tauFisher predicts circadian time from a single sample of bulk and single-cell pseudobulk transcriptomic data.

As the circadian clock regulates fundamental biological processes, disrupted clocks are often observed in patients and diseased tissues. Determining the circadian time of the patient or the tissue of focus is essential in circadian medicine and research. Here we present tauFisher, a computational pipeline that accurately predicts circadian time from a single transcriptomic sample by finding correlations between rhythmic genes within the sample. We demonstrate tauFisher's performance in adding timestamps to both bulk and single-cell transcriptomic samples collected from multiple tissue types and experimental settings. Application of tauFisher at a cell-type level in a single-cell RNAseq dataset collected from mouse dermal skin implies that greater circadian phase heterogeneity may explain the dampened rhythm of collective core clock gene expression in dermal immune cells compared to dermal fibroblasts. Given its robustness and generalizability across assay platforms, experimental setups, and tissue types, as well as its potential application in single-cell RNAseq data analysis, tauFisher is a promising tool that facilitates circadian medicine and research.

Suppression of TCF4 promotes a ZC3H12A-mediated self-sustaining inflammatory feedback cycle involving IL-17RA/IL-17RE epidermal signaling.

IL-17C is an epithelial cell-derived proinflammatory cytokine whose transcriptional regulation remains unclear. Analysis of the IL17C promoter region identified TCF4 as putative regulator, and siRNA knockdown of TCF4 in human keratinocytes (KCs) increased IL17C. IL-17C stimulation of KCs (along with IL-17A and TNF-α stimulation) decreased TCF4 and increased NFKBIZ and ZC3H12A expression in an IL-17RA/RE-dependent manner, thus creating a feedback loop. ZC3H12A (MCPIP1/Regnase-1), a transcriptional immune-response regulator, also increased following TCF4 siRNA knockdown, and siRNA knockdown of ZC3H12A decreased NFKBIZ, IL1B, IL36G, CCL20, and CXCL1, revealing a proinflammatory role for ZC3H12A. Examination of lesional skin from the KC-Tie2 inflammatory dermatitis mouse model identified decreases in TCF4 protein concomitant with increases in IL-17C and Zc3h12a that reversed following the genetic elimination of Il17c, Il17ra, and Il17re and improvement in the skin phenotype. Conversely, interference with Tcf4 in KC-Tie2 mouse skin increased Il17c and exacerbated the inflammatory skin phenotype. Together, these findings identify a role for TCF4 in the negative regulation of IL-17C, which, alone and with TNF-α and IL-17A, feed back to decrease TCF4 in an IL-17RA/RE-dependent manner. This loop is further amplified by IL-17C-TCF4 autocrine regulation of ZC3H12A and IL-17C regulation of NFKBIZ to promote self-sustaining skin inflammation.

Topical Ruxolitinib in the Treatment of Necrobiosis Lipoidica: A Prospective, Open-Label Study.

Necrobiosis lipoidica (NL) is a rare granulomatous disease. There are few effective treatments for NL. We sought to investigate the efficacy and safety of the Jak1/2 inhibitor, ruxolitnib, in the treatment of NL and identify the biomarkers associated with the disease and treatment response. We conducted an open-label, phase 2 study of ruxolitinib in 12 patients with NL. We performed transcriptomic analysis of tissue samples before and after treatment. At week 12, the mean NL lesion score decreased by 58.2% (SD = 28.7%, P = .003). Transcriptomic analysis demonstrated enrichment of type I and type II IFN pathways in baseline disease. Weighted gene coexpression network analysis demonstrated post-treatment changes in IFN pathways with key hub genes IFNG and signal transducer and activator of transcription 1 gene STAT1. Limitations include small sample size and a study group limited to patients with <10% body surface area. In conclusion, ruxolitinib is an effective treatment for NL and targets the key pathogenic mediators of the disease.

Lupus dermal fibroblasts are proinflammatory and exhibit a profibrotic phenotype in scarring skin disease.

