Journal of Investigative Dermatology

ECM composition and organization are greatly altered during inflammation but it is still elusive if ECM dynamics may protect tissue stem cells against aberrant inflammation. Fibulin 7 (encoded by Fbln7) is part of the basement membrane ECM where epidermal stem cells (EpSCs) reside. It supports the long-term potential of fast-cycling EpSCs and moderates aging-related inflammatory markers in keratinocytes. Here, we assessed fibulin 7s role during imiquimod (IMQ)-induced inflammation in 1-year-old mouse dorsal skin. We found that loss of Fbln7 aggravates epidermal inflammation, marked by increased epidermal thickness, proliferation, and phosphorylation of JNK (c-Jun N-terminal kinase). Fast-cycling EpSCs labeled with Slc1a3-creER-TdTomato demonstrated that IMQ-induced proliferation in Fbln7 KO mice is contributed by cell divisions in the suprabasal layers, a hallmark of inflammatory epidermal responses. EpSC transcriptomes further reveal IMQ-modulated genes that are more substantially affected in Fbln7 KO mice, including IL-17 pathway-related genes known in psoriasis pathogenesis. Mechanistically, fibulin 7 directly binds to IL-17A and decreases IL-17A-mediated p38 MAPK activation. In public human psoriasis datasets, FBLN7 is reduced in lesional skin compared with non-lesional or normal skin, and it is significantly correlated with common psoriasis-associated genes. Altogether, fibulin 7 is potentially beneficial to protect against skin inflammation.

Lichen Planopilaris (LPP) is a lymphocyte-mediated scarring alopecia characterized by progressive follicular destruction and fibrosis. In this clinical trial, patients with biopsy proven LPP were treated with deucravacitinib (an oral inhibitor of tyrosine kinase 2 (TYK2)) 6 mg BID for 24 weeks (NCT-06091956). Bulk and single-cell RNA sequencing was performed on paired pre- and post-treatment scalp biopsies from baseline and week 4. Patients (N=10) demonstrated improvements in PGA (88.9%, p=0.008), LPPAI (-2.3 points, SD 1.1, p=0.002) and Skindex-16 (-21.0 points, SD 22.1, p=0.014) scores at week 24. Bulk transcriptomic analysis of untreated LPP revealed upregulation of type I Interferon (IFN)-stimulated genes and pathways related to inflammation, immune activation, keratinization, and extracellular matrix remodelling, with downregulation of immune and inflammatory pathways following treatment. Single-cell RNA-seq of LPP was characterized by enrichment of CD8+GZMK+ T cells which showed downregulation of T-cell receptor signaling as well as antiviral pathways with treatment. Basal keratinocytes exhibited reduced cytokine and interferon signaling and decreased communication with NK cells following treatment. CCL19+ fibroblasts were prominent in untreated disease was attenuated after treatment, with downregulation of type I IFN signaling. Selective TYK2 inhibition with deucravacitinib effectively suppresses these inflammatory circuits in LPP and represents a promising therapeutic strategy.

Peptidase inhibitor 16 (Pi16)-expressing fibroblasts are found across tissues and species, but their functional role is unclear. As fibroblasts and macrophages have been proposed to exist in a reciprocal circuit, we hypothesized Pi16+ fibroblasts may regulate macrophage homeostasis. Flow cytometry revealed [~]80% of skin fibroblasts express Pi16, leading us to investigate the role of these cells in maintaining a macrophage niche in this tissue. We generated an in vivo system where fibroblast-derived Colony Stimulating Factor 1 (Csf1) was constitutively eliminated in Pi16+ fibroblasts by crossing animals with a Csf1fl/fl allele to mice in which the gene Pi16 drives an IresCre cassette. Deletion of Csf1 in Pi16+ fibroblasts resulted in significant diminishment of CD64+ and CD11c+ macrophages alongside expansion of PDPN+YFP+ fibroblasts. Alterations in cell population dynamics coincided with thickening of both the dermis and fascial compartments of the skin. Deletion of Csf1 in Pi16+ fibroblasts delayed early wound healing in a unsplinted mouse model. Loss of PI16+ fibroblasts was observed in individuals with limited (lSSc) and diffuse (dSSc) systemic Scleroderma compared to healthy controls. These findings suggest that loss of Csf1 in Pi16+ fibroblasts elicit changes in the population dynamics of skin macrophages and modifications to tissue architecture.

