tbuckley/cabot-search huggingface dataset:
P30 Exposure to particulate matter induces structural and pigmentary changes to human skin
Authors: S Deyvapriya, Poonam Halai, Orsolya Kiss, Bezaleel Mambwe, Rachel Watson, Abigail Langton
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims The majority of the world’s population reside in places with high levels of air pollution. The impact of air pollution on human health has been extensively studied and demonstrates a significant correlation between particulate matter (PM) and risk of cardiovascular and respiratory diseases. However, little is known about the impact of air pollution on human skin health. Here, we develop an ex vivo skin model to investigate cutaneous changes induced by the topical application of PM. Methods PM (0, 0.5, 3, 6, 10, 20 and 40 μg mL−1) was topically applied to Genoskin® NativeSkin Access® (Genoskin, Salem, MA, USA) human ex vivo skin samples of White European ancestry (n = 3) for 8 days, with reapplication on day 4. Resultant samples were assessed using haematoxylin and eosin staining (for morphometric analyses), immunofluorescence for Ki67 and terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) (to determine cellular turnover), while pigmentary changes were analysed by colourimetry, Warthin–Starry (WS) staining and immunohistochemistry for the melanin markers tyrosinase (TYR), gp100 and c-kit. Results Morphological assessment revealed that epidermal thickness was significantly reduced following exposure to higher doses of PM (r = −0.9163, P &amp;lt; 0.01). In addition, there was reduced epidermal cell proliferation (Ki67) and increased apoptosis (TUNEL) at higher doses of PM. Exposure to PM resulted in a dose-dependent decrease in individual typology angle, which is indicative of skin colour becoming darker (r = −0.7895, P &amp;lt; 0.05). Furthermore, WS staining demonstrated significantly increased epidermal melanin (r = 0.9583, P &amp;lt; 0.01) and significantly increased expression of melanin markers TYR (P &amp;lt; 0.01), gp100 (P &amp;lt; 0.001) and c-kit (P &amp;lt; 0.01) in response to PM exposure. Conclusions This study confirms that PM induces pigmentary changes to ex vivo human skin by increasing epidermal melanin. Increased expression of c-kit in response to PM exposure suggests that increased pigmentation occurs owing to the upregulation of the microphthalmia-associated transcription factor signalling pathway. Further investigation of the mechanistic drivers of increased pigmentation is warranted.
P18 Investigating the role of the microbiome in chronic wound skin barrier reformation
Authors: Alice Cannon, Holly Wilkinson, Michelle Rudden, Mat Hardman
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims The integrity of the skin barrier plays a pivotal role in dermatological health, particularly in elderly and individuals with diabetes who are more susceptible to injury. Our skin barrier is also in direct contact with the microbiota, a diverse community of microorganisms that play wide-ranging roles in cutaneous function. Although microbial dysbiosis has been observed in elderly and diabetic skin, few studies have investigated the mechanistic links between skin barrier perturbations and the microbiome. The aim of this study was to compare barrier reformation in healthy and pathological (chronic wound) skin and determine the role of microbial dysbiosis in skin barrier perturbations. Methods Here, we developed a novel human ex vivo skin barrier model to simulate skin barrier disruption in healthy and pathological skin. Following tape stripping (n = 6 per donor type), skin barrier disruption was confirmed by measuring transepidermal water loss (TEWL). Skin barrier reformation was assessed following culture using haematoxylin and eosin staining, Nile red (lipid expression and distribution), and immunohistochemistry for skin barrier proteins (e.g. claudins, keratins). Secreted products from mixed bacterial communities were also applied to healthy tape-stripped skin to assess the influence of a healthy vs. pathological microbiome on skin barrier reformation. Results Pathological skin showed higher TEWL following barrier disruption, in addition to delayed barrier reformation and altered skin barrier protein expression. Moreover, secreted products from a pathological microbiome, with high proportions of Staphylococcus aureus, significantly delayed skin barrier reformation. Conclusions Collectively, our data provide a novel skin barrier reformation model to assess the effect of cutaneous pathology and microbial dysbiosis on skin barrier repair. Further understanding of the role of the microbiome in skin barrier function is integral to reducing infections and barrier perturbation.
