Search

Breast cancer-related lymphedema (BCRL) is characterized by skin changes, swelling, fibrosis, and recurrent skin infections. Clinical studies have suggested that lymphedema results in skin barrier defects; however, the underlying cellular mechanisms and the effects of bacterial contamination on skin barrier function remain unknown. In matched biopsies from patients with unilateral BCRL, we observed decreased expression of FLG and the tight junction protein ZO-1 in skin affected by moderate lymphedema or by subclinical lymphedema in which dermal backflow of lymph was identified by indocyanine green lymphography, relative to those in the controls (areas without backflow and from the unaffected arm). In vitro stimulation of keratinocytes with lymph fluid obtained from patients undergoing lymphedema surgery led to the same changes as well as increased expression of keratin 14, a marker of immature keratinocytes. Finally, using mouse models of lymphedema, we showed that similar to the clinical scenario, the expression of skin barrier proteins was decreased relative to that in normal skin and that colonization with Staphylococcus epidermidis bacteria amplified this effect as well as lymphedema severity. Taken together, our findings suggest that lymphatic fluid stasis contributes to skin barrier dysfunction in lymphedema., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The skin epidermis provides a barrier that is imperative for preventing transepidermal water loss (TEWL) and protecting against environmental stimuli. The underlying molecular mechanisms for regulating barrier functions and sustaining its integrity remain unclear. RORα is a nuclear receptor highly expressed in the epidermis of normal skin. Clinical studies showed that the epidermal RORα expression is significantly reduced in the lesions of multiple inflammatory skin diseases. In this study, we investigate the central roles of RORα in stabilizing skin barrier function using mice with an epidermis-specific Rora gene deletion ( Rora EKO ). While lacking spontaneous skin lesions or dermatitis, Rora EKO mice exhibited an elevated TEWL rate and skin characteristics of barrier dysfunction. Immunostaining and Western blot analysis revealed low levels of cornified envelope proteins in the Rora EKO epidermis, suggesting disturbed late epidermal differentiation. In addition, an RNA-seq analysis showed the altered expression of genes related to "keratinization" and "lipid metabolism" in RORα deficient epidermis. A lipidomic analysis further uncovered an aberrant ceramide composition in the Rora EKO epidermis. Importantly, epidermal Rora ablation greatly exaggerated percutaneous allergic inflammatory responses to oxazolone in an allergic contact dermatitis (ACD) mouse model. Our results substantiate the essence of epidermal RORα in maintaining late keratinocyte differentiation and normal barrier function while suppressing cutaneous inflammation.

In vitro epidermis models are important to evaluate and study disease progression and possible dermal drug delivery. An in vitro epidermis model using floating paper chips as a scaffold for proliferation and differentiation of primary human keratinocytes is reported. The formation of the four main layers of the epidermis (i.e., basal, spinosum, granulose, and cornified layers) is confirmed. The development of a cornified layer and the tight junction formation are evaluated as well as the alterations of organelles during the differentiation process. Further, this in vitro model is used to assess keratinocyte migration. Finally, magnetic micromotors are assembled, and their ability to aid cell migration on paper chips is confirmed when a static magnetic field is present. Taken together, this attempt to combine bottom-up synthetic biology with dermatology offers interesting opportunities for studying skin disease pathologies and evaluate possible treatments., (© 2022 The Authors. Small published by Wiley-VCH GmbH.)

Twenty years after the cloning, characterization, and identification of interleukin (IL)-22 in 2000, the precise biological role of this cytokine in healthy and unhealthy skin is not completely known. The aim of this review is to provide an overview on the recent knowledge available in literature about the origin, sources, targets, molecular mechanism of action, and clinical issues regarding IL-22. Last but not least, recent experimental evidence obtained in a 3D model of organotypic culture of normal human skin highlights its homeostatic role and will be discussed in detail, as personal observations. As most of the data concerning IL-22 immunomodulating activity are obtained from mouse models, this work offers a new perspective on its clinical role. The hypothesis herein advanced is that IL-22 profoundly affects keratinocyte terminal differentiation, whereas, in order to induce a proliferation impairment, a more complex psoriatic-like microenvironment is needed., (© 2022 S. Karger AG, Basel.)

Aberrant differentiation of keratinocytes disrupts the skin barrier and causes a series of skin diseases. However, the molecular basis of keratinocyte differentiation is still poorly understood. In the present study, we examined the expression of C7ORF41 using tissue microarrays by immunohistochemistry and found that C7ORF41 is specifically expressed in the basal layers of skin epithelium and its expression is gradually decreased during keratinocytes differentiation. Importantly, we corroborated the pivotal role of C7ORF41 during keratinocyte differentiation by C7ORF41 knockdown or overexpression in TPA-induced Hacat keratinocytes. Mechanismly, we first demonstrated that C7ORF41 inhibited keratinocyte differentiation mainly through formatting a complex with IKKα in the cytoplasm, which thus blocked the nuclear translocation of IKKα. Furthermore, we also demonstrated that inhibiting the PKCα/ERK signaling pathway reversed the reduction in C7ORF41 in TPA-induced keratinocytes, indicating that C7ORF41 expression could be regulated by upstream PKCα/ERK signaling pathway during keratinocyte differentiation. Collectively, our study uncovers a novel regulatory network PKCα/ERK/C7ORF41/IKKα during keratinocyte differentiation, which provides potential therapeutic targets for skin diseases., (© 2021 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.)

Background: Cutaneous adverse drug reactions (CADR) associated with oncology therapy involve 45-100% of patients receiving kinase inhibitors. Such adverse reactions may include skin inflammation, infection, pruritus and dryness, symptoms that can significantly affect the patient's quality of life. To prevent severe skin damages dose adjustment or drug discontinuation is often required, interfering with the prescribed oncology treatment protocol. This is particularly the case of Epidermal Growth Factor Receptor inhibitors (EGFRi) targeting carcinomas. Since the EGFR pathway is pivotal for epidermal keratinocytes, it is reasonable to hypothesize that EGFRi also affect these cells and therefore interfere with the epidermal structure formation and skin barrier function., Methods: To test this hypothesis, the effects of EGFRi and Vascular Endothelial Growth Factor Receptor inhibitors (VEGFRi) at therapeutically relevant concentrations (3, 10, 30, 100 nM) were assessed on proliferation and differentiation markers of human keratinocytes in a novel 3D micro-epidermis tissue culture model., Results: EGFRi directly affect basal keratinocyte growth, leading to tissue size reduction and switching keratinocytes from a proliferative to a differentiative phenotype, as evidenced by decreased Ki67 staining and increased filaggrin, desmoglein-1 and involucrin expression compared to control. These effects lead to skin barrier impairment, which can be observed in a reconstructed human epidermis model showing a decrease in trans-epidermal water loss rates. On the other hand, pan-kinase inhibitors mainly targeting VEGFR barely affect keratinocyte differentiation and rather promote a proliferative phenotype., Conclusions: This study contributes to the mechanistic understanding of the clinically observed CADR during therapy with EGFRi. These in vitro results suggest a specific mode of action of EGFRi by directly affecting keratinocyte growth and barrier function.

The role of microRNAs (miRNAs) during keratinocyte (KC) differentiation and in skin diseases with epidermal phenotypes has attracted strong interest over the past few years. However, combined mRNA and miRNA expression analyses to elucidate the intricate mRNA-miRNA networks of KCs at different stages of differentiation have not been performed yet. In the present study, we investigated the dynamics of miRNA and mRNA expression during KC differentiation in vitro and in normal and psoriatic epidermis. While we identified comparable numbers of up- and downregulated mRNAs (49% and 51%, respectively), miRNAs were predominantly upregulated (76% vs 24%) during KC differentiation. Further bioinformatics analyses suggested an important inhibitory role for miR-155 in KC differentiation, as it was repressed during KC differentiation in normal skin but strongly upregulated in the epidermis of psoriatic skin lesions. Mimicking the inflammatory milieu of psoriatic skin in vitro, we could show that the pro-inflammatory cytokines IL17, IL1β and INFγ synergistically upregulated miR-155 expression in KCs. Forced over-expression of miR-155 in human in vitro skin models specifically reduced the expression of loricrin (LOR) in KCs, indicating that miR-155 interferes with the establishment of a normal epidermal barrier. Together, our data indicate that downregulation of miR-155 during KC differentiation is a crucial step for epidermal barrier formation. Furthermore, its strong upregulation in psoriatic lesions suggests a contributing role of miR-155 in the altered keratinocyte differentiation observed in psoriasis. Therefore, miR-155 represents as a potential target for treating psoriatic skin lesions.