Stem cell niches are dynamic microenvironments that regulate tissue homeostasis. Epidermal stem cells (EpiSC) preferentially localize to concave regions of epidermal rete ridges, which serve as primary niches for stem cell maintenance. EpiSC number and functional integrity decline during chronological aging. A defining feature of aged skin is epidermal atrophy, in which the prominent rete ridges present in young skin become flattened. Whether such topographical alterations influence EpiSC homeostasis and differentiation remains unclear. To address this, we generated anatomically accurate rete ridge structures using 3D bioprinting of collagen matrices as an ex vivo model and compared EpiSC cultured within concave topography to those maintained on a flat matrix resembling aged skin. Transcriptomic analysis revealed that concave niches promoted keratinocyte differentiation, marked by increased type I and II keratin gene expression and downregulation of cell cycle-associated genes. ATAC-seq identified topography-dependent chromatin accessibility changes enriched for transcription factors regulating epidermal differentiation, including upregulation of KLF4 and GRHL3 and downregulation of SOX9, HOXA1, and ETS1. Consistently, aged human skin showed reduced KLF4 and GRHL3 and increased SOX9 compared with young skin. Our findings demonstrate that concave niche topography imposes a spatially defined EpiSC microenvironment that promotes differentiation, alters cell cycle, and when perturbed, potentially contributes to the aging process. We conclude that spatial localization within rete ridge regions significantly affects epidermal progenitor stemness properties as fundamental differences in the physical microenvironment appear to influence cell fate decisions, thus, form shapes function of EpiSC.

Background: Human papillomaviruses (HPVs) pose a severe threat to global public health by driving nonmelanoma skin cancer (NMSC) and cervical cancer, with NMSC being one of the most common cancers worldwide. Epidermodysplasia verruciformis (EV) is an inborn error of immunity characterized by an increased susceptibility to persistent infection of cutaneous HPV and a high risk of NMSC. The genetic basis remains unknown in many patients with EV. Methods: We collected four unrelated pedigrees with EV. Genetic analysis identified five variants in JAK1 encoding the Janus kinase 1. Ex vivo models and patient-derived tissue were employed to evaluate the functional effects of JAK1 variants and delineate the pathogenic mechanisms. Results: We identified different variants in JAK1 in four pedigrees with dominant EV. Genetic analysis revealed five novel variants in JAK1, three of which resulted in nonsense-mediated mRNA decay (NMD). Functional assays identified a decreased phosphorylation of the signal transducers and activators of transcription (STATs), impaired interferon responses, and defective T cell activation. Immune dysregulation in patients, characterized by a reduced CD4/CD8 T cell ratio, decreased CD8 naive T cell proportion, and accumulated memory T cells, implies impaired antiviral immunity against HPV. Conclusions: Our findings confirm that JAK1 loss-of-function (LOF) variants underlie susceptibility to cutaneous HPV infection. [Funded by the National Natural Science Foundation of China (81788101, 81230015, 82394420, and 82394423), the National Key Research and Development Program of China (2022YFC2703900), the CAMS Innovation Fund for Medical Sciences (2021-I2M-1-018), and the Regione Lombardia, Italy (Innovative Research Project 1137-2010)].

