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Current inflammatory bowel disease (IBD) therapies are ineffective in a high proportion of patients. Combining bulk and single-cell transcriptomics, quantitative histopathology and in situ localization across three cohorts of patients with IBD (total n = 376), we identify coexpressed gene modules within the heterogeneous tissular inflammatory response in IBD that map to distinct histopathological and cellular features (pathotypes). One of these pathotypes is defined by high neutrophil infiltration, activation of fibroblasts and vascular remodeling at sites of deep ulceration. Activated fibroblasts in the ulcer bed display neutrophil-chemoattractant properties that are IL-1R, but not TNF, dependent. Pathotype-associated neutrophil and fibroblast signatures are increased in nonresponders to several therapies across four independent cohorts (total n = 343). The identification of distinct, localized, tissular pathotypes will aid precision targeting of current therapeutics and provides a biological rationale for IL-1 signaling blockade in ulcerating disease., Transcriptomic and histological profiling of gut biopsies from multiple independent cohorts of patients with inflammatory bowel disease identifies distinct histopathological, molecular and cellular features associated with treatment response, providing insights for patient stratification and precision therapy.

BACKGROUND: Pro-inflammatory fibroblasts are critical for pathogenesis in rheumatoid arthritis, inflammatory bowel disease, interstitial lung disease, and Sjögren’s syndrome and represent a novel therapeutic target for chronic inflammatory disease. However, the heterogeneity of fibroblast phenotypes, exacerbated by the lack of a common cross-tissue taxonomy, has limited our understanding of which pathways are shared by multiple diseases. METHODS: We profiled fibroblasts derived from inflamed and non-inflamed synovium, intestine, lungs, and salivary glands from affected individuals with single-cell RNA sequencing. We integrated all fibroblasts into a multi-tissue atlas to characterize shared and tissue-specific phenotypes. FINDINGS: Two shared clusters, CXCL10(+)CCL19(+) immune-interacting and SPARC(+)COL3A1(+) vascular-interacting fibroblasts, were expanded in all inflamed tissues and mapped to dermal analogs in a public atopic dermatitis atlas. We confirmed these human pro-inflammatory fibroblasts in animal models of lung, joint, and intestinal inflammation. CONCLUSIONS: This work represents a thorough investigation into fibroblasts across organ systems, individual donors, and disease states that reveals shared pathogenic activation states across four chronic inflammatory diseases. FUNDING: Grant from F. Hoffmann-La Roche (Roche) AG.

Lipids play a major role in inflammatory diseases by altering inflammatory cell functions, through their use as energy substrates or as lipid mediators such as oxylipins. Autophagy, a lysosomal degradation pathway that limits inflammation, is known to impact on lipid availability, however whether this controls inflammation remains unexplored. We found that upon intestinal inflammation visceral adipocytes upregulate autophagy and that adipocyte-specific loss of the autophagy gene Atg7 exacerbates inflammation. While autophagy decreased lipolytic release of free fatty acids, loss of the major lipolytic enzyme Pnpla2/Atgl in adipocytes did not alter intestinal inflammation, ruling out free fatty acids as anti- inflammatory energy substrates. Instead, Atg7-deficient adipose tissues exhibited an altered oxylipin balance, driven through an NRF2-mediated upregulation of Ephx1. This was accompanied by a shift in adipose tissue macrophage polarization with reduced secretion of IL-10, leading to lower circulating levels of IL-10. These results suggest an underappreciated fat-gut crosstalk through an autophagy- dependent regulation of anti-inflammatory oxylipins, indicating a protective effect of adipose tissues for distant inflammation.

Current inflammatory bowel disease (IBD) therapies are ineffective in a high proportion of patients. Combining bulk and single-cell transcriptomics, quantitative histopathology, and in situ localisation, we describe heterogeneity of the tissular inflammatory response in IBD treatment failure. Among inflammatory pathotypes, we found high neutrophil infiltration, activation of fibroblasts, and vascular remodelling at sites of deep ulceration was a feature of non-response to several anti-inflammatory therapies. Activated fibroblasts in the ulcer bed display neutrophil chemoattractant properties that are IL-1R- but not TNF-dependent. The identification of distinct, localised, tissular pathotypes associated with treatment non-response will aid precision targeting of current therapeutics and provide a biological rationale for IL-1 signalling blockade in ulcerating disease.