Fibroblasts are stromal cells known to regulate local immune responses important for wound healing and scar formation; however, the cellular mechanisms driving damage and scarring in patients with cutaneous lupus erythematosus (CLE) remain poorly understood. Dermal fibroblasts in patients with systemic lupus erythematosus (SLE) experience increased cytokine signaling in vivo, but the effect of inflammatory mediators on fibroblast responses in nonscarring versus scarring CLE subtypes is unclear. Here, we examined responses to cytokines in dermal fibroblasts from nonlesional skin of 22 patients with SLE and CLE and 34 individuals acting as healthy controls. Notably, inflammatory cytokine responses were exaggerated in SLE fibroblasts compared with those from individuals acting as healthy controls. In lesional CLE biopsies, these same inflammatory profiles were reflected in single-cell RNA-Seq of SFRP2+ and inflammatory fibroblast subsets, and TGF-ß was identified as a critical upstream regulator for inflammatory fibroblasts in scarring discoid lupus lesions. In vitro cytokine stimulation of nonlesional fibroblasts from patients who scar from CLE identified an upregulation of collagens, particularly in response to TGF-ß, whereas inflammatory pathways were more prominent in nonscarring patients. Our study revealed that SLE fibroblasts are poised to hyperrespond to inflammation, with differential responses among patients with scarring versus nonscarring disease, providing a potential skin-specific target for mitigating damage.

HERC6 regulates STING activity in a sex-biased manner through modulation of LATS2/VGLL3 Hippo signaling.

Interferon (IFN) activity exhibits a gender bias in human skin, skewed toward females. We show that HERC6, an IFN-induced E3 ubiquitin ligase, is induced in human keratinocytes through the epidermal type I IFN; IFN-κ. HERC6 knockdown in human keratinocytes results in enhanced induction of interferon-stimulated genes (ISGs) upon treatment with a double-stranded (ds) DNA STING activator cGAMP but not in response to the RNA-sensing TLR3 agonist. Keratinocytes lacking HERC6 exhibit sustained STING-TBK1 signaling following cGAMP stimulation through modulation of LATS2 and TBK1 activity, unmasking more robust ISG responses in female keratinocytes. This enhanced female-biased immune response with loss of HERC6 depends on VGLL3, a regulator of type I IFN signature. These data identify HERC6 as a previously unrecognized negative regulator of ISG expression specific to dsDNA sensing and establish it as a regulator of female-biased immune responses through modulation of STING signaling.

Comparative analyses of Netherton syndrome patients and Spink5 conditional knock-out mice uncover disease-relevant pathways.

Netherton syndrome (NS) is a rare skin disease caused by loss-of-function mutations in the serine peptidase inhibitor Kazal type 5 (SPINK5) gene. Disease severity and the lack of efficacious treatments call for a better understanding of NS mechanisms. Here we describe a novel and viable, Spink5 conditional knock-out (cKO) mouse model, allowing to study NS progression. By combining transcriptomics and proteomics, we determine a disease molecular profile common to mouse models and NS patients. Spink5 cKO mice and NS patients share skin barrier and inflammation signatures defined by up-regulation and increased activity of proteases, IL-17, IL-36, and IL-20 family cytokine signaling. Systemic inflammation in Spink5 cKO mice correlates with disease severity and is associated with thymic atrophy and enlargement of lymph nodes and spleen. This systemic inflammation phenotype is marked by neutrophils and IL-17/IL-22 signaling, does not involve primary T cell immunodeficiency and is independent of bacterial infection. By comparing skin transcriptomes and proteomes, we uncover several putative substrates of tissue kallikrein-related proteases (KLKs), demonstrating that KLKs can proteolytically regulate IL-36 pro-inflammatory cytokines. Our study thus provides a conserved molecular framework for NS and reveals a KLK/IL-36 signaling axis, adding new insights into the disease mechanisms and therapeutic targets.

Significance of stress keratin expression in normal and diseased epithelia.

