ResearchPad - keratinocytes https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[Application of co-culture technology of epithelial type cells and mesenchymal type cells using nanopatterned structures]]> https://www.researchpad.co/article/elastic_article_7654 Various nanopatterning techniques have been developed to improve cell proliferation and differentiation efficiency. As we previously reported, nanopillars and pores are able to sustain human pluripotent stem cells and differentiate pancreatic cells. From this, the nanoscale patterns would be effective environment for the co-culturing of epithelial and mesenchymal cell types. Interestingly, the nanopatterning selectively reduced the proliferative rate of mesenchymal cells while increasing the expression of adhesion protein in epithelial type cells. Additionally, co-cultured cells on the nanopatterning were not negatively affected in terms of cell function metabolic ability or cell survival. This is in contrast to conventional co-culturing methods such as ultraviolet or chemical treatments. The nanopatterning appears to be an effective environment for mesenchymal co-cultures with typically low proliferative rates cells such as astrocytes, neurons, melanocytes, and fibroblasts without using potentially damaging treatments.

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<![CDATA[Identification of Merkel cells associated with neurons in engineered skin substitutes after grafting to full thickness wounds]]> https://www.researchpad.co/article/5c8823d9d5eed0c484639153

Engineered skin substitutes (ESS), prepared using primary human fibroblasts and keratinocytes with a biopolymer scaffold, were shown to provide stable closure of excised burns, but relatively little is known about innervation of ESS after grafting. This study investigated innervation of ESS and, specifically, whether Merkel cells are present in healed grafts. Merkel cells are specialized neuroendocrine cells required for fine touch sensation in skin. We discovered cells positive for keratin 20 (KRT20), a general marker for Merkel cells, in the basal epidermis of ESS after transplantation to mice, suggesting the presence of Merkel cells. Cells expressing KRT20 were not observed in ESS in vitro. However, widely separated KRT20-positive cells were observed in basal epidermis of ESS by 2 weeks after grafting. By 4 weeks, these cells increased in number and expressed keratins 18 and 19, additional Merkel cells markers. Putative Merkel cell numbers increased further between weeks 6 and 14; their densities varied widely and no specific pattern of organization was observed, similar to Merkel cell localization in human skin. KRT20-positive cells co-expressed epidermal markers E-cadherin and keratin 15, suggesting derivation from the epidermal lineage, and neuroendocrine markers synaptophysin and chromogranin A, consistent with their identification as Merkel cells. By 4 weeks after grafting, some Merkel cells in engineered skin were associated with immature afferents expressing neurofilament-medium. By 8 weeks, Merkel cells were complexed with more mature neurons expressing neurofilament-heavy. Positive staining for human leukocyte antigen demonstrated that the Merkel cells in ESS were derived from grafted human cells. The results identify, for the first time, Merkel cell-neurite complexes in engineered skin in vivo. This suggests that fine touch sensation may be restored in ESS after grafting, although this must be confirmed with future functional studies.

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<![CDATA[RPGRIP1L is required for stabilizing epidermal keratinocyte adhesion through regulating desmoglein endocytosis]]> https://www.researchpad.co/article/5c58d63dd5eed0c48403195b

Cilia-related proteins are believed to be involved in a broad range of cellular processes. Retinitis pigmentosa GTPase regulator interacting protein 1-like (RPGRIP1L) is a ciliary protein required for ciliogenesis in many cell types, including epidermal keratinocytes. Here we report that RPGRIP1L is also involved in the maintenance of desmosomal junctions between keratinocytes. Genetically disrupting the Rpgrip1l gene in mice caused intraepidermal blistering, primarily between basal and suprabasal keratinocytes. This blistering phenotype was associated with aberrant expression patterns of desmosomal proteins, impaired desmosome ultrastructure, and compromised cell-cell adhesion in vivo and in vitro. We found that disrupting the RPGRIP1L gene in HaCaT cells, which do not form primary cilia, resulted in mislocalization of desmosomal proteins to the cytoplasm, suggesting a cilia-independent function of RPGRIP1L. Mechanistically, we found that RPGRIP1L regulates the endocytosis of desmogleins such that RPGRIP1L-knockdown not only induced spontaneous desmoglein endocytosis, as determined by AK23 labeling and biotinylation assays, but also exacerbated EGTA- or pemphigus vulgaris IgG-induced desmoglein endocytosis. Accordingly, inhibiting endocytosis with dynasore or sucrose rescued these desmosomal phenotypes. Biotinylation assays on cell surface proteins not only reinforced the role of RPGRIP1L in desmoglein endocytosis, but also suggested that RPGRIP1L may be more broadly involved in endocytosis. Thus, data obtained from this study advanced our understanding of the biological functions of RPGRIP1L by identifying its role in the cellular endocytic pathway.