Heat shock proteins (HSPs), a large group of highly evolutionary conserved proteins, are considered to be main elements of the cellular proteoprotection system. HSPs are encoded by genes activated during the exposure of cells to proteotoxic factors, as well as by genes that are expressed constitutively under physiological conditions. HSPs, having properties of molecular chaperones, are involved in controlling/modulation of multiple cellular and physiological processes. In the presented review, we summarize the current knowledge on HSPs in the biology of epidermis, the outer skin layer composed of stratified squamous epithelium. This tissue has a vital barrier function preventing from dehydratation due to passive diffusion of water out of the skin, and protecting from infection and other environmental insults. We focused on HSPB1 (HSP27), HSPA1 (HSP70), HSPA2, and HSPC (HSP90), because only these HSPs have been studied in the context of physiology and pathophysiology of the epidermis. The analysis of literature data shows that HSPB1 plays a role in the regulation of final steps of keratinization; HSPA1 is involved in the cytoprotection, HSPA2 contributes to the early steps of keratinocyte differentiation, while HSPC is essential in the re-epithelialization process. Since HSPs have diverse functions in various types of somatic tissues, in spite of multiple investigations, open questions still remain about detailed roles of a particular HSP isoform in the biology of epidermal keratinocytes.

Epidermis, the outermost layer of the skin, plays a critical role as both a physical and immunological barrier protecting the internal tissues from external environmental insults, such as pathogenic bacteria, fungi, viruses, UV irradiation, and water loss. Epidermal keratinocytes (KC), the predominant cell type in the skin epidermis, are in the front line of skin defense. Here we describe methods to isolate and culture primary epidermal KC from neonatal and adult mouse skin and describe in vitro assays to study and characterize KC proliferation and differentiation and pro-inflammatory responses to viral products and UVB irradiation. These methods will be useful for researchers in the field of epidermal biology to set up in vitro assays to study the barrier and pro-inflammatory function of epidermal keratinocytes.

Background: Ultraviolet (UV) radiation is known to be one of the most important environmental hazards acting on the skin. The most part of UV radiation is absorbed in the epidermis, where keratinocytes are the most abundant and exposed cell type. Lipids have an important role in skin biology, not only for their important contribution to the maintenance of the permeability barrier but also for the production and storage of energy, membrane organization and cell signalling functions. However, the effects on the lipid composition of keratinocytes under UV radiation are little explored., Objective: The present work aims to explore the effects on the phenotype and lipid content of primary human keratinocytes exposed to simulated solar UV radiation., Methods: Keratinocytes were exposed to a single (acute exposure) and repeated simulated solar UV irradiations for 4 weeks (chronic exposure). Cell viability and morphology were explored, as well as the production of reactive oxygen species. Then, lipid extracts were analysed through liquid chromatography coupled to mass spectrometry (LC-MS) and the data generated was processed using the ROIMCR chemometric methodology together with partial least squares discriminant analysis (PLS-DA), to finally reveal the most relevant lipid changes that occurred in keratinocytes upon UV irradiation. Also, the potential induction of keratinocyte differentiation was explored by measuring the increase of involucrin., Results: Under acute irradiation, cell viability and morphology were not altered. However, a general increase of phosphatidylcholines (PC) phosphatidylethanolamines (PE) and phosphatidylglycerol (PG) together with a slight sphingomyelin (SM) decrease were found in UV irradiated cells, among other changes. In addition, keratinocyte cultures did not present any differentiation hallmark. Contrary to acute-irradiated cells, in chronic exposures, cell viability was reduced and keratinocytes presented an altered morphology. Also, hallmarks of differentiation, such as the increase of involucrin protein and the autophagy induction were detected. Among the main lipid changes that accompanied this phenotype, the increase of long-chain ceramides, lysoPC and glycerolipid species were found., Conclusion: Important lipid changes were detected under acute and chronic UV irradiation. The lipid profile under chronic exposure may represent a lipid fingerprint of the keratinocyte differentiation phenotype., (Copyright © 2018 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)

Deimination or citrullination is a post-translational modification catalysed by a family of calcium-dependent enzymes called peptidylarginine deiminases (PADs). It corresponds to the transformation of arginine residues within a peptide sequence into citrulline residues. Deimination induces a decreased net charge of targeted proteins; therefore, it alters their folding and changes intra- and intermolecular ionic interactions. Deimination is involved in several physiological processes (inflammation, gene regulation, etc.) and human diseases (rheumatoid arthritis, neurodegenerative diseases, cancer, etc.). Here, we describe the PADs expressed in the epidermis and their known substrates, focusing on their role in the epidermal barrier function., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: A variety of human skin disorders is characterized by defects in the epidermal barrier, leading to dehydration, itchiness, and rashes. Previously published literature suggests that microtubule stabilization at the cortex of differentiating keratinocytes is necessary for the formation of the epidermal barrier., Objectives: We tested whether stabilization of microtubules with paclitaxel or epothilone B can repair barrier defects that were experimentally induced in three-dimensional culture models of epidermis., Methods: We established two models of defective epidermis in vitro, using three-dimensional cultures of primary human keratinocytes on filter supports: immature reconstructed human epidermis (RHE), and RHE that was compromised by treatment with inflammatory cytokines, the latter mimicking defects seen in atopic dermatitis., Results: Both paclitaxel and epothilone B promoted keratinocyte differentiation, accumulation of junctional proteins at the cell cortex, and the early appearance of lamellar bodies in immature RHE, whereas destabilization of microtubules by nocodazole had the reverse effect. Moreover, stabilization of microtubules rescued the barrier after cytokine treatment. The rescued barrier function correlated with the restoration of filaggrin and loricrin protein levels, the cortical accumulation of junctional proteins (E-cadherin, β-catenin, and claudin-1), and with the secretion of lamellar bodies., Conclusions: Our data suggest that the microtubule network is important for the formation of the epidermis, and that stabilization of microtubules promotes barrier formation. Microtubule stabilization may support regeneration of damaged skin, by restoring or improving the barrier., (Copyright © 2018 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)

Mammalian epidermis is composed of four morphologically and functionally distinct layers of keratinocytes. The innermost basal layer consists of proliferating self-renewing keratinocytes, which also undergo asymmetric cell division to differentiate into postmitotic suprabasal cells throughout life. Control of the balance between growth and differentiation of basal cells is important for epidermal homeostasis to prevent skin disorders including malignancies; however, the underlying mechanism remains to be elucidated. Recently, MafB was identified as one of the transcription factors that regulate epidermal keratinocyte differentiation. MafB is expressed in postmitotic differentiating keratinocytes, and epidermal differentiation is partially impaired in MafB-deficient mice. To further establish the roles of MafB in the epidermis in vivo, we generated mice transgenic for MafB under the control of the basal cell-specific keratin (Krt) 14 promoter. In the epidermis of transgenic mice at embryonic day 18.5, the number of proliferating Krt14-positive basal-like cells was increased, and the granular and cornified layers were thickened. Furthermore, these MafB transgenic mice developed papillomas spontaneously with age. Therefore, MafB promotes differentiation in postmitotic keratinocytes and simultaneously has potential to promote growth when ectopically expressed in undifferentiated basal keratinocytes., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Transglutaminases (TGs) are a family of enzymes that catalyse the formation of isopeptide bonds between the γ-carboxamide groups of glutamine residues and the ε-amino groups of lysine residues leading to cross-linking reactions among proteins. Four members, TG1, TG2, TG3, and TG5, of the nine mammalian enzymes are expressed in the skin. TG1, TG3 and TG5 crosslinking properties are fundamental for cornified envelope assembly. In contrast, the role of TG2 in keratinization has never been studied at biochemical level in vivo. In this study, taking advantage of the TG2 knock-out (KO) and TG1 heterozygous mice, we generated and characterized the epidermis of TG1-TG2 double knock-out (DKO) mice. We performed morphological analysis of the epidermis and evaluation of the expression of differentiation markers. In addition, we performed analysis of the amino acid composition from isolated corneocytes. We found a significant change in amino acid composition in TG1KO cornified cell envelopes (CEs) while TG2KO amino acid composition was similar to wild-type CEs. Our results confirm a key role of TG1 in skin differentiation and CE assembly and demonstrate that TG2 is not essential for CE assembly and skin formation.