A group of keratin intermediate filament genes, the type II KRT6A-C and type I KRT16 and KRT17, are deemed stress responsive as they are induced in keratinocytes of surface epithelia in response to environmental stressors, in skin disorders (e.g., psoriasis) and in carcinomas. Monitoring stress keratins is widely used to identify keratinocytes in an activated state. Here, we analyze single-cell transcriptomic data from healthy and diseased human skin to explore the properties of stress keratins. Relative to keratins occurring in healthy skin, stress-induced keratins are expressed at lower levels and show lesser type I-type II pairwise regulation. Stress keratins do not "replace" the keratins expressed during normal differentiation nor reflect cellular proliferation. Instead, stress keratins are consistently co-regulated with genes with roles in differentiation, inflammation, and/or activation of innate immunity at the single-cell level. These findings provide a roadmap toward explaining the broad diversity and contextual regulation of keratins.

Decline in Cancer Diagnoses during the 'Zero COVID' Policy in Hong Kong: Indirect Spillover Impact of the COVID-19 Pandemic.

AIMS: Despite a largely successful 'zero COVID' policy in 2020, the COVID-19 pandemic disrupted routine cancer services in the city of Hong Kong. The aims of this study were to examine the trends in cancer incidence before and during the COVID-19 pandemic and estimate missed cancer diagnoses. MATERIALS AND METHODS: We used population-based data from the Hong Kong Cancer Registry 1983-2020 to examine the trends of age- and sex-standardised cancer incidence before and during the COVID-19 pandemic. We applied: (i) the annual average percentage change (AAPC) calculated using the Joinpoint regression model and (ii) the autoregressive integrated moving average (ARIMA) model to forecast cancer incidence rates in 2020. Missed cancer diagnoses in 2020 were estimated by comparing forecasted incidence rates to reported rates. A subgroup analysis was conducted by sex, age and cancer site. RESULTS: The cancer incidence in Hong Kong declined by 4.4% from 2019 to 2020 (male 8.1%; female 1.1%) compared with the long-term AAPC of 0.5% from 2005 to 2019 (95% confidence interval 0.3, 0.7). The gap between the reported and forecasted incidence for 2020 ranged from 5.1 to 5.7% (male 8.5%, 9.8%; female 2.3%, 3.5%). We estimated 1525-1596 missed cancer diagnoses (ARIMA estimate -98, 3148; AAPC 514, 1729) in 2020. Most missed diagnoses were in males (ARIMA 1361 [327, 2394]; AAPC 1401 [1353, 1460]), with an estimated 479-557 missed cases of colorectal cancer (ARIMA 112, 837; AAPC 518, 597) and 256-352 missed cases of prostate cancer (AAPC 231, 280; ARIMA 110, 594). CONCLUSION: The incidence of new cancer diagnoses declined in 2020 contrary to the long-term increase over the previous decades. Significantly lower diagnoses than expected were observed in males, particularly for colorectal and prostate cancers. Fewer reported cancer cases indicate missed diagnoses and could lead to delayed treatment that could impact future health outcomes.

Differential effects of teriparatide, denosumab and zoledronate on hip structural and mechanical parameters in osteoporosis; a real-life study.