Background: Atopic dermatitis (AD) is a prevalent chronic inflammatory skin disease associated with clinical, psychosocial, and economic burden. Accurate severity assessment is essential for guiding treatment escalation and monitoring disease activity, yet clinician-based scoring systems such as the Eczema Area and Severity Index (EASI) are limited by subjectivity and considerable inter- and intra-rater variability. Erythema, a key driver of AD severity grading, is particularly prone to inconsistent evaluation due to differences in ambient lighting, device quality, skin tone, and rater experience, underscoring the need for objective, reproducible assessment tools. Objective: To develop and validate an artificial intelligence (AI) pipeline for grading erythema, excoriation, and lichenification severity in AD from clinical photographs. The study evaluated the level of agreement between AI severity ratings in each category against dermatologists, non-specialists, and a consensus reference standard, with erythema as the primary outcome of interest. Methods: A two-stage AI pipeline was developed using EfficientNet B7 convolutional neural networks (CNNs). The first CNN was trained as a binary AD classifier on 451 AD and 601 non-AD images for lesion detection and segmentation. The second CNN was trained on 173 dermatologist-annotated AD images which were scored on a 0-3 ordinal scale for erythema, excoriation, and lichenification. This CNN had a downstream feature extraction algorithms such red channel contrast for erythema, Law's E5L5 for excoriation, and S5L5 texture maps for lichenification. In a cross-sectional validation study, 41 independent test images were scored by two blinded dermatologists and two blinded physicians. AI predictions were compared to individual rater groups and mode-derived consensus scores using weighted Cohen's kappa, classification accuracy, confusion matrices, and error direction analyses. Results: On internal validation, the severity CNN achieved 84% overall accuracy (averaged across all three attributes), 86% sensitivity, 87% specificity, and a macro-averaged area under the receiver operating characteristic curve (AUC) of 0.90. In the external comparison with blinded human raters, erythema agreement between the AI and dermatologist consensus was substantial (accuracy 80.7%; kappa = 0.68), with no large (>2-point) misclassifications. Physician consensus agreement was lower (accuracy 54.8%; kappa = 0.34), reflecting greater variability among primary care physicians (non-specialists). For excoriation, AI-dermatologist agreement was moderate (accuracy 72.4%; kappa = 0.62); for lichenification, agreement was similar (accuracy 71.4%; kappa = 0.59). Across all features, disagreements were predominantly between adjacent severity categories. The AI was able to generate erythema severity grades for images of darker skin tones that dermatologists typically would not rate and were marked as "unable to assess". Limitations: The validation set was small (41 images), severe cases (score 3) were underrepresented, one rater participated in both training annotation and validation scoring, and sample size was insufficient for robust stratification by skin tone or body site. Conclusion: The AI pipeline demonstrated dermatologist-level accuracy for erythema scoring, consistent moderate agreement for excoriation and lichenification, and a potential advantage in assessing erythema on darker skin tones. These findings support its potential as a standardized, objective tool for AD severity assessment. Prospective validation in larger, more diverse cohorts is warranted.

Vitiligo is an acquired pigmentary disorder of the skin and mucus membranes. Previous study has demonstrated that autologous cultured epithelial grafts (ACEG) is an effective treatment for stable vitiligo. However, extraction of full-thickness skin might result in scar formation at donor site, which have hindered the wider application of this technology, especially for patients requiring large-area transplantation. Hair follicle as a source of keratinocyte and melanocyte, could be potential source of cells for preparation of autologous cultured sheet. Through culture system optimization, we have demonstrated maintenance of undifferentiated hair follicle-derived cells in feeder-independent culture system. After expansion, the hair follicle cells were directed to differentiate into a multi-layered, epidermis-like sheet. Cell identity, viability, purity, genomic stability, and antiseptic testing for hair follicle-derived epithelial sheet (HFES) were evaluated to ensure its safety. Immunofluorescence staining showed that basal keratinocytes were the main cell type of the autologous HFES. Optimization of culture conditions leads to increased melanocyte proliferation and functionality. Transcriptomic analysis confirmed upregulation of melanosome maturation genes. The proportions of cells are also similar to composition of cells under physiological conditions. Transplantation of HFES to depigmented areas in patients with stable vitiligo results in skin repigmentation. This technology provides a novel therapeutic option for vitiligo management.

Chronic or excessive inflammation are hallmarks of many human diseases, but the endogenous factors that limit inflammatory responses are insufficiently characterized. We discovered a broad anti-inflammatory function of fibroblast growth factor 7 (FGF7), which is mediated via epithelial cels. FGF7 suppressed the expression of pro-inflammatory and immunomodulatory genes in cultured human keratinocytes in the absence or presence of pro-inflammatory stimuli and in a psoriasiform inflammation model in mice. Mechanistically, this involves an FGF7-FGF receptor 2 (FGFR2)-MAPK-Kruppel-like factor 4 (KLF4) signaling axis. FGF7 induced significant alterations in the KLF4 interactome in human keratinocytes and suppressed the transcriptional activity of KLF4 at its pro-inflammatory target genes. Concomitantly, expression of FGFR2 and downstream signaling components was promoted by KLF4, identifying a KLF4-dependent regulatory feedback loop that sustains anti-inflammatory FGF signaling. These results suggest activation of FGF7-FGFR2-KLF4 signaling as a strategy for the treatment of inflammatory diseases involving the skin or other epithelial tissues and highlight the role of epithelial cells in the control of inflammation.