SummaryPro-inflammatory fibroblasts are critical to pathogenesis in rheumatoid arthritis, inflammatory bowel disease, interstitial lung disease, and Sjögren’s syndrome, and represent a novel therapeutic target for chronic inflammatory disease. However, the heterogeneity of fibroblast phenotypes, exacerbated by the lack of a common cross-tissue taxonomy, has limited the understanding of which pathways are shared by multiple diseases. To investigate, we profiled patient-derived fibroblasts from inflamed and non-inflamed synovium, intestine, lung, and salivary glands with single-cell RNA-sequencing. We integrated all fibroblasts into a multi-tissue atlas to characterize shared and tissue-specific phenotypes. Two shared clusters, CXCL10+CCL19+ immune-interacting and SPARC+COL3A1+ vascular-interacting fibroblasts were expanded in all inflamed tissues and additionally mapped to dermal analogues in a public atopic dermatitis atlas. We further confirmed these human pro-inflammatory fibroblasts in animal models of lung, joint, and intestinal inflammation. This work represents the first cross-tissue, single-cell fibroblast atlas revealing shared pathogenic activation states across four chronic inflammatory diseases.

Background A relationship between alcohol consumption and psoriasis has been reported, but it is unclear whether alcohol consumption aggravates psoriasis. Here, we studied the effect of chronic ethanol (EtOH) consumption in the murine model of Aldara-induced psoriasiform dermatitis. Methods C57BL/6 mice received 5% EtOH in their drinking water for 10 weeks. Dermatitis was induced from weeks 9 to 10, by applying Aldara to the shaved patch of skin on the back. Inflammation was characterized by histological and transcriptomic analyses. Results EtOH consumption aggravated Aldara-induced dermatitis. The scales were more severe, epidermal thickening was more pronounced, and cutaneous expression of Th17-related cytokines was exacerbated. Control mice simply receiving EtOH displayed minimal cutaneous inflammation, characterized by epidermal infiltrates of T lymphocytes and the overexpression of IL-17A and the Th17-recruiting chemokine CCL20. In vitro studies showed that low concentrations of EtOH induce the expression of CCL20 by murine epidermal keratinocytes. Conclusion Alcohol consumption leads to subliminar skin inflammation, which is revealed by the exacerbation of Aldara-induced experimental psoriasiform dermatitis, likely through Th17-type minimal skin inflammation. These results favor the systematic management of alcohol consumption in psoriatic patients.

SummaryBackground Psoriasis associates several extra-cutaneous manifestations. Little is known about hepatic parameters specifically associated with psoriasis. Objective To study whether psoriasiform dermatitis is associated with liver injury. Methods We studied liver parameters of inflammation and fibrosis in a murine model of psoriasiform dermatitis induced by topical application of imiquimod for nine weeks. Results Topical treatment with imiquimod induced a form of psoriasiform dermatitis reminiscent of the human disorder, characterized by thickened and scaly skin, psoriasiform epidermal hyperplasia, altered keratinocyte differentiation, and cutaneous overexpression of interleukin-17A. Mice with dermatitis displayed hepatitis, as shown by elevation of plasma transaminase levels, as well as portal and periportal hepatitis characterized by T lymphocyte (CD3e+) and polymorphonuclear cell (Gr1+) infiltrates. The hepatitis progressed toward liver fibrogenesis, as shown by excessive Sirius red staining, consistent with the expression of alpha-smooth muscle actin by hepatic stellate cells. Conclusions These results indicate that liver inflammation and fibrosis is associated with experimental psoriasiform dermatitis. Our results suggest that psoriatic inflammation may be associated with specific liver injury. This article is protected by copyright. All rights reserved.