PURPOSE: The aim of this study was to evaluate changes in hip geometry parameters following treatment with teriparatide (TPD), denosumab (Dmab) and zoledronate (ZOL) in real-life setting. METHODS: We studied 249 patients with osteoporosis (OP) with mean [SD] age of 71.5 [11.1] years divided into 3 treatment groups; Group A received TPD; n = 55, Group B (Dmab); n = 116 and Group C (ZOL); n = 78 attending a routine metabolic bone clinic. Bone mineral density (BMD) was measured by DXA at the lumbar spine (LS), total hip (TH) and femoral neck (FN) prior to treatment and after 2 years (Group A), after a mean treatment duration of 3.3 [1.3] years (Group B) and after 1, 2 and 3 doses of ZOL (Group C) to assess treatment response. Hip structural analysis (HSA) was carried out retrospectively from DXA-acquired femur images at the narrow neck (NN), the intertrochanter (IT) and femoral shaft (FS). RESULTS: Changes in parameters of hip geometry and mechanical strength were seen in the following treatment. Percentage change in cross-sectional area (CSA): 3.56[1.6] % p = 0.01 and cross-sectional moment of inertia (CSMI): 4.1[1.8] % p = 0.029 increased at the NN only in Group A. Improvement in HSA parameters at the IT were seen in group B: CSA: 3.3[0.67]% p < 0.001, cortical thickness (Co Th): 2.8[0.78]% p = 0.001, CSMI: 5.9[1.3]% p < 0.001, section modulus (Z):6.2[1.1]% p < 0.001 and buckling ratio (BR): - 3.0[0.86]% p = 0.001 with small changes at the FS: CSA: 1.2[0.4]% p = 0.005, Z:1.6 [0.76]%, p = 0.04. Changes at the IT were also seen in Group C (after 2 doses): CSA: 2.5[0.77]% p = 0.017, Co Th: 2.4[0.84]% p = 0.012, CSMI: 3.9[1.3]% p = 0.017, Z:5.2[1.16]% p < 0.001 and BR: - 3.1[0.88]% p = 0.001 and at the NN (following 3 doses): outer diameter (OD): 4.0[1.4]% p = 0.0005, endocortical diameter(ED): 4.3[1.67% p = 0.009, CSA:5.2[1.8]% p = 0.003, CSMI: 9.3[3.8]% p = 0.019. CONCLUSIONS: Analysis of the effect of OP therapies on hip geometry is useful in understanding the mechanisms of their anti-fracture effect and may provide additional information on their efficacy.

Systems-based identification of the Hippo pathway for promoting fibrotic mesenchymal differentiation in systemic sclerosis.

Systemic sclerosis (SSc) is a devastating autoimmune disease characterized by excessive production and accumulation of extracellular matrix, leading to fibrosis of skin and other internal organs. However, the main cellular participants in SSc skin fibrosis remain incompletely understood. Here using differentiation trajectories at a single cell level, we demonstrate a dual source of extracellular matrix deposition in SSc skin from both myofibroblasts and endothelial-to-mesenchymal-transitioning cells (EndoMT). We further define a central role of Hippo pathway effectors in differentiation and homeostasis of myofibroblast and EndoMT, respectively, and show that myofibroblasts and EndoMTs function as central communication hubs that drive key pro-fibrotic signaling pathways in SSc. Together, our data help characterize myofibroblast differentiation and EndoMT phenotypes in SSc skin, and hint that modulation of the Hippo pathway may contribute in reversing the pro-fibrotic phenotypes in myofibroblasts and EndoMTs.

Oral Baricitinib in the Treatment of Cutaneous Lichen Planus.

Background: Cutaneous lichen planus (LP) is a recalcitrant, difficult-to-treat, inflammatory skin disease characterized by pruritic, flat-topped, violaceous papules on the skin. Baricitinib is an oral Janus kinase (JAK) 1/2 inhibitor that interrupts the signaling pathway of interferon (IFN)-γ, a cytokine implicated in the pathogenesis of LP. Methods: In this phase II trial, twelve patients with cutaneous LP received baricitinib 2 mg daily for 16 weeks, accompanied by in-depth spatial, single-cell, and bulk transcriptomic profiling of pre-and post-treatment samples. Results: An early and sustained clinical response was seen with 83.3% of patients responsive at week 16. Our molecular data identified a unique, oligoclonal IFN-γ, CD8+, CXCL13+ cytotoxic T-cell population in LP skin and demonstrate a rapid decrease in interferon signature within 2 weeks of treatment, most prominent in the basal layer of the epidermis. Conclusion: This study demonstrates the efficacy and molecular mechanisms of JAK inhibition in LP. Trial Registration Number : NCT05188521.

Single-cell profiling of prurigo nodularis demonstrates immune-stromal crosstalk driving profibrotic responses and reversal with nemolizumab.