O14 Monocytes and macrophages in the melanoma tumour microenvironment express Fc receptors which can be engaged with monoclonal antibodies of different isotypes
Authors: Rebecca Adams, Gabriel Osborn, Roman Laddach, Melanie Grandits, Alicia Chenoweth, Victoria Sanz-Moreno, Katie Lacy, Sophia Karagiannis
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims Melanoma is the deadliest form of skin cancer and has an increasing worldwide incidence. It is well established that the melanoma tumour microenvironment (TME) has a large immune cell infiltrate, including a prominent monocytic compartment. Monocytic cells within the TME support melanoma growth, survival and metastasis. Contrastingly, such cells can be stimulated to promote tumour killing when engaged with monoclonal antibodies (mAbs) against cancer antigens, owing to their expression of multiple Fc receptors (FcRs). However, little is known about the monocytic subsets present within melanoma, the FcRs they express and how engaging their FcRs may influence function. Methods Flow cytometry on peripheral monocytes isolated from patients with melanoma and healthy volunteers enabled the characterization of cell surface markers. Next, publicly available single-cell RNA-Seq datasets were analysed to explore the transcriptome of melanoma-associated macrophages and these phenotypes were recapitulated in vitro. Finally, monocytes and macrophages were cocultured with melanoma cells, with or without the addition of mAbs against a melanoma antigen, in order to assess the effect of FcR crosslinking on monocyte and macrophage phenotype and function. Results Circulating monocytes in patients with melanoma feature an inhibitory phenotype, and express multiple FcRs, including Fcε and Fcγ receptors, which could be targeted with mAbs of multiple isotypes. Secondly, monocytic cells within the melanoma TME express protumour genes, including metalloproteinases and angiogenic factors. However, FcR expression suggests that tumour-promoting subsets can be targeted with mAbs. Finally, in ex vivo cocultures with melanoma cells, the FcRs of both human monocytes and macrophages are able to engage antimelanoma mAbs of multiple isotypes, inducing differential phenotype change. This is demonstrated by the upregulation of activation markers and costimulatory cell surface markers. Conclusions Together, these data suggest that tumour-promoting monocytic cells within the melanoma TME can be engaged and harnessed using different isotypes of mAbs targeting melanoma.
P15 Understanding the role of cathepsin H in atopic dermatitis pathogenesis
Authors: Theresa Tachie-Menson, Aaron Hughes, Edel O’Toole, Ryan O’Shaughnessy
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims Atopic dermatitis (AD) is a chronic skin condition ranked the 15th highest global burden for all nonfatal diseases. Our lab has found that patients with AD have a reduction in the gene and protein expression of cathepsin H, a lysosomal cysteine protease with aminopeptidase activity. Ctsh-/- mice have a reduced integrity of the epidermal barrier, including reduced filaggrin processing, reduced corneocyte integrity and a change in the expression of several proinflammatory mediators including interleukin-1α. We aim to elucidate the roles of cathepsin H in keratinocytes to understand its contribution to the pathogenesis of AD. Methods We utilized rat epidermal and N/TERT keratinocytes transiently overexpressing procathepsin B, H or L in addition to incubating keratinocytes with cathepsin H inhibitor (CTSHi) for 24 h to identify potential substrates and proteins upregulated or downregulated with cathepsin H expression. We also performed colocalization studies using indirect immunofluorescence in rat epidermal keratinocytes (REKs) and N/TERTs to investigate the biology of cathepsin H. Results REKs transiently overexpressing procathepsin H-FLAG or FLAG and REKs treated with the cathepsin H inhibitor H2N-Ser(OBzl)-CHN2 were analysed by liquid chromatography-mass spectrometry. This analysis indicated that keratin 1 and keratin 9 were upregulated with cathepsin H expression and downregulated by cathepsin H inhibition. Overexpressed procathepsin H and endogenous cathepsin H do not colocalize with the lysosomal membrane protein, LAMP1, whereas endogenous cathepsin B does. Conclusions Keratin 1 and keratin 9 could be coregulated with cathepsin H. Cathepsin H may not primarily localize to lysosomes unlike cathepsin B.
