGFR ligands are important modifiers of IL-1 activity, synergizing with IL-1 to stimulate epidermal production of hBD-2, S100A7 and CCL20, three of the most upregulated transcripts in psoriatic plaques.