Therapeutic blockade of IL-17 and TNF can effectively treat inflammatory skin diseases such as hidradenitis suppurativa and psoriasis, yet the relative importance of the different cell types that respond to IL-17 and TNF remains unresolved. Keratinocytes are viewed as the dominant effector cells, whereas fibroblasts have recently emerged as important contributors. In mice, topical imiquimod induces IL-17- and TNF-dependent skin inflammation and is frequently used to model psoriasis. Here, we demonstrate that intradermal injection of recombinant IL-17 and TNF elicits skin inflammation with features of hidradenitis suppurativa, including a gene expression program that is distinct from psoriasis and imiquimod-induced inflammation. Single-cell transcriptomic network analysis identified dermal fibroblasts as the dominant cell communication hub in hidradenitis suppurativa and in mice injected with IL-17 and TNF. In contrast, fibroblasts and keratinocytes both show strong network involvement in psoriasis and in mice challenged with imiquimod. Cell-type-specific deletion of IL-17 receptor A in mice revealed that imiquimod-induced inflammation depends equally on IL-17 signaling in fibroblasts and keratinocytes, whereas inflammation induced by intradermal IL-17 and TNF only requires fibroblasts to recognize IL-17 and is independent of keratinocyte IL-17 sensing. Single-cell transcriptomic analysis of these conditional knockout mice further demonstrated that keratinocytes and fibroblasts activate divergent and disease-dependent transcriptional programs following activation by IL-17. Together, these findings introduce a new conceptual framework wherein IL-17 signaling is routed through distinct cellular and molecular pathways depending on disease context and establish complementary experimental systems for interrogating type 17 skin inflammation.

A phase 2, randomised, double-blind, placebo controlled trial has been conducted in mild to moderate patchy alopecia areata (AA). This demonstrated significant and dose related improvements in hair regrowth with STS01, a controlled release, topical formulation of dithranol. Here we report the results of the Alopecia Areata Symptom Impact Scale (AASIS) that assesses the severity of symptoms, daily functioning and feelings. Similar to trials in severe AA, significant improvements in hair regrowth did not translate into significant health-related quality of life (HRQoL) improvements, even in patients with complete hair regrowth, although there was some treatment-related correlation between changes in AASIS scores from baseline and clinical assessment SALT scores. The use of current HRQoL methods or indeed new measures in development for future trials, will have considerable challenges: patients may not have a true baseline at entry, may develop coping mechanisms, and there may be a delay between physical and psychological improvement.

The primary route of SARS-CoV-2 entry is via respiratory epithelium. However, many COVID-19 patients developed dermatological lesions, and SARS-CoV-2 RNA has been detected in the patients skin. Inflammatory skin diseases, psoriasis and atopic dermatitis (AD), significantly increased the risk of COVID-19. To evaluate the potential role of skin in SARS-CoV-2 host interactions, we utilized 3D human skin organoids (HSO) generated from human epidermal keratinocytes, as well as neonatal skin explants. HSO were treated with cytokines involved in acute and chronic skin inflammation and cytokine storm in severe COVID-19 disease, TNF-, IL-6, IL-1{beta}, and IFN-{gamma}, individually and in combination. HSO were also treated with Th1 (TNF- + IL-17) and Th2 (IL-4 + IL-13) cocktails inducing pro-psoriasis and pro-AD HSO changes, respectively. All individual cytokines, and especially their combinations, elevated the expression of ACE2 and TMPRSS2 at mRNA/protein levels. The Th2 induced only TMPRSS2, the Th1 predominantly induced ACE2. Topically applied Spike-pseudotyped lentiviral Tomato reporter, which binds ACE2 similarly to SARS-CoV-2, successfully infected control and cytokine-treated HSO as well as neonatal skin explants. Cytokine treatment, especially TNF- + IL-6 + IL-1{beta} + IFN-{gamma} and the Th1, significantly increased viral entry. Transcriptomic analysis further revealed partial overlap between gene expression signatures induced by Spike-mediated entry in inflamed HSO and those observed in lung tissue from COVID-19 patients, supporting the biological relevance of skin models. Together, these findings demonstrate that inflammation enhances the permissiveness of human skin to SARS-CoV-2 entry, suggesting that the skin may represent a previously underappreciated interface in viral host interactions.