Omega-hydroxyceramides (ω-OH-Cer) play a crucial role in maintaining the integrity of skin barrier. ω-OH-Cer are the primary lipid constituents of the corneocyte lipid envelope (CLE) covalently attached to the outer surface of the cornified envelope linked to involucrin to become bound form lipids in stratum corneum (SC). CLE becomes a hydrophobic impermeable layer of matured corneocyte preventing loss of natural moisturizing factor inside the corneocytes. More importantly, CLE may also play an important role in the formation of proper orientation of intercellular lipid lamellar structure by interdigitating with the intercellular lipids in a comb-like fashion. Abnormal barrier conditions associated with atopic dermatitis but also UVB-irradiated skins are known to have lowered level of bound lipids, especially ω-OH-Cer, which indicate that ω-OH-Cer play an important role in maintaining the integrity of skin barrier. In this study, protective effects of a novel synthetic C16 omega-hydroxyphytoceramides (ω-OH-phytoceramide) on skin barrier function were investigated. Epidermal barrier disruption was induced by UVB irradiation, tape-stripping in hairless mouse and human skin. Protective effect of damaged epidermis was evaluated using the measurement of transepidermal water loss and cohesion of SC. Increased keratinocyte differentiation was verified using cultured keratinocyte through western blot. Results clearly demonstrated that a synthetic C16 ω-OH-phytoceramide enhanced the integrity of SC and accelerated the recovery of damaged skin barrier function by stimulating differentiation process. In a conclusion, a synthetic C16 ω-OH-phytoceramide treatment improved epidermal homeostasis in several disrupted conditions.

The well-established human epidermal keratinocyte (HEK) differentiation model was investigated to determine possible alterations in chromosome territory (CT) association during differentiation. The seven human chromosomes (1, 4, 11, 12, 16, 17, and 18) selected for this analysis are representative of the chromosome size and gene density range of the overall human genome as well as including a majority of genes involved in epidermal development and differentiation (CT1, 12, and 17). Induction with calcium chloride (Ca(2+)) resulted in morphological changes characteristic of keratinocyte differentiation. Combined multi-fluorescence in situ hybridization (FISH) and computational image analysis on the undifferentiated (0 h) and differentiated (24 h after Ca(2+) treatment) HEK revealed that (a) increases in CT volumes correspond to overall nuclear volume increases, (b) radial positioning is gene density-dependent at 0 h but neither gene density- nor size-dependent at 24 h, (c) the average number of interchromosomal associations for each CT is gene density-dependent and similar at both time points, and (d) there are striking differences in the single and multiple pairwise interchromosomal association profiles. Probabilistic network models of the overall interchromosomal associations demonstrate major reorganization of the network during differentiation. Only ~40 % of the CT pairwise connections in the networks are common to both 0 and 24 h HEK. We propose that there is a probabilistic chromosome positional code which can be significantly altered during cell differentiation in coordination with reprogramming of gene expression.

The human epidermis provides a very effective barrier function against chemical, physical and microbial insults from the environment. This is only possible as the epidermis renews itself constantly. Stem cells located at the basal lamina which forms the dermoepidermal junction provide an almost inexhaustible source of keratinocytes which differentiate and die during their journey to the surface where they are shed off as scales. Despite the continuous renewal of the epidermis it nevertheless succumbs to aging as the turnover rate of the keratinocytes is slowing down dramatically. Aging is associated with such hallmarks as thinning of the epidermis, elastosis, loss of melanocytes associated with an increased paleness and lucency of the skin and a decreased barrier function. As the differentiation of keratinocytes is strictly calcium dependent, calcium also plays an important role in the aging epidermis. Just recently it was shown that the epidermal calcium gradient in the skin that facilitates the proliferation of keratinocytes in the stratum basale and enables differentiation in the stratum granulosum is lost in the process of skin aging. In the course of this review we try to explain how this calcium gradient is built up on the one hand and is lost during aging on the other hand. How this disturbed calcium homeostasis is affecting the gene expression in aged skin and is leading to dramatic changes in the composition of the cornified envelope will also be discussed. This loss of the epidermal calcium gradient is not only specific for skin aging but can also be found in skin diseases such as Darier disease, Hailey-Hailey disease, psoriasis and atopic dermatitis, which might be very helpful to get a deeper insight in skin aging., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Expression of the human dermokine gene (DMKN) leads to the production of four dermokine isoform families. The secreted α, β and γ isoforms have an epidermis-restricted expression pattern, with Dmkn β and γ being specifically expressed by the granular keratinocytes. The δ isoforms are intracellular and ubiquitous. Here, we performed an in-depth characterization of Dmkn expression in mouse skin and found an expression pattern that was less complex than in humans. In particular, mRNA coding for the δ family were absent. Homozygous mice null for the Dmkn β and γ isoforms had no obvious phenotype but only a temporary scaly skin during the first week of life. The pups null for the Dmkn β and γ isoforms had smaller keratohyalin granules and their cornified envelopes were more sensitive to mechanical stress. At the molecular level, amounts of profilaggrin and filaggrin monomers were reduced whereas amino acid components of the natural moisturizing factor were increased. In addition, the electrophoretic mobility of involucrin was modified, suggesting post-translational modifications. Finally, the mice null for the Dmkn β and γ isoforms strongly overexpressed Dmkn α. These data are evocative of compensatory mechanisms relevant to the temporary phenotype. Overall, we improved the knowledge of Dmkn expression in mouse and highlighted a role for Dmkn β and γ in cornification., (© 2014. Published by The Company of Biologists Ltd.)

Objective: Hyaluronan (HA) plays a role in keratinocyte proliferation and differentiation. In addition, HA has been shown to have different biological activities depending on its molecular weight. It has been reported that HA-mediated CD44 activation regulates keratinocyte differentiation. Therefore, the aim of this study was to investigate the influence of HA tetrasaccharides (HA4) on the regulation of keratinocyte differentiation, CD44 gene expression and CD44-phosphorylated protein in human keratinocytes, and compare HA4 with high molecular weight HA., Methods: Normal human epidermal keratinocytes (NHEKs) were treated at doses of 1 μg mL(-1) HA or HA oligosaccharides (HA4). After treatment, cell viability was checked using an MTT (3-(4,5-di-methylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Each differentiation marker and CD44 mRNA expression was detected by real-time PCR. Each differentiation marker and CD44-phosphorylated protein was assessed by Western blotting., Results: Hyaluronan and HA4 showed no cytotoxicity up to a dose of 1 μg mL(-1) . On day 3 after HA4 treatment, each differentiation marker mRNA and K10 protein level was higher than that of the control. On day 9, late differentiation marker mRNA and protein levels were increased with HA and HA4 treatment. In addition, HA4 treatment increased the expression of CD44 mRNA, CD44-phosphorylated protein and intracellular calcium concentrations. HA4 enhanced keratinocyte differentiation and increased CD44-phosphorylated protein levels., Conclusion: HA4 may induce epidermal differentiation through phosphorylation of CD44., (© 2013 Society of Cosmetic Scientists and the Société Française de Cosmétologie.)