International audience; Recent studies suggest that psoriasis may be more severe in patients with nonalcoholic fatty liver disease, particularly in those with the inflammatory stage of steatohepatitis [nonalcoholic steatohepatitis (NASH)]. Herein, we investigated the impact of diet-induced steatohepatitis on the severity of imiquimod-induced psoriasiform dermatitis. Mice fed with a high-fat diet developed steatohepatitis reminiscent of human NASH with ballooning hepatocytes and significant liver fibrosis. Mice with steatohepatitis also displayed moderate cutaneous inflammation characterized by erythema, dermalinfiltrates of CD45(+) leukocytes, and a local production of IL-17A. Moreover, steatohepatitis was associated with an epidermal activation of caspase-1 and cutaneous overexpression of IL-1 beta. Imiquimod-induced psoriasiform dermatitis was exacerbated in mice with steatohepatitis as compared to animals fed with a standard diet. Scale formation and acanthosis were aggravated, in correlation with increased IL-17A and IL-22 expression in inflamed skins. Finally, intradermal injection of IL-17A in standard diet-fed mice recapitulated the cutaneous pathology of mice with steatohepatitis. The results show that high-fat diet induced steatohepatitis aggravates the inflammation in psoriasiform dermatitis, via the cutaneous production of IL-17A. In agreement with clinical data, this description of a novel extrahepatic manifestation of NASH should sensitize dermatologists to the screening and the management of fatty liver in psoriatic patients.

International audience; The development of three‐dimensional models of reconstituted mouse epidermis (RME) has been hampered by the difficulty to maintain murine primary keratinocyte cultures and to achieve a complete epidermal stratification. In this study, a new protocol is proposed for the rapid and convenient generation of RME, which reproduces accurately the architecture of a normal mouse epidermis. During RME morphogenesis, the expression of differentiation markers such as keratins, loricrin, filaggrin, E‐cadherin and connexins was followed, showing that RME structure at day 5 was similar to those of a normal mouse epidermis, with the acquisition of the natural barrier function. It was also demonstrated that RME responded to skin‐relevant proinflammatory cytokines by increasing the expression of antimicrobial peptides and chemokines, and inhibiting epidermal differentiation markers, as in the human system. This new model of RME is therefore suitable to investigate mouse epidermis physiology further and opens new perspectives to generate reconstituted epidermis from transgenic mice.

The pathogenesis of inflammatory skin diseases such as psoriasis involves the release of numerous proinflammatory cytokines, including members of the IL-1 family. Here we report overexpression of IL-1α, IL-1β, and IL-1 receptor antagonist mRNA, associated to expression of IL-23p19, IL-17A, and IL-22 in skin cells, upon topical application of the TLR7 agonist imiquimod (IMQ) in C57BL/6J mice. IMQ-induced skin inflammation was partially reduced in mice deficient for both IL-1α/IL-1β or for IL-1 receptor type 1 (IL-1R1), but not in IL-1α- or IL-1β-deficient mice, demonstrating the redundant activity of IL-1α and IL-1β for skin inflammation. NLRP3 or apoptosis-associated Speck-like protein containing a Caspase recruitment domain-deficient mice had no significant reduction of skin inflammation in response to IMQ treatment, mainly due to the redundancy of IL-1α. However, IMQ-induced skin inflammation was abolished in the absence of MyD88, the adaptor protein shared by IL-1R and TLR signaling pathways. These results are consistent with the TLR7 dependence of IMQ-induced skin inflammation. Thus, IL-1R1 contributes to the IMQ-induced skin inflammation, and disruption of MyD88 signaling completely abrogates this response.

Background Acute-serum Amyloid A (A-SAA), one of the major acute-phase proteins, is mainly produced in the liver but extra-hepatic synthesis involving the skin has been reported. Its expression is regulated by the transcription factors NF-κB, C/EBPβ, STAT3 activated by proinflammatory cytokines. Objectives We investigated A-SAA synthesis by resting and cytokine-activated Normal Human Epidermal Keratinocytes (NHEK), and their inflammatory response to A-SAA stimulation. A-SAA expression was also studied in mouse skin and liver in a model mimicking psoriasis and in the skin and sera of psoriatic and atopic dermatitis (AD) patients. Methods NHEK were stimulated by A-SAA or the cytokines IL-1α, IL-17A, IL-22, OSM, TNF-α alone or in combination, previously reported to reproduce features of psoriasis. Murine skins were treated by imiquimod cream. Human skins and sera were obtained from patients with psoriasis and AD. A-SAA mRNA was quantified by RT qPCR. A-SAA proteins were dosed by ELISA or immunonephelemetry assay. Results IL-1α, TNF-α and mainly IL-17A induced A-SAA expression by NHEK. A-SAA induced its own production and the synthesis of hBD2 and CCL20, both ligands for CCR6, a chemokine receptor involved in the trafficking of Th17 lymphocytes. A-SAA expression was increased in skins and livers from imiquimod-treated mice and in patient skins with psoriasis, but not significantly in those with AD. Correlations between A-SAA and psoriasis severity and duration were observed. Conclusion Keratinocytes could contribute to psoriasis pathogenesis via A-SAA production, maintaining a cutaneous inflammatory environment, activating innate immunity and Th17 lymphocyte recruitment.