Background There are no licensed treatments for patients with mild to moderate patchy alopecia areata (AA). Objectives To evaluate the efficacy, safety and dose response of STS01, a novel nanoparticle controlled release, topical formulation of dithranol/Prosilic. Methods In a phase 2, double blind study, adult patients with mild to moderate AA (guideline 10% to 50% of scalp hair loss) were randomly assigned to STS01 at doses of 0.25%, 0.5%, 1%, 2% or placebo, daily for 6 months. The primary endpoints included the proportion of patients achieving a >=30% improvement in Severity of Alopecia Tool (SALT) score, and percentage change from baseline in SALT score. This minimum level of improvement is generally accepted as an indicator of the population likely to progress to complete regrowth Results A total of 155 patients were randomized and treated (placebo, n=32; STS01 groups, n=30 to 31). STS01 1% met the primary efficacy endpoint of >=30% SALT score improvement compared to placebo: 75.9% (95% CI, 60.3 to 91.4%) vs 36.7% (95% CI, 19.4 to 53.9%) at 6 months; p=0.0037. The least squares (LS) mean percentage change in SALT score from baseline to end of treatment showed a clear dose response relationship; STS01 0.5% was the minimally effective dose and 2% the maximum tolerated dose, and there was a statistically significant improvement in the STS01 1% group (minus 55.0% vs +0.6% with placebo; p<0.01). Significant improvements (p<0.05) in LS mean percentage changes from baseline in SALT scores were demonstrated in the STS01 1% group at 2 months (-28.6% vs 12.8%), 4 months (-57.2% vs 1.5%), and 6 months (minus 67.0% vs 0.6%). Clinical Global Impression improvement was reported in 72.0% of patients with STS01 1% vs 41.7% with placebo (p<0.05). The most commonly reported treatment emergent adverse events were skin irritation reactions, but were mostly mild (STS01: 56.7% to 71.0%; placebo: 21.9%) or moderate (STS01:13.3% to 35.5%; placebo: 0%) and manageable by reduced frequency of application. There were 15 skin-related discontinuations with STS01 (12.2%) and 2 (6.3%) with placebo. Conclusions STS01 demonstrated a clear dose response, with STS01 1% dose optimally more effective than placebo for hair regrowth with minimal tolerance concerns in mild to moderate patchy AA. Skin irritation reactions were generally manageable and there were no new safety signals. Further characterisation of the STS01 1% dose is planned in a phase 3 study. Chief Investigator AGM reports fees from Soterios Ltd. Chief Statistician DMF is an employee of Soterios Ltd. All other authors were Principal Investigators in the trial and their clinics were reimbursed for the work involved. Most also had sponsorship in the form of consultancies, investigational roles or lecturing roles on behalf of other Dermatological pharmaceutical companies

Skin infections are among the most common types of infections in the United States and pose a serious health threat. Effective resolution requires both bacterial clearance and tissue repair, however the mechanisms underlying tissue repair remain poorly understood. Here, we investigate the cellular responses in a murine model of S. aureus dermonecrotic skin infection using single cell transcriptomics. Within the infected lesions, we identified a major influx of heterogeneous neutrophils, including a distinct subtype expressing high levels of Spp1, the gene encoding osteopontin (OPN). Cell-cell interaction analysis identified Spp1 signaling as the most enriched pathway in infected tissue. In OPN knockout (KO) mice, transcriptomic profiling revealed a lower proportion of fibroblast and keratinocyte cells at the infection site, and a reduction in associated repair signaling pathways. In vivo, OPN KO mice developed significantly larger lesions during skin infection, whereas WT mice treated with recombinant OPN showed accelerated healing relative to untreated controls. Together, these findings highlight the importance of specific neutrophil subtypes and OPN signaling as critical mediators in infected tissue repair. Further our study suggests that modulating OPN may represent a promising therapeutic strategy to accelerate infected tissue repair.