The epidermis is a stratified squamous epithelium composed of proliferating basal and differentiated suprabasal keratinocytes. It serves as the body's major physical and chemical barrier against infection and harsh environmental insults, as well as preventing excess water loss from the body into the atmosphere. Calcium is a key regulator of the proliferation and differentiation in keratinocytes. Elevated extracellular Ca(2+) concentration ([Ca(2+)]o) raises the levels of intracellular free calcium ([Ca(2+)]i), promotes cell-cell adhesion, and activates differentiation-related genes. Keratinocytes deficient in the calcium-sensing receptor fail to respond to [Ca(2+)]o stimulation and to differentiate, indicating a role for the calcium-sensing receptor in transducing the [Ca(2+)]o signal during differentiation. The concepts derived from in vitro gene knockdown experiments have been evaluated and confirmed in three mouse models in vivo., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Objective: Human skin is the first line of defence from environmental factors such as solar radiation and is susceptible to premature ageing, including a disruption in epidermal differentiation and homeostasis. We evaluated the impact of a Galactomyces Ferment Filtrate (GFF) on epidermal differentiation and response to oxidative stress. Methods: We used transcriptomics, both spatial and traditional, to assess the impact of GFF on epidermal biology and homeostasis in keratinocytes (primary or immortalized) and in ex vivo skin explant tissue. The effect of GFF on cell adhesion rates, cellular ATP levels and proliferation rates were quantitated. Oxidative phosphorylation and glycolytic rates were measured under normal and stress‐induced conditions. Results: Transcriptomics from keratinocytes and ex vivo skin explants from multiple donors show GFF induces keratinocyte differentiation, skin barrier development and cell adhesion while simultaneously repressing cellular stress and inflammatory related processes. Spatial transcriptomics profiling of ex vivo skin indicated basal keratinocytes at the epidermal‐dermal junction and cornifying keratinocytes in the top layer of the epidermis as the primary cell types influenced by GFF treatment. Additionally, GFF significantly increases crosstalk between suprabasal and basal keratinocytes. To support these findings, we show that GFF can significantly increase cell adhesion and proliferation in keratinocytes. GFF also protected overall cellular bioenergetics under metabolic or oxidative stress conditions. Conclusion: Our findings provide novel insights into cellular differences and epidermal spatial localization in response to GFF, supporting previous findings that this filtrate has a significant impact on epidermal biology and homeostasis, particularly on spatially defined crosstalk. We propose that GFF can help maintain epidermal health by enhancing keratinocyte crosstalk and differentiation/proliferation balance as well as promoting an enhanced response to stress. [ABSTRACT FROM AUTHOR]

Epidermal keratinocytes undergo morphological and functional changes during differentiation, eventually being enucleated to become corneocytes. Calcium has been shown to be involved in various cellular functions of epidermal cells, including proliferation, differentiation, and apoptosis. Cerium is a lanthanide-series element and rare earth metal. For skin, cerium oxide has been investigated for use in absorbing UV and promoting wound healing. However, the functions and physiological effects of inorganic cerium on the skin have rarely been investigated. Here, we focused on cerium's function in epidermal keratinocytes and its interaction with calcium by investigating their effects on cell differentiation and intracellular calcium concentration. This study showed that applying cerium chloride to epidermal keratinocytes altered calcium signaling. It also suggested that cerium and calcium induced an increase in intracellular calcium concentration and promoted keratinocyte differentiation. [ABSTRACT FROM AUTHOR]

Background: Conspicuous facial pores are benign but represent a cosmetic concern for men and women. Recent works described dermal and epidermal impairments as clinical causes of enlarged pores. Morphological modifications of skin at the site of pores were associated with collagen density loss, possible alteration of extracellular matrix and abnormal differentiation of keratinocytes. Aims: A composition containing mannose‐6‐phosphate (Active Complex) was designed to address these different aspects of pore enlargement. In vitro and ex vivo evaluations were conducted in different models mimicking disturbance of dermal and epidermal functions. The pore refining activity of Active Complex was assessed in two clinical trials studying a Caucasian women cohort and an Asian men cohort. Results: At the dermal level, Active Complex upregulated collagen I and decorin synthesis, and genes encoding collagens I, III, V, VII, XVII; suggesting its ability to favor collagen fiber organization and anchorage. The downregulation of matrix metalloprotease, involved in extracellular matrix degradation, reinforced the protective effect of Active Complex in the dermis. Active Complex down modulated differentiation markers in keratinocytes as well as genes involved in cell renewal. Study of reconstructed human epidermis modeling keratinocyte hyperproliferation revealed that Active Complex mitigated two markers of this state: number of nuclei in the stratum corneum and involucrin expression. Clinical trials confirmed the pore refining activity of Active Complex on men and women of different ages and ethnicities; −24% total skin pore area after 56 days of application on women, and −30.2% on men after 7 days. Conclusions: This work demonstrates the interest to target dermal and epidermal modifications described in conspicuous pore area, especially dermis fiber organization, to address this cosmetic concern. [ABSTRACT FROM AUTHOR]

The list of standard abbreviations for JDS is available at adsa.org/jds-abbreviations-24. Nonstandard abbreviations are available in the Notes. Pododermatitis aseptica hemorrhagica circumscripta is associated with metalloproteinase 2 weakening of distal phalangeal suspensory structures and sinkage of the distal phalanx in the claw capsule. Pressure from the tuberculum flexorium on the sole epidermis and dermis produces hemorrhagic tissue injury and defective horn production appearing as yellow-red, softened claw horn in region 4 of the sole. A model of the MAPK/ERK signal cascade orchestrating epidermal-dermal homeostasis was employed to determine if sterile inflammatory responses are linked to disturbed signal transduction for epidermal homeostasis in sole epidermis and dermis. The objective was to assess shifts in target genes of inflammation, up- and downstream MAPK/ERK signal elements, and targeted genes supporting epidermal proliferation and differentiation. Sole epidermis and dermis were removed from lateral claws bearing lesions of PAHC, medial claws from the same limb and lateral claws from completely normal limbs of multiparous, lactating Holstein cows. The abundance levels of targeted transcripts were evaluated by real-time PCR. Lesion effects were assessed by ANOVA, and mean comparisons were performed with t -tests to assess variations between mean expression in ulcer-bearing or medial claw dermis and epidermis and completely normal lateral claw dermis and epidermis or between ulcer-bearing dermis and epidermis and medial claw dermis and epidermis. The lesions were sterile and showed losses across multiple growth factors, their receptors, several downstream AP1 transcription components, CMYC, multiple cell-cycle and terminal differentiation elements conducted by MAPK/ERK signals and β 4, α 6, and collagen 17A hemidesmosome components. These losses coincided with increased cytokeratin 6, β 1 integrin, proinflammatory metalloproteinases 2 and 9, IL1B and physiologic inhibitors of IL1B, the decoy receptor, and receptor antagonist. Medial claw epidermis and dermis from limbs with lateral claws bearing PAHC showed reductions in upstream MAPK/ERK signal elements and downstream targets that paralleled those in hemorrhagic lesions. Inhibitors of IL1B increased in the absence of real increases in inflammatory targets in the medial claw dermis and epidermis. Losses across multiple signal path elements and downstream targets were associated with negative effects on targeted transcripts supporting claw horn production and wound repair across lesion-bearing lateral claws and lesion-free medial claw dermis and epidermis. It was unclear if the sterile inflammation was causative or a consequence of these perturbations. [ABSTRACT FROM AUTHOR]