O05 Single-cell transcriptomic signature of paradoxical eczema in patients with psoriasis treated with biologics
Authors: Ali Al-Janabi, Paul Martin, Catherine Simpson, Hefin Rhys, Adnan Khan, Steve Eyre, Maria Christofi, Amy Foulkes, Andrew Skelton, Sebastien Viatte, Anne Barton, Andrew Morris, Catherine Smith, Christopher Griffiths, Richard Warren
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims Biologics targeting the tumour necrosis factor (TNF) and the interleukin (IL)-17/23 axis are highly effective treatments for psoriasis but can result in cutaneous adverse events. The pathogenesis of paradoxical eczema, an atopic dermatitis (AD) phenotype occurring after biologic initiation in people with psoriasis, is unknown. This study explored the systemic disease signature of paradoxical eczema. Methods We used single-cell RNA-Seq on peripheral blood mononuclear cells (PBMCs) from three patients with psoriasis with paradoxical eczema, and three matched, biologic-treated controls who had psoriasis. Differential gene expression, gene network expression and cell–cell interaction (using MultiNicheNet) analyses allowed comparison of cases and controls. We assessed overlap between paradoxical eczema and AD using six AD transcriptome datasets. Results Overall, 32 827 PBMCs (14 521 from cases, 18 306 from controls) were included and categorized into 27 clusters. Between cases and controls, there were 727 differentially expressed genes (DEGs) (log fold-change &amp;gt; 0.25, adjusted P &amp;lt; 0.05). A total of 94 of the top 100 DEGs were from monocytes. Significant DEG networks (P &amp;lt; 0.05) mostly mapped to T-cell and monocyte clusters, with commonly enriched pathways in cases including interferon (IFN)-γ, IFN-α, TNF, IL-2/signal transducer and activator of transcription 5 and T-cell differentiation pathways. Cell–cell interaction analysis revealed that the most prioritized interactions in controls, based on differential expression data and known ligand-receptor interactions, included immune regulatory interactions (including ligands such as transforming growth factor β1 and lymphocyte-activation gene 3). In cases, functions of prioritized interactions included chemotaxis, leucocyte activation, T-cell priming and IFN-γ signalling. There was significant enrichment of concordance in the direction of effect of DEGs (65%, P = 5.39 × 10−6) and enriched pathways (85%, P = 3.62 × 10−13) between AD and paradoxical eczema. Conclusions Patients with paradoxical eczema have reduced immune regulatory mechanisms and evidence of monocyte and T-cell activation systemically, with increased production IFN-γ, IFN-α and TNF. There is a significant overlap with the AD transcriptome.
P09 Next-generation sequencing: on-and off-target analysis of gene editing in fibroblasts from patients with recessive dystrophic epidermolysis bullosa
Authors: Aidin Kazemizadeh, Imogen Brooks, Ina Guri, Yara Alrokh, Katarzyna Balon, Lukasz Laczmanski, John McGrath, Joanna Jackow-Malinowska
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims Base editors (BEs) and prime editors (PEs) have proved to have advantages over traditional clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 in correcting pathogenic mutations. We previously demonstrated 94.6% efficiency using adenine base editor (ABE)8e in correcting nonsense mutation (c.5047 C&amp;gt;T) in fibroblasts, which causes recessive dystrophic epidermolysis bullosa (RDEB). However, safety profile and cell-type-specific efficiencies of gene editing techniques have not been characterized. The aim of our research is to establish corrections of c.5047 C&amp;gt;T pathogenic variant in fibroblasts and compare off-target effects and efficiency of CRISPR-Cas9, BE and PE systems. Methods Primary fibroblasts harbouring heterozygous C&amp;gt;T pathogenic variant on exon 54 will be transfected with CRISPR-Cas9, ABE8e and PE using electroporation or lipofectamine. Both Sanger sequencing and next-generation sequencing (NGS) will be conducted to confirm pathogenic variant corrections. Galaxy platform which incorporates different NGS analysis tools will be used to analyse the NGS data. Subsequently, whole-exome sequencing and targeted amplicon sequencing will be used for off-target analysis. Results On-target efficiency for ABE8e to correct c.5047 C&amp;gt;T pathogenic variant in patient primary fibroblasts was determined first by Sanger sequencing showing 100% editing in contrast to 84% determined by NGS. Moving forward, we plan to correct the same pathogenic variant with PEs. Initially experiments are under way with PEmax mRNA and engineered PE guide RNAs (epegRNA) to compare different PEs. Furthermore, we will investigate the off-target effects of CRISPR-Cas9, ABE8e and PEs to edit RDEB primary fibroblasts using NGS Methods and conduct functional studies. Conclusions Here, we showed that the c.5047 C&amp;gt;T pathogenic variant in RDEB fibroblasts could be successfully edited using ABE8e. We demonstrated that NGS is required to precisely determine the editing efficacy. Taken together, we have established a pipeline for base editing in RDEB and we aim to achieve the same for prime editing in the future.