Background In clinical trials for alopecia areata (AA) the treatment effect (percentage of hair loss) is estimated using the Severity of Alopecia Tool (SALT) score. Trials in patients with severe AA (>=50% hair loss) employed a local rating of the SALT score performed at trial sites by different investigators. However, in mild-to-moderate AA (<= 50% hair loss) where SALT scores are lower, potential inter rater variability and margin of error may compromise the results. Objectives To compare Centralised and Local measurement of hair loss in mild moderate AA. Methods In a Phase 2 clinical trial a centralised measurement of hair loss was performed from photographic images taken using a standardised protocol and professional camera equipment. Local scoring was also undertaken at screening/baseline for eligibility. We assessed: the repeatability of the central system (screening vs baseline values), the reproducibility of the central versus the local rating system and the potential impact of each method on the endpoints using a Monte-Carlo simulation method. Results There was good agreement and consistency of scoring with Central rating. This provided much smaller margins of error, 50% lower than Local rating. The simulations demonstrated that substituting Local rating for Central rating would result in a reduction of the likelihood of a statistically significant outcome by at least 50% depending on the SALT score defined clinical response endpoint. Conclusions Central rating is most appropriate in the Phase 2 learning stage of clinical development and provides an accurate representation of the quantity of hair loss, minimising error and ensuring consistency in measurements.

The human skin is repeatedly exposed to mechanical and environmental stress, particularly in common skin diseases such as eczema, and yet the determinants of recovery remain poorly understood. Using longitudinal, multimodal profiling of skin physiology, structure (Raman spectroscopy), and microbial communities (shotgun metagenomics), we investigated in a human cohort (n=36 subjects, x2 sites, x6 timepoints) how host-microbe interactions could jointly shape recovery. Despite baseline variability in physiological parameters, we established that our protocol enables a defined disruption of the stratum corneum. While recovery trajectories for host attributes were notably consistent across age groups and body sites, individual-specific differences in recovery timelines were observed. To assess the role of the skin microbiome, several key time-dependent changes in microbial species were identified including enrichment of select Cutibacterium and Staphylococcus species and depletion of Corynebacterium and Malassezia species. Clustering of microbiome stability profiles across subjects and sites identified 6 distinct groups which associate with varying host-recovery patterns and microbial functions. Finally, joint hazards modelling of recovery timing revealed significant contributions from microbial taxa, functions and stability groups, highlighting the under-appreciated role of host-microbial interactions in response to skin stress and in the recovery process.

Tolerogenic dendritic cells (TolDCs) are essential for immune tolerance and offer promise for treating autoimmune diseases. Despite the clinical evidence of their therapeutic potential, the key molecular pathways guiding their differentiation and tolerogenic phenotype remain elusive due to complex interactions identified in functional assays. Here we investigated the molecular profiles and regulatory programs underlying the functional status of tolerogenic dendritic cell populations in response to known tolerizing agents. We identified CD86 as a consistent and robust marker downregulated in tolerogenic state. Using CD86 blocking antibodies or CRISPR-mediated gene inactivation we demonstrated that CD86 is functionally required for TolDC-mediated suppression of T cell proliferation and cytokine secretion, establishing CD86 as both a consensus phenotypic and functional screening marker and a mechanistic regulator of tolerance. Leveraging CD86 as a scalable readout, we performed a pooled genome-wide CRISPR-Cas9 knockout screen to identify regulators of TolDC function. This approach uncovered UBE2L6 as a novel modulator that promotes the tolerogenic phenotype and restricts TolDC-mediated T cell activation. Mechanistically, UBE2L6 deficiency leads to coordinated upregulation of ISG15 and USP18, indicating a possible ISGylation-dependent pathway regulating CD86 expression and tolerogenic function. Together, this study identifies pathways that can be targeted to promote immune tolerance in immune-mediated inflammatory diseases. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=105 SRC="FIGDIR/small/713621v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@4c3636org.highwire.dtl.DTLVardef@17b27e5org.highwire.dtl.DTLVardef@783d95org.highwire.dtl.DTLVardef@1337ce_HPS_FORMAT_FIGEXP M_FIG C_FIG