Psoriasis is a common chronic inflammatory skin disease that significantly impacts the patients' quality of life. Recent studies highlighted the function of the interleukin (IL)-1 family member IL-38 in skin homeostasis and suggested an anti-inflammatory role for this cytokine in psoriasis. In this study, we generated mice specifically overexpressing the IL-38 protein in epidermal keratinocytes. We confirmed IL-38 overexpression in the skin by Western blotting. We further detected the protein by ELISA in the plasma, as well as in conditioned media of skin explants isolated from IL-38 overexpressing mice, indicating that IL-38 produced in the epidermis is released from keratinocytes and can be found in the circulation. Unexpectedly, epidermal IL-38 overexpression did not impact the global severity of imiquimod (IMQ)-induced skin inflammation, Similarly, keratinocyte activation and differentiation in IMQ-treated skin were not affected by increased IL-38 expression and there was no global effect on local or systemic inflammatory responses. Nevertheless, we observed a selective inhibition of CXCL1 and IL-6 production in response to IMQ in IL-38 overexpressing skin, as well as reduced Ly6g mRNA levels, suggesting decreased neutrophil infiltration. Epidermal IL-38 overexpression also selectively affected the desquamation process during IMQ-induced psoriasis, as illustrated by reduced plaque formation. Taken together, our results validate the generation of a new mouse line allowing for tissue-specific IL-38 overexpression. Interestingly, epidermal IL-38 overexpression selectively affected specific disease-associated readouts during IMQ-induced psoriasis, suggesting a more complex role of IL-38 in the inflamed skin than previously recognized. In particular, our data highlight a potential involvement of IL-38 in the regulation of skin desquamation. [ABSTRACT FROM AUTHOR]

A 3-dimensional culture system of keratinocytes achieves cornification as a terminal differentiation that can mimic the formation of stratified epidermis. At the onset of keratinocyte differentiation, air-exposure treatment is essential for promotion. We have previously reported that the stimulation of differentiation is accompanied by downregulation of the transcriptional activity of the hypoxia-inducible factor (HIF) and also found that rocking treatment of cultured keratinocytes in the submerged condition restored their differentiation. A comparative study of cultured keratinocytes with and without rocking was then carried out to investigate the characteristics of the recovered differentiation by morphological and biochemical analyses. In addition, transcriptome analysis revealed the expected similar pattern between air-exposed and rocking cultures, including HIF-regulating transcripts. Furthermore, the promotive effect of rocking treatment was impaired under hypoxic culture conditions (1% O2). We showed that the restored promotion of differentiation by rocking culture is mainly due to the abrogation of transcriptional events by hypoxia. [ABSTRACT FROM AUTHOR]

The dermis and epidermis, crucial structural layers of the skin, encompass appendages, hair follicles (HFs), and intricate cellular heterogeneity. However, an integrated spatiotemporal transcriptomic atlas of embryonic skin has not yet been described and would be invaluable for studying skin‐related diseases in humans. Here, single‐cell and spatial transcriptomic analyses are performed on skin samples of normal and hairless fetal pigs across four developmental periods. The cross‐species comparison of skin cells illustrated that the pig epidermis is more representative of the human epidermis than mice epidermis. Moreover, Phenome‐wide association study analysis revealed that the conserved genes between pigs and humans are strongly associated with human skin‐related diseases. In the epidermis, two lineage differentiation trajectories describe hair follicle (HF) morphogenesis and epidermal development. By comparing normal and hairless fetal pigs, it is found that the hair placode (Pc), the most characteristic initial structure in HFs, arises from progenitor‐like OGN+/UCHL1+ cells. These progenitors appear earlier in development than the previously described early Pc cells and exhibit abnormal proliferation and migration during differentiation in hairless pigs. The study provides a valuable resource for in‐depth insights into HF development, which may serve as a key reference atlas for studying human skin disease etiology using porcine models. [ABSTRACT FROM AUTHOR]

An important hallmark of radiation dermatitis is the impairment of the mitotic ability of the stem/progenitor cells in the basal cell layers due to radiation-induced DNA damage, leading to suppressed cell renewal in the epidermis. However, this mechanism alone does not adequately explain the complex pathogenesis of radiation-induced skin injury. In this review, we summarize the latest findings on the complex pathogenesis of radiation dermatitis and correlate these with the clinical features of radiation-induced skin reactions. The current studies show that skin exposure to ionizing radiation induces cellular senescence in the epidermal keratinocytes. As part of their epithelial stress response, these senescent keratinocytes secrete pro-inflammatory mediators, thereby triggering skin inflammation. Keratinocyte-derived cytokines and chemokines modulate intercellular communication with the immune cells, activating skin-resident and recruiting skin-infiltrating immune cells within the epidermis and dermis, thereby orchestrating the inflammatory response to radiation-induced tissue damage. The increased expression of specific chemoattractant chemokines leads to increased recruitment of neutrophils into the irradiated skin, where they release cytotoxic granules that are responsible for the exacerbation of an inflammatory state. Moreover, the importance of IL-17-expressing γδ-T cells to the radiation-induced hyperproliferation of keratinocytes was demonstrated, leading to reactive hyperplasia of the epidermis. Radiation-induced, reactive hyperproliferation of the keratinocytes disturbs the fine-tuned keratinization and cornification processes, leading to structural dysfunction of the epidermal barrier. In summary, in response to ionizing radiation, epidermal keratinocytes have important structural and immunoregulatory barrier functions in the skin, coordinating interacting immune responses to eliminate radiation-induced damage and to initiate the healing process. [ABSTRACT FROM AUTHOR]

The mammalian skin and its appendages depend on tightly coordinated differentiation of epithelial cells. Epidermal growth factor receptor (EGFR) pathway substrate 8 (EPS8) like 1 (EPS8L1) is enriched in the epidermis among human tissues and has also been detected in the epidermis of lizards. Here, we show by the analysis of single-cell RNA-sequencing data that EPS8L1 mRNA is co-expressed with filaggrin and loricrin in terminally differentiated human epidermal keratinocytes. Comparative genomics indicated that EPS8L1 is conserved in all main clades of mammals, whereas the orthologous gene has been lost in birds. Using a polyclonal antibody against EPS8L1, we performed an immunohistochemical screening of skin from diverse mammalian species and immuno-electron microscopy of human skin. EPS8L1 was detected predominantly in the granular layer of the epidermis in monotremes, marsupial, and placental mammals. The labeling was partly associated with cell membranes, and it was evident along the perimeter of keratinocytes at the transition with the cornified layer of the epidermis, similar to involucrin distribution. Basal, spinous, and the fully mature cornified layers lacked immunolabeling of EPS8L1. In addition to the epidermis, the hair follicle inner root sheath (IRS) was immunolabeled. Both epidermal granular layer and IRS contribute to the barrier function of the skin, suggesting that EPS8L1 is involved in the regulation of these barriers. [ABSTRACT FROM AUTHOR]

The epidermal differentiation complex (EDC) is a cluster of genes that encode protein components of the outermost layers of the epidermis in mammals, reptiles and birds. The development of the stratified epidermis from a single-layered ectoderm involves an embryo-specific superficial cell layer, the periderm. An additional layer, the subperiderm, develops in crocodilians and over scutate scales of birds. Here, we review the expression of EDC genes during embryonic development. Several EDC genes are expressed predominantly or exclusively in embryo-specific cell layers, whereas others are confined to the epidermal layers that are maintained in postnatal skin. The S100 fused-type proteins scaffoldin and trichohyalin are expressed in the avian and mammalian periderm, respectively. Scaffoldin forms the so-called periderm granules, which are histological markers of the periderm in birds. Epidermal differentiation cysteine-rich protein (EDCRP) and epidermal differentiation protein containing DPCC motifs (EDDM) are expressed in the avian subperiderm where they are supposed to undergo cross-linking via disulfide bonds. Furthermore, a histidine-rich epidermal differentiation protein and feather-type corneous beta-proteins, also known as beta-keratins, are expressed in the subperiderm. The accumulating evidence for roles of EDC genes in the development of the epidermis has implications on the evolutionary diversification of the skin in amniotes. [ABSTRACT FROM AUTHOR]

Epidermis is the outer skin layer built of specialized cells called keratinocytes. Keratinocytes undergo a unique differentiation process, also known as cornification, during which their gene expression pattern, morphology and other properties change remarkably to the effect that the terminally differentiated, cornified cells can form a physical barrier, which separates the underlying tissues from the environment. Many genes encoding proteins that are important for epidermal barrier formation are located in a gene cluster called epidermal differentiation complex (EDC). Recent data provided valuable information on the dynamics of the EDC locus and the network of interactions between EDC gene promoters, enhancers and other regions, during keratinocytes differentiation. These data, together with results concerning changes in epigenetic modifications, provide a valuable insight into the mode of regulation of EDC gene expression. [ABSTRACT FROM AUTHOR]