P16 Fibroblast signalling crosstalk drives junctional epidermolysis bullosa disease progression
Authors: Mohammad R A Khan, Thomas Kirk, Matthew P Caley, Emanuel Rognoni
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims Junctional epidermolysis bullosa (JEB) is a devastating hereditary skin disorder characterized by severe fragility of skin and mucus membrane, emerges as blistering with little or no trauma. The major classification of JEB includes JEB intermediate and JEB severe; the latter is the most common subtype, which is the result of loss of function mutations in the skin basement membrane protein laminin-332. Lacking this protein impairs basement membrane adhesion of epidermal keratinocytes and leads to deregulated epidermal–dermal crosstalk. The skin dermis is composed of multiple fibroblast subpopulations with different functions in skin homeostasis and repair; however, their specific involvement in skin diseases is largely unknown. Here, we investigate how fibroblast subpopulations, namely papillary and reticular fibroblasts, impact JEB disease progression and skin regeneration. Methods Three-dimensional (3D) organotypic (OT) combining healthy (control) and diseased (JEB) keratinocytes cells with distinct fibroblast subpopulations from different anatomical sites were developed and analysed by immunohistochemistry and RNA-Seq. Results JEB OTs with papillary fibroblasts showed increased epidermal growth, invagination of basal layer and deregulated differentiation compared with reticular fibroblasts. RNA-Seq analysis revealed altered activity in key signalling including immune, cytokine, gap junction, mitogen-activated protein kinase and remodelling of keratinization and ECM proteins, particularly in JEB OT cultured with papillary fibroblast. JEB OTs and cells also demonstrated variation in cell proliferation and activation of transforming growth factor-β1 and immune cell signalling. Currently, we are targeting identified signalling pathways in our two-dimensional and 3D cell culture models and identifying druggable targets, before testing selected candidates in our JEB mouse disease model for further preclinical validation. Conclusions By dissecting the pathological signalling interactions between keratinocytes and distinct fibroblast subpopulations, our goal is to identify new therapeutic target to improve the skin health and regeneration in patients with JEB.
P13 Keratinocyte heterogeneity and dynamics in human skin wound healing
Authors: Jia Jun Lee, Claire Higgins
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims Human chronic skin wounds have been extensively characterized, but ethical limitations of collecting and studying wounds from healthy individuals means there is a sparsity of data regarding re-epithelialization in normal wound healing. Here, we characterize ex vivo models of wound closure in human skin to better understand the keratinocyte dynamics of normal healing. Methods We conducted single-cell RNA sequencing (scRNA-Seq) from ex vivo skin explants 2 days postwounding (n = 4) to interrogate heterogeneity in keratinocytes relative to the wound edge. Immunostaining was also conducted on sections from skin explants at different timepoints during wound closure. Scratch assays were conducted using isolated human keratinocytes. Specific zones of keratinocytes were separated relative to the original scratch, 6 h postscratch, and used for real-time polymerase chain reaction analyses. Results We found that partial re-epithelialization and full healing of 2-mm wounds occurred 2–3 days and 4 days postwounding, respectively. Analyses of the scRNA-Seq data showed an increase in S100A7 expression in suprabasal keratinocyte subclusters in wounded skin compared with unwounded skin. Using immunostaining we also showed that S100A7 is expressed at the leading edge of the regenerating epidermis. In an in vitro scratch wound model, the keratinocytes nearest to the wound edge also show an upregulation of S100A7, EGFR, EPHA4 and TIMP1, as well as a downregulation in PCNA, implying a decrease in cell proliferation in leading keratinocytes. Conclusions S100A7 was previously reported to be increased at the wound edge in human keratinocytes, highlighting that transcriptional changes in our ex vivo and in vitro human wound healing models reflect those occurring in vivo. The decrease in PCNA in keratinocytes in the leading edge also recapitulates Results previously shown in mouse models of wound healing. These data on normal re-epithelialization will be compared with published data on human chronic wounds and other skin diseases.