BACKGROUND: Prurigo nodularis (PN) is a chronic neuroimmune skin disease characterized by bilaterally distributed pruritic hyperkeratotic nodules on extremities and trunk. Neuroimmune dysregulation and chronic scratching are believed to both induce and maintain the characteristic lesions. OBJECTIVES: This study sought to provide a comprehensive view of the molecular pathogenesis of PN at the single-cell level to identify and outline key pathologic processes and the cell types involved. Features that distinguish PN skin from the skin of patients with atopic dermatitis were of particular interest. We further aimed to determine the impact of the IL31RA antagonist, nemolizumab, and its specificity at the single-cell level. METHODS: Single-cell RNA-sequencing of skin from 15 healthy donors and nonlesional and lesional skin from 6 patients each with PN and atopic dermatitis, combined with spatial-sequencing using the 10x Visium platform. Integration with bulk RNA-sequencing data from patients treated with nemolizumab. RESULTS: This study demonstrates that PN is an inflammatory skin disease characterized by both keratinocyte proliferation and activation of profibrotic responses. This study also demonstrates that the COL11A1+ fibroblast subset is a major contributor to fibrosis and is predominantly found in the papillary dermis of PN skin. Activation of fibrotic responses is the main distinguishing feature between PN and atopic dermatitis skin. This study further shows the broad effect of nemolizumab on PN cell types, with a prominent effect driving COL11A1+ fibroblast and keratinocyte responses toward normal. CONCLUSIONS: This study provides a high-resolution characterization of the cell types and cellular processes activated in PN skin, establishing PN as a chronic fibrotic inflammatory skin disease. It further demonstrates the broad effect of nemolizumab on pathological processes in PN skin.

Single-cell sequencing reveals Hippo signaling as a driver of fibrosis in hidradenitis suppurativa.

Hidradenitis suppurativa (HS) is a chronic inflammatory disease characterized by abscesses, nodules, dissecting/draining tunnels, and extensive fibrosis. Here, we integrate single-cell RNA sequencing, spatial transcriptomics, and immunostaining to provide an unprecedented view of the pathogenesis of chronic HS, characterizing the main cellular players and defining their interactions. We found a striking layering of the chronic HS infiltrate and identified the contribution of 2 fibroblast subtypes (SFRP4+ and CXCL13+) in orchestrating this compartmentalized immune response. We further demonstrated the central role of the Hippo pathway in promoting extensive fibrosis in HS and provided preclinical evidence that the profibrotic fibroblast response in HS can be modulated through inhibition of this pathway. These data provide insights into key aspects of HS pathogenesis with broad therapeutic implications.

Monogenic etiologies of persistent human papillomavirus infections: A comprehensive systematic review.

PURPOSE: Persistent human papillomavirus infection (PHPVI) causes cutaneous, anogenital, and mucosal warts. Cutaneous warts include common warts, Treeman syndrome, and epidermodysplasia verruciformis, among others. Although more reports of monogenic predisposition to PHPVI have been published with the development of genomic technologies, genetic testing is rarely incorporated into clinical assessments. To encourage broader molecular testing, we compiled a list of the various monogenic etiologies of PHPVI. METHODS: We conducted a systematic literature review to determine the genetic, immunological, and clinical characteristics of patients with PHPVI. RESULTS: The inclusion criteria were met by 261 of 40,687 articles. In 842 patients, 83 PHPVI-associated genes were identified, including 42, 6, and 35 genes with strong, moderate, and weak evidence for causality, respectively. Autosomal recessive inheritance predominated (69%). PHPVI onset age was 10.8 ± 8.6 years, with an interquartile range of 5 to 14 years. GATA2,IL2RG,DOCK8, CXCR4, TMC6, TMC8, and CIB1 are the most frequently reported PHPVI-associated genes with strong causality. Most genes (74 out of 83) belong to a catalog of 485 inborn errors of immunity-related genes, and 40 genes (54%) are represented in the nonsyndromic and syndromic combined immunodeficiency categories. CONCLUSION: PHPVI has at least 83 monogenic etiologies and a genetic diagnosis is essential for effective management.

Genetic and Immunological Pathogenesis of Atopic Dermatitis.