BackgroundCutaneous melanoma (CM) is an aggressive skin cancer with rising incidence, representing a growing public health concern. Despite the remarkable success of immune-checkpoint inhibitors (ICIs) in the management of advanced disease, mortality remains high due to therapy resistance. Identifying reliable prognostic and predictive biomarkers is therefore essential to improve patient stratification, optimize treatment selection, and minimize unnecessary toxicity. MethodsWe comprehensively profiled the circulating immune landscape of 54 treatment-naive CM patients by integrating flow cytometry immunophenotyping with clinicopathological data, and performed tumor gene expression analysis in a subset of 26 patients. ResultsElevated HLA-DR and CD69 expression on circulating CD4+ T cells, together with reduced circulating CD8+ T cell frequency, emerged as candidate prognostic biomarkers associated with improved survival. Prognostic models combining these immune variables with clinical covariates accurately stratified patients by overall survival (89.5% sensitivity, 72.7% specificity; AUC = 0.872, p < 0.0001) and progression/recurrence risk (75% sensitivity and 71.4% specificity; AUC = 0.763, p = 0.001). In a subset of 43 patients subsequently treated with ICIs, elevated baseline HLA-DR and CD69 expression on circulating CD4+ T cells was also associated with therapeutic benefit. A predictive model integrating these markers with clinical covariates achieved good discriminatory performance (65.2% sensitivity, 88.9% specificity; AUC = 0.775, p = 0.0027). Tumor gene expression profiling supported the role of IFN-{gamma}-related signatures, previously linked to ICI response, as complementary prognostic and predictive tools. ConclusionThese findings highlight systemic CD4+ T cell activation status as a promising, easily measurable biomarker in CM, laying the foundation for future strategies to refine patient stratification and guiding immunotherapy decisions.

ImportanceDigital Phenotyping (DP) utilizes digital technologies to assess observable phenotypic traits, enhancing our understanding of various disease states. It aims to support equitable interventions through diverse digital data, fostering inclusivity by integrating various digital devices. Central to DP is identifying digital biomarkers (DBMs), which offer real-time health monitoring and personalized insights into disease conditions. Applying machine learning (ML) techniques on non-invasive signals sets the ideal platform for precision medicine application, especially in rare diseases (RDs). People living with rare diseases (PLWRD) often face significant challenges in receiving timely and accurate diagnoses, leading to what is known as a diagnostic odyssey. Digital phenotyping (DP) offers a promising solution by leveraging advanced technology, such as 3D facial photography, to capture unique digital signatures associated with various rare diseases. This innovative approach not only aids in the identification of these conditions but also facilitates the detection of digital biomarkers (DBM). These biomarkers enable healthcare providers to monitor the progression of the disease over time, enhancing patient care and potentially shortening the duration of the diagnostic odyssey. By utilizing DP and DBMs, we can improve both the diagnosis and management of RDs, ultimately leading to better health outcomes for affected participants. ObjectiveTo identify whether DBMs can be identified by DP utilizing 3D facial imaging techniques in outpatient settings in participants with RDs. The primary objective of this study was to determine if specific facial measurements in participants with RD who experience transient episodes of facial swelling (oedema) differ from established ethnically matched norms. The secondary objective was to assess peri-orbital and/or facial swelling as a potential biomarker for identifying flare-ups in hereditary angioedema (HAE). Design, setting, and participantsThis multicentre observational study was conducted in 3 hospitals in Singapore. The eligible participants were male and female RD participants of various age groups. The study duration was 4 years and 8 months. Interventions: Twenty participants of Chinese genetic ancestry were photographed using a 3D camera. Additionally, two participants with hereditary angioedema (HAE) were photographed during acute stages of disease flare-ups. Main outcomes and measuresThe obtained facial scans of participants (that included participants with HAE in non-acute phase) were plotted using Artificial Intelligence-powered software - Cliniface. The growth curves and facial landmarks obtained were compared against the growth curves of normal RD-unaffected individuals of Chinese genetic ancestry. The two participants with HAE were photographed qualitatively over a longer period of time, and their scans were plotted, yielding growth curves. ResultsDistinct facial markers such as periorbital swelling were identified in two qualitatively assessed HAE participants during flare-up stages. This provides an opportunity to explore and validate further if these facial signatures in a disease condition can be assigned as DBM for HAE. Conclusions and relevanceThis study explores the utility of 3D facial analysis as a DBM in rare diseases such as HAE. Applying non-invasive signals coupled with AI may open new vistas for precision medicine in real-world settings. The individual measurements that yielded small p-values demonstrate significant relevance and potential utility. These findings offer preliminary objective evidence that supports existing subjective reports of facial features in the literature. Additionally, while DPs diagnostic capabilities may be limited, it successfully identified DBM, which could facilitate disease monitoring in conditions such as HAE. Author SummaryRare diseases pose a significant challenge to all stakeholders, including clinicians, patients families, care providers, and the healthcare system. Diagnostic delays are integral and impose a massive financial and emotional burden on everyone involved in care delivery, beyond the patients themselves. A universally acceptable, scalable, and replicable non-invasive mechanism to detect distinct biomarkers associated with a rare disease can help identify the diseases signs and symptoms far earlier and ease the burden. An easy-to-deploy approach could be 3D facial imaging of patients with rare diseases, which are associated with distinct or subtle facial changes at different stages of disease progression. A rare disease, such as hereditary angioedema, which is known to exhibit facial swelling in patients during the acute disease state, is a prime example. The facial changes can be identified and assigned as specific disease markers, also known as facial biomarkers. These facial biomarkers can be identified and measured using 3D facial imaging when patients present to the clinic. Subsequently, these initial signs can be correlated clinically to establish a firm diagnosis earlier than traditional approaches.