P14 Characterizing the hair follicle–nervous system connection
Authors: Anna Rhodes, Julià Agramunt, Summik Limbu, Rimma Laufer, Francisco Jiménez, Greg Williams, Leigh Knight, Parastoo Hashemi, Claire Higgins
Journal: British Journal of Dermatology (2024)

Abstract Introduction and aims The human hair follicle (HF) is a miniorgan in the skin that possesses a rich microenvironment composed of a complex assembly of sensory nerves that surround it. Recent studies have explored the effects of this perineural niche on the HF, but the influence of HF on the nervous system from a folliculocentric and skin-centric perspective remains underexplored. To explore the potential bidirectionality of the HF–nervous system connection in the context of nerve development, we asked whether the HF influences the differentiation and maturation of sensory neuron subtypes around it through the release of distinct neurotransmitters and neurotrophic factors. Methods To first identify which cell types might be involved in directing sensory neuron development, we reanalysed available skin and HF single-cell RNA sequencing (scRNA-Seq) datasets. Next, to evaluate neurotransmitter release from these cell types, we used fast-scan cyclic voltammetry (FSCV). Lastly, to interrogate the role of the HF on directing neuronal development in vitro, we used induced pluripotent stem cell-derived sensory neurons as a model for developing nerves. Results We found that a higher percentage of KRT14- (epithelial) and PDGFRA-expressing (mesenchymal) cells in the HF coexpressed genes encoding specific neurotrophic factors and enzymatic machinery for neurotransmitter production compared with equivalent cells in the skin, suggesting that the HF possesses a greater capacity to signal to the nervous system compared with interfollicular skin. Using FSCV, we then found that follicular outer root sheath and directionally selective cells are capable of releasing neurotransmitters such as serotonin and histamine. Conclusions We postulate that HF-derived neurotransmitters are important for aspects of sensory neuron development such as axonal growth, as serotonin has previously been shown to be an attractive guidance cue for neuronal growth cones. Overall, our work sheds light on the HF–nervous system connection and how the HF may regulate its microenvironment in the skin.
An operational guide to translational clinical machine learning in academic medical centers
Authors: Mukund Poddar, Jayson S. Marwaha, William Yuan, Santiago Romero-Brufau, Gabriel A. Brat
Journal: npj Digital Medicine (2024)

Abstract Few published data science tools are ever translated from academia to real-world clinical settings for which they were intended. One dimension of this problem is the software engineering task of turning published academic projects into tools that are usable at the bedside. Given the complexity of the data ecosystem in large health systems, this task often represents a significant barrier to the real-world deployment of data science tools for prospective piloting and evaluation. Many information technology companies have created Machine Learning Operations (MLOps) teams to help with such tasks at scale, but the low penetration of home-grown data science tools in regular clinical practice precludes the formation of such teams in healthcare organizations. Based on experiences deploying data science tools at two large academic medical centers (Beth Israel Deaconess Medical Center, Boston, MA; Mayo Clinic, Rochester, MN), we propose a strategy to facilitate this transition from academic product to operational tool, defining the responsibilities of the principal investigator, data scientist, machine learning engineer, health system IT administrator, and clinician end-user throughout the process. We first enumerate the technical resources and stakeholders needed to prepare for model deployment. We then propose an approach to planning how the final product will work from data extraction and analysis to visualization of model outputs. Finally, we describe how the team should execute on this plan. We hope to guide health systems aiming to deploy minimum viable data science tools and realize their value in clinical practice.