International audience; Oncostatin M (OSM) has been reported to be overexpressed in psoriasis skin lesions and to exert proinflammatory effects in vitro on human keratinocytes. Here, we report the proinflammatory role of OSM in vivo in a mouse model of skin inflammation induced by intradermal injection of murine OSM-encoding adenovirus (AdOSM) and compare with that induced by IL-6 injection. Here, we show that OSM potently regulates the expression of genes involved in skin inflammation and epidermal differentiation in murine primary keratinocytes. In vivo, intradermal injection of AdOSM in mouse ears provoked robust skin inflammation with epidermal thickening and keratinocyte proliferation, while minimal effect was observed after AdIL-6 injection. OSM overexpression in the skin increased the expression of the S100A8/9 antimicrobial peptides, CXCL3, CCL2, CCL5, CCL20, and Th1/Th2 cytokines, in correlation with neutrophil and macrophage infiltration. In contrast , OSM downregulated the expression of epidermal differentiation genes, such as cytokeratin-10 or filaggrin. Collectively, these results support the proinflammatory role of OSM when it is overexpressed in the skin. However, OSM expression was not required in the murine model of psoriasis induced by topical application of imiquimod, as demonstrated by the inflammatory phenotype of OSM-deficient mice or wild-type mice treated with anti-OSM antibodies.

Introduction Les cultures de keratinocytes en monocouche sont couramment utilisees dans les etudes in vitro, mais celles-ci ne reproduisent pas fidelement l’architecture de l’epiderme et les caracteristiques physiologiques des differents etats de differenciation cellulaire. Dans cette etude, nous decrivons un protocole simple et rapide pour la generation d’epidermes reconstruits murins (RME) a partir de keratinocytes primaires de souriceaux. Apres avoir caracterise les differentes etapes de leur differenciation, nous analysons leur reponse a la stimulation par des cytokines proinflammatoires. Materiel et methodes Les RME sont realises a partir de keratinocytes primaires issus de peaux de souriceaux nouveau-nes cultives dans des inserts en polycarbonate dans un milieu d’ensemencement. Des l’obtention d’une monocouche cellulaire, les inserts sont passes a l’interface air-liquide au contact d’un milieu de differenciation. La caracterisation des RME est effectuee par histologie et par immunohistofluorescence pour la detection de marqueurs de differenciation incluant les keratines 14, 6, 10, la loricrine, la filaggrine, l’E-Cadherine et des connexines. Leur permeabilite est eprouvee par des tests de diffusion de la sonde fluorescente Lucifer Yellow . Enfin, la reponse des RME aux cytokines IL-17A, TNFα, IL-22, IL-1α et oncostatine M est evaluee par RT-qPCR par la mesure de l’expression de marqueurs de differenciation, de peptides antimicrobiens et de chimiokines. Resultats Notre protocole permet d’obtenir des RME qui reproduisent les caracteristiques morphologiques et phenotypiques d’un epiderme de souris en seulement 5 jours apres leur passage a l’interface air-liquide comme l’atteste l’expression des marqueurs de differenciation conforme a celle d’un epiderme natif. Les RME acquierent egalement leur fonction de barriere et sont impermeables au passage de la Lucifer Yellow . Enfin, suite a leur stimulation par des cytokines proinflammatoires, les RME repondent de maniere similaire a un epiderme in vivo par une diminution d’expression de la keratine 10, de l’involucrine et de la filaggrine et une augmentation de l’expression de mediateurs de l’inflammation tels que la chimiokine CXCL-3 et les peptides antimicrobiens S100A9 et β-defensine-3. Discussion Ce nouveau protocole permet donc de realiser rapidement et facilement des RME reproduisant fidelement l’architecture et les caracteristiques physiologiques d’un epiderme, y compris leur reponse dans un contexte inflammatoire. Conclusion Ce modele de RME permet d’etudier les mecanismes physiologiques de la peau et ouvre la perspective de realiser des RME a partir de keratinocytes issus de souris genetiquement modifiees.