Type 2 immune-mediated diseases give a clear answer to the issue of nature (genetics) versus nurture (environment). Both genetics and environment play vital complementary roles in the development of atopic dermatitis (AD). As a key component of the atopic march, AD demonstrates the interactive nature of genetic and environmental contributions to atopy. From sequence variants in the epithelial barrier gene encoding FLG to the hygiene hypothesis, AD combines a broad array of contributions into a single syndrome. This review will focus on the genetic contribution to AD and where genetics facilitates the elicitation or enhancement of AD pathogenesis.

Large-scale functional inference for skin-expressing lncRNAs using expression and sequence information.

Long noncoding RNAs (lncRNAs) regulate the expression of protein-coding genes and have been shown to play important roles in inflammatory skin diseases. However, we still have limited understanding of the functional impact of lncRNAs in skin, partly due to their tissue specificity and lower expression levels compared with protein-coding genes. We compiled a comprehensive list of 18,517 lncRNAs from different sources and studied their expression profiles in 834 RNA-Seq samples from multiple inflammatory skin conditions and cytokine-stimulated keratinocytes. Applying a balanced random forest to predict involvement in biological functions, we achieved a median AUROC of 0.79 in 10-fold cross-validation, identifying significant DNA binding domains (DBDs) for 39 lncRNAs. G18244, a skin-expressing lncRNA predicted for IL-4/IL-13 signaling in keratinocytes, was highly correlated in expression with F13A1, a protein-coding gene involved in macrophage regulation, and we further identified a significant DBD in F13A1 for G18244. Reflecting clinical implications, AC090198.1 (predicted for IL-17 pathway) and AC005332.6 (predicted for IFN-γ pathway) had significant negative correlation with the SCORAD metric for atopic dermatitis. We also utilized single-cell RNA and spatial sequencing data to validate cell type specificity. Our research demonstrates lncRNAs have important immunological roles and can help prioritize their impact on inflammatory skin diseases.

Serial KinderMiner (SKiM) discovers and annotates biomedical knowledge using co-occurrence and transformer models.

BACKGROUND: The PubMed archive contains more than 34 million articles; consequently, it is becoming increasingly difficult for a biomedical researcher to keep up-to-date with different knowledge domains. Computationally efficient and interpretable tools are needed to help researchers find and understand associations between biomedical concepts. The goal of literature-based discovery (LBD) is to connect concepts in isolated literature domains that would normally go undiscovered. This usually takes the form of an A-B-C relationship, where A and C terms are linked through a B term intermediate. Here we describe Serial KinderMiner (SKiM), an LBD algorithm for finding statistically significant links between an A term and one or more C terms through some B term intermediate(s). The development of SKiM is motivated by the observation that there are only a few LBD tools that provide a functional web interface, and that the available tools are limited in one or more of the following ways: (1) they identify a relationship but not the type of relationship, (2) they do not allow the user to provide their own lists of B or C terms, hindering flexibility, (3) they do not allow for querying thousands of C terms (which is crucial if, for instance, the user wants to query connections between a disease and the thousands of available drugs), or (4) they are specific for a particular biomedical domain (such as cancer). We provide an open-source tool and web interface that improves on all of these issues. RESULTS: We demonstrate SKiM's ability to discover useful A-B-C linkages in three control experiments: classic LBD discoveries, drug repurposing, and finding associations related to cancer. Furthermore, we supplement SKiM with a knowledge graph built with transformer machine-learning models to aid in interpreting the relationships between terms found by SKiM. Finally, we provide a simple and intuitive open-source web interface ( https://skim.morgridge.org ) with comprehensive lists of drugs, diseases, phenotypes, and symptoms so that anyone can easily perform SKiM searches. CONCLUSIONS: SKiM is a simple algorithm that can perform LBD searches to discover relationships between arbitrary user-defined concepts. SKiM is generalized for any domain, can perform searches with many thousands of C term concepts, and moves beyond the simple identification of an existence of a relationship; many relationships are given relationship type labels from our knowledge graph.