IκBζ (NFKBIZ) has been implicated as a key co-transcription factor in psoriasis pathogenesis. While its role in keratinocytes is well established, the involvement in dermal fibroblasts, another critical skin cell type, remains underexplored. This study characterizes cytokine-induced NFKBIZ regulation in human dermal fibroblasts in vitro and integrates spatial transcriptomics to determine NFKBIZ expression patterns in psoriatic skin biopsies. Primary dermal fibroblasts were stimulated with IL-17A, IL-17F, and TNF. Signaling pathways and gene regulation were examined using chemical inhibitors, siRNA knockdown, qPCR, and Western blotting. Additionally, spatial transcriptomics (CosMx™) assessed NFKBIZ expression in paired lesional and non-lesional psoriatic skin biopsies. Results showed significant upregulation of IκBζ expression in dermal fibroblasts following stimulation with both IL-17A and IL-17F. The NF-κB signaling pathway was identified as the primary regulator of NFKBIZ induction. NFKBIZ knockdown significantly reduced cytokine-induced expression of inflammatory mediators (CXCL8, CCL20, CCL2), confirming its regulatory role. Spatial transcriptomics further confirmed NFKBIZ expression in dermal fibroblasts in vivo, particularly in lesional psoriatic skin. This study establishes IκBζ as a critical modulator of inflammatory responses in dermal fibroblasts, expanding its recognized role beyond keratinocytes and immune cells, and highlights IκBζ inhibition as a potential therapeutic strategy.

Recombinant proteins are critical tools in research; however, their purity is often assumed rather than verified, leading to potential experimental errors. This study aimed to investigate the inflammatory role of recombinant human IL-17F in dermal fibroblasts. Unexpectedly, we discovered with Western blot that recombinant IL-17F from the supplier was contaminated with IL-4, leading to unintended stimulatory effects such as STAT6 phosphorylation and gene induction of CCL26 and IL4R. This contamination led to misinterpretation of data, loss of research time, and erroneous conclusions about IL-17F activity. These findings underscore the critical need for stringent quality control in recombinant protein production and highlight the risks of relying on single-source suppliers. Researchers should remain cautious about potential contamination, ideally validating proteins from multiple suppliers. Our experience illustrates a broader requirement for suppliers to strengthen quality assurance measures, as contaminants can propagate misleading data in the literature and undermine research reproducibility.

Chronic inflammatory skin diseases such as psoriasis and hidradenitis suppurativa are driven by cytokines, including IL‐17A and TNF. Although biologics targeting these cytokines have transformed therapy, the transcriptional contributions of individual skin‐resident cell types remain unclear, and dermal fibroblasts have been largely overlooked compared with keratinocytes. To address this, we compared the transcriptional responses of primary human dermal fibroblasts and keratinocytes following in vitro stimulation with IL‐17A, TNF, or both, using bulk RNA sequencing and Western blotting. Dermal fibroblasts mounted a stronger and broader proinflammatory response than keratinocytes. This was particularly evident in response to TNF and combined TNF/IL‐17A stimulation, with enrichment for immune signalling and chemotaxis pathways and robust induction of chemokine genes, including CCL20 , CXCL8 , and IL6 . Keratinocytes primarily upregulated genes associated with epithelial differentiation, barrier function, and protein regulation, including IL36G , S100A7A , and DEFB4A . The heightened fibroblast responsiveness correlated with increased TNF sensitivity and substantially higher TNFR2 ( TNFRSF1B ) expression and signalling compared with keratinocytes, suggesting a fibroblast‐specific mechanism amplifying inflammatory responses. These findings challenge the keratinocyte‐centric view of skin inflammation and identify dermal fibroblasts as active contributors and potential therapeutic targets in TNF‐ and Th17‐driven skin diseases.

T helper 17 (Th17)-mediated skin diseases, such as psoriasis and hidradenitis suppurativa, are driven by a dysregulated cytokine network where the specific contribution of Interleukin (IL)-17F has historically been underestimated due to its lower intrinsic potency compared to IL-17 A. To elucidate the role of IL-17F in the dermal compartment, we performed bulk RNA sequencing on primary human dermal fibroblasts stimulated with recombinant IL-17 A or IL-17F, either alone or in combination with Tumor Necrosis Factor (TNF). While single stimulation confirmed the superior potency of IL-17 A, the addition of TNF completely abolished this hierarchy. Transcriptional profiling demonstrated that TNF synergizes potently with IL-17F, amplifying its inflammatory output to levels almost indistinguishable from the IL-17 A and TNF combination. This synergistic interplay drove a robust upregulation of pathogenic mediators, including neutrophil-attracting chemokines (CXCL1, CXCL8) and tissue-destructive enzymes (MMP1). These findings challenge the paradigm of IL-17F as a redundant homolog, demonstrating that within a TNF-rich pro-inflammatory environment, IL-17F overcomes its limited basal potency to mirror the inflammatory drive of IL-17 A functionally. This mechanism provides a biological rationale for the superior clinical efficacy observed with dual IL-17 A/F inhibition in chronic inflammatory skin diseases.

Chronic inflammatory skin diseases such as psoriasis and hidradenitis suppurativa are driven by cytokines, including IL-17A and TNF. Although biologics targeting these cytokines have transformed therapy, the transcriptional contributions of individual skin-resident cell types remain unclear, and dermal fibroblasts have been largely overlooked compared with keratinocytes. To address this, we compared the transcriptional responses of primary human dermal fibroblasts and keratinocytes following in vitro stimulation with IL-17A, TNF, or both, using bulk RNA sequencing and Western blotting. Dermal fibroblasts mounted a stronger and broader proinflammatory response than keratinocytes. This was particularly evident in response to TNF and combined TNF/IL-17A stimulation, with enrichment for immune signalling and chemotaxis pathways and robust induction of chemokine genes, including CCL20, CXCL8, and IL6. Keratinocytes primarily upregulated genes associated with epithelial differentiation, barrier function, and protein regulation, including IL36G, S100A7A, and DEFB4A. The heightened fibroblast responsiveness correlated with increased TNF sensitivity and substantially higher TNFR2 (TNFRSF1B) expression and signalling compared with keratinocytes, suggesting a fibroblast-specific mechanism amplifying inflammatory responses. These findings challenge the keratinocyte-centric view of skin inflammation and identify dermal fibroblasts as active contributors and potential therapeutic targets in TNF- and Th17-driven skin diseases.