We generated interferon signaling reporters in human and mouse melanoma cells and observed heterogeneity in interferon responses among cells in the same tumors. This was marked by inflamed regions within primary tumors that contained increased numbers of interferon-expressing macrophages/monocytes and elevated type I interferon signaling in melanoma cells. Melanoma cells that expressed GFP-Interferon stimulated gene 15 (ISG15) or GFP-Interferon-induced protein with tetraticopeptide repeats 3 (IFIT3) fusion reporters exhibited a profoundly increased ability to form metastatic tumors as compared to GFP-ISG15- melanoma cells from the same tumors, particularly when transplanted into immunocompetent mice. The increased metastatic potential of GFP-ISG15+ cells was driven partly by increased CD47 expression, which protected metastasizing cells from phagocytosis by macrophages. Macrophages are thus a double-edged sword, inhibiting the development of metastatic disease by phagocytosing disseminated melanoma cells, but promoting the emergence of a hyper-metastatic subpopulation of cells in inflamed regions of primary tumors as a consequence of sustained interferon production. One sentence summarySustained interferon exposure within inflamed regions of primary tumors dramatically increases the metastatic potential of a subpopulation of melanoma cells, partly by promoting CD47 expression, which protects against phagocytosis by macrophages.

Humans are tight-skinned mammals who typically fail to regenerate large full-thickness skin wounds, instead healing with substantial scarring and concomitant loss of function. Mechanical context is a major determinant of this outcome: elevated tissue tension or stiffness promotes fibrotic repair associated with hypertrophic or keloid scarring. Accordingly, regenerative medicine research has relied on diverse animal models to understand scar development and skin regeneration. Loose-skinned mammals exhibit greater regeneration ability. Furthermore, spiny mouse skin is significantly less stiff and associated with enhanced regenerative ability. Interestingly, this skin wound stiffness can be modulated to shift healing toward more regenerative or more fibrotic trajectories. Despite of this progress, the restoration of normal skin architecture after large-full thickness injury has not been elucidated in tight-skinned mammals. Can large full-thickness wounds regenerate with minimal scarring in tight-skinned mammals? Here we show the tight-skinned mammal Frasers Dolphin regenerates de novo a complex rete ridge architecture with associated vasculature and minimal scar following large full-thickness wound healing. Counterintuitively, this skin regeneration occurs in an aqueous, high-shear stress and high-tension environment. Complete rete ridge regeneration in tight-skinned mammals has not been documented and not observed in humans except in utero. This unique ability to rebuild elaborate rete ridges under tension is an opportunity to uncover molecular, cellular, and tissue-level mechanisms that enable regenerative wound healing in a mechanical regime typically associated with fibrosis.