Because of the crucial role of epidermal permeability barrier in regulation of cutaneous and extracutaneous functions, great efforts have been made to identify and develop the regimens that can improve epidermal permeability barrier function. Studies have demonstrated that oral administration of natural ingredients can improve epidermal permeability barrier in various skin conditions, including inflammatory dermatoses and UV-irradiation. Moreover, topical applications of some natural ingredients can also accelerate the repair of epidermal permeability barrier after acute barrier disruption and lower transepidermal water loss in the intact skin. Natural ingredient-induced improvements in epidermal permeability barrier function can be attributable to upregulation of keratinocyte differentiation, lipid production, antioxidant, hyaluronic acid production, expression of aquaporin 3 and sodium-hydrogen exchanger 1. In this review, we summarize the benefits of topical natural ingredients in epidermal permeability barrier in normal skin with or without acute barrier disruption and the underlying mechanisms. [ABSTRACT FROM AUTHOR]

Background: The skin, particularly the epidermis, is subjected to various external stresses, including ultraviolet (UV) irradiation. UV irradiation, mainly UVB at wavelength of 280–315 nm, can alter several epidermal functions, including cutaneous inflammation, epidermal hyperproliferation, DNA damage, disruption of epidermal permeability barrier and reduction in stratum corneum hydration levels. Because of the negative impacts of UVB irradiation on epidermal functions, great efforts have been made to develop regimens for the protection of alterations in epidermal function induced by UV irradiation. Summary: While sunscreen can provide physical barrier to UV light, some natural ingredients can also effectively protect the skin from UVB irradiation-induced damages. Studies have demonstrated that either topical or oral administrations of some natural ingredients attenuate UVB irradiation-induced alterations in the epidermal function. The underlying mechanisms by which natural ingredients improve epidermal functions are attributable to antioxidation, stimulation of keratinocyte differentiation, increases in the content of epidermal natural moisturizers and inhibition of inflammation. Key Message: Some natural ingredients exhibit protective and therapeutical benefits in photo-induced epidermal dysfunctions via divergent mechanisms. [ABSTRACT FROM AUTHOR]

The epidermis serves many vital roles, including protecting the body from external influences and healing eventual injuries. It is maintained by an incredibly complex and perfectly coordinated keratinization process. In this process, desquamation is essential for the differentiation of epidermal basal progenitor cells into enucleated corneocytes, which subsequently desquamate through programmed death. Numerous factors control keratinocyte differentiation: epidermal growth factor, transforming growth factor-α, keratinocyte growth factor, interleukins IL-1-β and IL-6, elevated vitamin A levels, and changes in Ca2+ concentration. The backbone of the keratinocyte transformation process from mitotically active basal cells into fully differentiated, enucleated corneocytes is the expression of specific proteins and the creation of a Ca2+ and pH gradient at precise locations within the epidermis. Skin keratinization disorders (histologically characterized predominantly by dyskeratosis, parakeratosis, and hyperkeratosis) may be categorized into three groups: defects in the α-helical rod pattern, defects outside the α-helical rod domain, and disorders of keratin-associated proteins. Understanding the process of keratinization is essential for the pathogenesis of many dermatological diseases because improper desquamation and epidermopoiesis/keratinization (due to genetic mutations of factors or due to immune pathological processes) can lead to various conditions (ichthyoses, palmoplantar keratodermas, psoriasis, pityriasis rubra pilaris, epidermolytic hyperkeratosis, and others). [ABSTRACT FROM AUTHOR]

Keratinocytes are one of the primary cells affected by psoriasis inflammation. Our study aimed to delve deeper into their morphology, transcriptome, and epigenome changes in response to psoriasis-like inflammation. We created a novel cytokine mixture to mimic mild and severe psoriasis-like inflammatory conditions in cultured keratinocytes. Upon induction of inflammation, we observed that the keratinocytes exhibited a mesenchymal-like phenotype, further confirmed by increased VIM mRNA expression and results obtained from confocal microscopy. We performed RNA sequencing to achieve a more global view, revealing 858 and 6987 DEGs in mildly and severely inflamed keratinocytes, respectively. Surprisingly, we found that the transcriptome of mildly inflamed keratinocytes more closely mimicked that of the psoriatic epidermis transcriptome than the severely inflamed keratinocytes. Genes involved in the IL-17 pathway were a major contributor to the similarities of the transcriptomes between mildly inflamed KCs and psoriatic epidermis. Mild and severe inflammation led to the gene regulation of epigenetic modifiers such as HATs, HDACs, DNMTs, and TETs. Immunofluorescence staining revealed distinct 5-hmC patterns in inflamed versus control keratinocytes, and consistently low 5-mC intensity in both groups. However, the global DNA methylation assay detected a tendency of decreased 5-mC levels in inflamed keratinocytes versus controls. This study emphasizes how inflammation severity affects the transcriptomic similarity of keratinocytes to psoriatic epidermis and proves dynamic epigenetic regulation and adaptive morphological changes in inflamed keratinocytes. [ABSTRACT FROM AUTHOR]

The skin is a highly organized tissue composed of multiple layers and cell types that require coordinated cell to cell communication to maintain tissue homeostasis. In skin cancer, this organized structure and communication is disrupted, prompting the malignant transformation of healthy cells into melanoma, basal cell carcinoma or squamous cell carcinoma tumours. One such family of channel proteins critical for cellular communication is pannexins (PANX1, PANX2, PANX3), all of which are present in the skin. These heptameric single-membrane channels act as conduits for small molecules and ions like ATP and Ca2+ but have also been shown to have channel-independent functions through their interacting partners or action in signalling pathways. Pannexins have diverse roles in the skin such as in skin development, aging, barrier function, keratinocyte differentiation, inflammation, and wound healing, which were discovered through work with pannexin knockout mice, organotypic epidermis models, primary cells, and immortalized cell lines. In the context of cutaneous cancer, PANX1 is present at high levels in melanoma tumours and functions in melanoma carcinogenesis, and both PANX1 and PANX3 expression is altered in non-melanoma skin cancer. PANX2 has thus far not been implicated in any skin cancer. This review will discuss pannexin isoforms, structure, trafficking, post-translational modifications, interactome, and channel activity. We will also outline the expression, localization, and function of pannexin channels within the diverse cell types of the epidermis, dermis, hypodermis, and adnexal structures of the skin, and how these properties are exploited or abrogated in instances of skin cancer. [ABSTRACT FROM AUTHOR]

Epidermal development is responsible for the formation of the outermost layer of the skin, the epidermis. The establishment of the epidermal barrier is a critical aspect of mammalian development. Proper formation of the epidermis, which is composed of stratified squamous epithelial cells, is essential for the survival of terrestrial vertebrates because it acts as a crucial protective barrier against external threats such as pathogens, toxins, and physical trauma. In mammals, epidermal development begins from the embryonic surface ectoderm, which gives rise to the basal layer of the epidermis. This layer undergoes a series of complex processes that lead to the formation of subsequent layers, including the stratum intermedium, stratum spinosum, stratum granulosum, and stratum corneum. The stratum corneum, which is the topmost layer of the epidermis, is formed by corneoptosis, a specialized form of cell death. This process involves the transformation of epidermal keratinocytes in the granular layer into flattened dead cells, which constitute the protective barrier. In this review, we focus on the intricate mechanisms that drive the development and establishment of the mammalian epidermis to gain insight into the complex processes that govern this vital biological system. [ABSTRACT FROM AUTHOR]