De nombreuses etudes in vitro sont realisees a partir de cultures primaires de keratinocytes en monocouche. Ces cultures cellulaires sont composees majoritairement de cellules progenitrices de la couche basale, presentant un potentiel proliferatif important mais un etat de differenciation epidermique peu avance. Afin d’etudier in vitro la physiologie du keratinocyte dans un systeme plus differencie, nous avons mis au point un protocole d’obtention d’epidermes reconstruits murins (RME) a partir de keratinocytes primaires issus de souriceaux nouveau-nes. La caracterisation des RME en immuno-histofluorescence montre une expression similaire a celle de la peau de souris des marqueurs de differenciation epidermique, tels que les cytokeratines 6/10, la loricrine ou la filaggrine, de cadherines et de connexines des j5. Ces RME permettent egalement de mettre en evidence le blocage de la differenciation induit par les cytokines pro-inflammatoires telles que l’oncostatine M ou l’IL-22. Nous avons egalement compare les profils d’expression des Toll-like Receptors des RME avec les cultures de keratinocytes en monocouche et la peau de souris. Outre l’etude des reponses des RME en conditions infectieuses et/ou inflammatoires, ce protocole ouvre la possibilite de generer des RME a partir de souris transgeniques knock-out ou knock-in.

The pathogenesis of inflammatory skin diseases such as psoriasis involves the release of numerous proinflammatory cytokines, including members of the IL-1 family. Here we report overexpression of IL-1α, IL-1β, and IL-1 receptor antagonist mRNA, associated to expression of IL-23p19, IL-17A, and IL-22 in skin cells, upon topical application of the TLR7 agonist imiquimod (IMQ) in C57BL/6J mice. IMQ-induced skin inflammation was partially reduced in mice deficient for both IL-1α/IL-1β or for IL-1 receptor type 1 (IL-1R1), but not in IL-1α- or IL-1β-deficient mice, demonstrating the redundant activity of IL-1α and IL-1β for skin inflammation. NLRP3 or apoptosis-associated Speck-like protein containing a Caspase recruitment domain-deficient mice had no significant reduction of skin inflammation in response to IMQ treatment, mainly due to the redundancy of IL-1α. However, IMQ-induced skin inflammation was abolished in the absence of MyD88, the adaptor protein shared by IL-1R and TLR signaling pathways. These results are consistent with the TLR7 dependence of IMQ-induced skin inflammation. Thus, IL-1R1 contributes to the IMQ-induced skin inflammation, and disruption of MyD88 signaling completely abrogates this response.

La steatopathie hepatique metabolique (SHM), dont la forme inflammatoire correspond a la steatohepatite non alcoolique (NASH), est 2 fois plus frequente dans la population psoriasique ; sa presence pourrait aggraver l’atteinte cutanee. Notre objectif est d’etudier l’impact de la NASH dans un modele murin de psoriasis induit par l’application topique d’imiquimod (IMQ). La NASH est induite par un regime hyperlipidique enrichi en acide cholique de 11 semaines ; de la 9e a la 11e semaine, les souris recoivent tous les 2 jours une application cutanee d’IMQ. Avant induction du psoriasis, les souris recevant le regime hyperlipidique presentent une peau erythemateuse associee a une surexpression cutanee d’IL-17A. Apres traitement par IMQ, l’inflammation cutanee est plus severe chez les souris NASH que chez les souris recevant un regime standard : le score clinique est augmente, l’epaississement epidermique est plus important (62 ± 9 μm vs 90 ± 10 μm, p