Peptidylarginine deiminases (PADs) transform a protein arginine residue into the non-standard amino acid citrulline. This calcium-dependent post-translational modification of proteins is called citrullination or deimination. As described in this special issue, PADs play a role in various physiological processes, and PAD deregulations are involved in many human diseases. Three PADs are expressed in the epidermis, where their roles begin to be deciphered. PAD1 and PAD3 are involved in keratinocyte differentiation, particularly in the epidermal barrier function, keratins, filaggrin and filaggrin-related proteins being the most abundant deiminated epidermal proteins. Reduced amounts of deiminated proteins and PAD1 expression may be involved in the pathogenesis of psoriasis and atopic dermatitis, two very frequent and chronic skin inflammatory diseases. The trichohyalin/PAD3/transglutaminase three pathway is important for hair shaft formation. Mutations of the PADI3 gene, leading to a decreased activity or abnormal localization of the corresponding isotype, are the cause of a rare hair disorder called uncombable hair syndrome, and are associated with the central centrifugal cicatricial alopecia, a frequent alopecia mainly affecting women of African ancestry. This article is part of the Theo Murphy meeting issue 'The virtues and vices of protein citrullination'. [ABSTRACT FROM AUTHOR]

The transparent corneal epithelium in the eye is maintained through the homeostasis regulated by limbal stem cells (LSCs), while the nontransparent epidermis relies on epidermal keratinocytes for renewal. Despite their cellular similarities, the precise cell fates of these two types of epithelial stem cells, which give rise to functionally distinct epithelia, remain unknown. We performed a multi-omics analysis of human LSCs from the cornea and keratinocytes from the epidermis and characterized their molecular signatures, highlighting their similarities and differences. Through gene regulatory network analyses, we identified shared and cell type-specific transcription factors (TFs) that define specific cell fates and established their regulatory hierarchy. Single-cell RNA-seq (scRNA-seq) analyses of the cornea and the epidermis confirmed these shared and cell type-specific TFs. Notably, the shared and LSC-specific TFs can cooperatively target genes associated with corneal opacity. Importantly, we discovered that FOSL2, a direct PAX6 target gene, is a novel candidate associated with corneal opacity, and it regulates genes implicated in corneal diseases. By characterizing molecular signatures, our study unveils the regulatory circuitry governing the LSC fate and its association with corneal opacity. This study uses multiomics and gene regulatory network analysis to compare the limbal stem cells that maintain the cornea with keratinocytes which maintain the skin, revealing an intriguing interplay of transcription factors that drive epithelial similarities and differences in health and disease. [ABSTRACT FROM AUTHOR]

A complete in vitro skin model, containing resident cell types is needed to understand physiology and to consider the role of immune and endothelial cells in dermal drug testing. In this study, a cell extraction technique was developed to isolate resident skin cells from the same human donor while preserving the immune and endothelial cells. Then those cells were used to reconstruct an autologous, vascularized, and immunocompetent Tissue-Engineered Skin model, aviTES. Phenotypic characterization of the viable cells was performed on freshly isolated cells and after thawing through flow cytometry. Dermal cell extracts were characterized as fibroblasts, endothelial and immune cells, and the average amount of each cell type represents 4, 0.5, and 1 million viable cells per g of the dermis, respectively. The 3D models, TES and aviTES, were characterized by a fully differentiated epidermis that showed an increase in the presence of Ki67+ cells in the basolateral layer of the aviTES model. Capillary-like network formation, through the self-assembly of endothelial cells, and the presence of functional immune cells were identified through immunofluorescence staining in aviTES. In addition, the aviTES model was immunocompetent, as evidenced by its capacity to increase the production of pro-inflammatory cytokines TNF-α, MIP-1α, and GM-CSF following LPS stimulation. This study describes an autologous skin model containing a functional resident skin immune system and a capillary network. It provides a relevant tool to study the contribution of the immune system to skin diseases and inflammatory responses and to investigate resident skin cell interactions and drug development. There is an urgent need for a complete in vitro skin model containing the resident cell types to better understand the role of immune and endothelial cells in skin and to be able to use it for drug testing. Actual 3D models of human skin most often contain only fibroblasts and keratinocytes with a limited number of models containing endothelial cells or a limited variety of immune cells. This study describes an autologous skin model containing a functional resident skin immune system and a capillary network. It provides a relevant tool to study the contribution of the immune system to skin diseases and inflammatory responses and to investigate interactions between resident skin cell, improving our capacity to develop new drugs. [Display omitted] [ABSTRACT FROM AUTHOR]

The skin epidermis exhibits an asymmetric structure composed of multilayered keratinocytes and those in the outer layers form two-way physical barriers, cornified cell envelope (CCE), and tight junctions (TJs). While undifferentiated keratinocytes in the basal layer continuously deliver daughter cells outward, which undergo successive differentiation with losing their polarized characteristics, they retain the expression of several polarity proteins. In the present study, we revealed that the t-SNARE protein syntaxin3, a critical element for the formation of the apical compartment in simple epithelial cells, is required to confer the ability to organize the physical barriers on "poorly polarized" keratinocytes in epidermal outer layers. HaCaT keratinocytes with genetic ablation of syntaxin3 readily succumbed to hydrogen peroxide-induced cell death. Additionally, they lost the ability to organize TJ and CCE structures, accompanied by notable downregulation of transglutaminase1 and caspase14 (a cornification regulator) expression. These syntaxin3-knockout cells appeared to restore oxidative stress tolerance and functional TJ formation ability, in response to the inducible re-expression of exogenous syntaxin3. While plausible mechanisms underlying these phenomena remain unclear, syntaxin3, an apical polarity protein in the simple epithelia, has emerged as a potentially crucial element for barrier formation in poorly polarized keratinocytes in polarized epidermal tissue. [ABSTRACT FROM AUTHOR]

Atopic dermatitis (AD) is a chronic inflammatory skin disease featuring skin barrier dysfunction and immune dysregulation. Previously, we reported that the retinoid-related orphan nuclear receptor RORα was highly expressed in the epidermis of normal skin. We also found that it positively regulated the expression of differentiation markers and skin barrier-related genes in human keratinocytes. In contrast, epidermal RORα expression was downregulated in the skin lesions of several inflammatory skin diseases, including AD. In this study, we generated mouse strains with epidermis-specific Rora ablation to understand the roles of epidermal RORα in regulating AD pathogenesis. Although Rora deficiency did not cause overt macroscopic skin abnormalities at the steady state, it greatly amplified MC903-elicited AD-like symptoms by intensifying skin scaliness, increasing epidermal hyperproliferation and barrier impairment, and elevating dermal immune infiltrates, proinflammatory cytokines, and chemokines. Despite the normal appearance at the steady state, Rora-deficient skin showed microscopic abnormalities, including mild epidermal hyperplasia, increased TEWL, and elevated mRNA expression of Krt16, Sprr2a, and Tslp genes, indicating subclinical impairment of epidermal barrier functions. Our results substantiate the importance of epidermal RORα in partially suppressing AD development by maintaining normal keratinocyte differentiation and skin barrier function. [ABSTRACT FROM AUTHOR]

The mammalian epidermis undergoes constant renewal, replenished by a pool of stem cells and terminal differentiation of their progeny. This is accompanied by changes in gene expression and morphology that are orchestrated, in part, by epigenetic modifiers. Here, we define the role of the histone acetyltransferase KAT2A in epidermal homeostasis and provide a comparative analysis that reveals key functional divergence with its paralog KAT2B. In contrast to the reported function of KAT2B in epidermal differentiation, KAT2A supports the undifferentiated state in keratinocytes. RNA-seq analysis of KAT2A- and KAT2B- depleted keratinocytes revealed dysregulated epidermal differentiation. Depletion of KAT2A led to premature expression of epidermal differentiation genes in the absence of inductive signals, whereas loss of KAT2B delayed differentiation. KAT2A acetyltransferase activity was indispensable in regulating epidermal differentiation gene expression. The metazoan-specific N terminus of KAT2A was also required to support its function in keratinocytes. We further showed that the interplay between KAT2A- and KAT2B-mediated regulation was important for normal cutaneous wound healing in vivo. Overall, these findings reveal a distinct mechanism in which keratinocytes use a pair of highly homologous histone acetyltransferases to support divergent functions in self-renewal and differentiation processes. [ABSTRACT FROM AUTHOR]

The article focuses on the innovative use of fibroblasts from volar skin to improve the quality of life for prosthetic limb users by enhancing the properties of nonvolar skin. Topics include the unique mechanical properties of volar skin compared to nonvolar skin, the clinical trial demonstrating the efficacy of injecting volar fibroblasts into nonvolar skin, and the potential for this technique to address issues like skin breakdown and discomfort for prosthesis wearers.

NRF2 is a master regulator of the cellular protection against oxidative damage in mammals and of multiple pathways relevant in the mammalian aging process. In the epidermis of the skin NRF2 contributes additionally to the formation of an antioxidant barrier to protect from environmental insults and is involved in the differentiation process of keratinocytes. In chronological aging of skin, the capacity for antioxidant responses and the ability to restore homeostasis after damage are impaired. Surprisingly, in absence of extrinsic stressors, NRF2 deficient mice do not show any obvious skin phenotype, not even at old age. We investigated the differences in chronological epidermal aging of wild type and NRF2‐deficient mice to identify the changes in aged epidermis that may compensate for absence of this important transcriptional regulator. While both genotypes showed elevated epidermal senescence markers (increased Lysophospholipids, decreased LaminB1 expression), the aged NRF2 deficient mice displayed disturbed epidermal differentiation manifested in irregular keratin 10 and loricrin expression. The tail skin displayed less age‐related epidermal thinning and a less pronounced decline in proliferating basal epidermal cells compared to the wildtype controls. The stratum corneum lipid composition also differed, as we observed elevated production of barrier protective linoleic acid (C18:2) and reduced abundance of longer chain saturated lignoceric acid (C24:0) among the stratum corneum fatty acids in the aged NRF2‐deficient mice. Thus, despite epidermal differentiation being disturbed in aged NRF2‐deficient animals in homeostasis, adaptations in keratinocyte proliferation and barrier lipid synthesis could explain the lack of a more severe phenotype. [ABSTRACT FROM AUTHOR]

Formation of the human skin epidermis can be reproduced by a three‐dimensional (3D) keratinocyte culture system, in which air‐exposure is inevitable upon initiation of differentiation. In the continuous submerged culture without air‐exposure, even with a differentiation‐compatible medium, several keratinocyte‐specific proteins were not induced resulting in the formation of aberrant epidermal layers. To clarify the mechanism by which air‐exposure promotes keratinocyte differentiation, we performed a comparative analysis on biological properties between submerged and air–liquid interphase culture systems. By transcriptomic analysis, hypoxia‐inducible factor (HIF)‐related genes appeared to significantly change in these cultured cells. In submerged culture, the transcriptional activity of HIF on its canonical response element was enhanced, while air‐exposure treatment drastically reduced the transcriptional activity despite the high HIF protein level. Regulating HIF activity through reagents and genetic manipulation revealed that the reduced but retained HIF‐transcriptional activity was essentially involved in differentiation. Furthermore, we showed, for the first time, that artificial supplementation of oxygen in the submerged culture system could restore keratinocyte differentiation as observed in the air‐exposed culture. Thus, we mechanistically evaluated how HIF regulates the air‐exposure‐dependent differentiation of keratinocytes in a 3D culture system. [ABSTRACT FROM AUTHOR]

The epidermis is a stratified squamous epithelium distributed in the outermost layer of the skin and is intimately involved in the formation of a physical barrier to pathogens. Basal keratinocytes possess the properties of stem cells and play an essential role in epidermal development and skin damage recovery. Therefore, understanding the molecular mechanism of how basal keratinocytes participate in epidermal development and stratification is vital for preventing and treating skin lesions. During epidermal morphogenesis, the symmetric division of basal keratinocytes contributes to the extension of skin tissues, while their asymmetric division and migration facilitate epidermal stratification. In this review, we summarize the process of epidermal stratification and illustrate the molecular mechanisms underlying epidermal morphogenesis. Furthermore, we discuss the coordination of multiple signaling pathways and transcription factors in epidermal stratification, together with the roles of cell polarity and cell dynamics during the process. [ABSTRACT FROM AUTHOR]

The disease-residual transcriptomic profile (DRTP) within psoriatic healed/resolved skin and epidermal tissue-resident memory T (TRM) cells have been proposed to be crucial for the recurrence of old lesions. However, it is unclear whether epidermal keratinocytes are involved in disease recurrence. There is increasing evidence regarding the importance of epigenetic mechanisms in the pathogenesis of psoriasis. Nonetheless, the epigenetic changes that contribute to the recurrence of psoriasis remain unknown. The aim of this study was to elucidate the role of keratinocytes in psoriasis relapse. The epigenetic marks 5-methylcytosine (5-mC) and 5-hydroxymethylcytosine (5-hmC) were visualized using immunofluorescence staining, and RNA sequencing was performed on paired never-lesional and resolved epidermal and dermal compartments of skin from psoriasis patients. We observed diminished 5-mC and 5-hmC amounts and decreased mRNA expression of the ten-eleven translocation (TET) 3 enzyme in the resolved epidermis. SAMHD1, C10orf99, and AKR1B10: the highly dysregulated genes in resolved epidermis are known to be associated with pathogenesis of psoriasis, and the DRTP was enriched in WNT, TNF, and mTOR signaling pathways. Our results suggest that epigenetic changes detected in epidermal keratinocytes of resolved skin may be responsible for the DRTP in the same regions. Thus, the DRTP of keratinocytes may contribute to site-specific local relapse. [ABSTRACT FROM AUTHOR]

The epidermal barrier of mammals is initially formed during embryonic development and continuously regenerated by the differentiation and cornification of keratinocytes in postnatal life. Cornification is associated with the breakdown of organelles and other cell components by mechanisms which are only incompletely understood. Here, we investigated whether heme oxygenase 1 (HO-1), which converts heme into biliverdin, ferrous iron and carbon monoxide, is required for normal cornification of epidermal keratinocytes. We show that HO-1 is transcriptionally upregulated during the terminal differentiation of human keratinocytes in vitro and in vivo. Immunohistochemistry demonstrated expression of HO-1 in the granular layer of the epidermis where keratinocytes undergo cornification. Next, we deleted the Hmox1 gene, which encodes HO-1, by crossing Hmox1-floxed and K14-Cre mice. The epidermis and isolated keratinocytes of the resulting Hmox1f/f K14-Cre mice lacked HO-1 expression. The genetic inactivation of HO-1 did not impair the expression of keratinocyte differentiation markers, loricrin and filaggrin. Likewise, the transglutaminase activity and formation of the stratum corneum were not altered in Hmox1f/f K14-Cre mice, suggesting that HO-1 is dispensable for epidermal cornification. The genetically modified mice generated in this study may be useful for future investigations of the potential roles of epidermal HO-1 in iron metabolism and responses to oxidative stress. [ABSTRACT FROM AUTHOR]

Pemphigus vulgaris is a life-threatening blistering skin disease caused by autoantibodies destabilizing desmosomal adhesion. Current therapies focus on suppression of autoantibody formation and thus treatments directly stabilizing keratinocyte adhesion would fulfill an unmet medical need. We here demonstrate that apremilast, a phosphodiesterase 4 inhibitor used in psoriasis, prevents skin blistering in pemphigus vulgaris. Apremilast abrogates pemphigus autoantibody-induced loss of keratinocyte cohesion in ex-vivo human epidermis, cultured keratinocytes in vitro and in vivo in mice. In parallel, apremilast inhibits keratin retraction as well as desmosome splitting, induces phosphorylation of plakoglobin at serine 665 and desmoplakin assembly into desmosomal plaques. We established a plakoglobin phospho-deficient mouse model that reveals fragile epidermis with altered organization of keratin filaments and desmosomal cadherins. In keratinocytes derived from these mice, intercellular adhesion is impaired and not rescued by apremilast. These data identify an unreported mechanism of desmosome regulation and propose that apremilast stabilizes keratinocyte adhesion and is protective in pemphigus. Pemphigus vulgaris is a life-threatening blistering skin disease caused by autoantibodies which destabilize cell adhesion of keratinocytes. The phosphodiesterase 4 inhibitor apremilast prevents skin blistering by stabilizing the keratin filament anchorage of desmosomes. [ABSTRACT FROM AUTHOR]