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<![CDATA[Orf virus (ORFV) infection in a three-dimensional human skin model: Characteristic cellular alterations and interference with keratinocyte differentiation]]> https://www.researchpad.co/article/5c5b523fd5eed0c4842bc547

ORF virus (ORFV) is the causative agent of contagious ecthyma, a pustular dermatitis of small ruminants and humans. Even though the development of lesions caused by ORFV was extensively studied in animals, only limited knowledge exists about the lesion development in human skin. The aim of the present study was to evaluate a three-dimensional (3D) organotypic culture (OTC) as a human skin model for ORFV infection considering lesion development, replication of the virus, viral gene transcription and modulation of differentiation of human keratinocytes by ORFV. ORFV infection of OTC was performed using the ORFV isolate B029 derived from a human patient. The OTC sections showed a similar structure of stratified epidermal keratinocytes as human foreskin and a similar expression profile of the differentiation markers keratin 1 (K1), K10, and loricrin. Upon ORFV infection, OTCs exhibited histological cytopathic changes including hyperkeratosis and ballooning degeneration of the keratinocytes. ORFV persisted for 10 days and was located in keratinocytes of the outer epidermal layers. ORFV-specific early, intermediate and late genes were transcribed, but limited viral spread and restricted cell infection were noticed. ORFV infection resulted in downregulation of K1, K10, and loricrin at the transcriptional level without affecting proliferation as shown by PCNA or Ki-67 expression. In conclusion, OTC provides a suitable model to study the interaction of virus with human keratinocytes in a similar structural setting as human skin and reveals that ORFV infection downregulates several differentiation markers in the epidermis of the human skin, a hitherto unknown feature of dermal ORFV infection in man.

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<![CDATA[Epithelial stratification shapes infection dynamics]]> https://www.researchpad.co/article/5c52182bd5eed0c48479763d

Infections of stratified epithelia contribute to a large group of common diseases, such as dermatological conditions and sexually transmitted diseases. To investigate how epithelial structure affects infection dynamics, we develop a general ecology-inspired model for stratified epithelia. Our model allows us to simulate infections, explore new hypotheses and estimate parameters that are difficult to measure with tissue cell cultures. We focus on two contrasting pathogens: Chlamydia trachomatis and Human papillomaviruses (HPV). Using cervicovaginal parameter estimates, we find that key infection symptoms can be explained by differential interactions with the layers, while clearance and pathogen burden appear to be bottom-up processes. Cell protective responses to infections (e.g. mucus trapping) generally lowered pathogen load but there were specific effects based on infection strategies. Our modeling approach opens new perspectives for 3D tissue culture experimental systems of infections and, more generally, for developing and testing hypotheses related to infections of stratified epithelia.

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<![CDATA[Innate antimicrobial immunity in the skin: A protective barrier against bacteria, viruses, and fungi]]> https://www.researchpad.co/article/5c12cfa9d5eed0c484914b78 ]]> <![CDATA[Novelty upon novelty visualized by rotational scanning electron micrographs (rSEM): Laboulbeniales on the millipede order Chordeumatida]]> https://www.researchpad.co/article/5c0841c9d5eed0c484fcabc2

Laboulbeniales are highly specific ectoparasitic fungi of arthropods (insects, millipedes, and arachnids). The first Laboulbeniales parasitizing the millipede order Chordeumatida (Diplopoda) were discovered and described as a new dioecious genus of Laboulbeniales, Thaxterimyces, to accommodate the new species T. baliensis. Also the millipede host is a new species and is described as Metopidiothrix sheari. This is the first time Laboulbeniales fungus and its millipede host are described as new together. Males of Metopidiothrix have the most extensive secondary sexual modifications in the entire class Diplopoda. Although nothing is known about the function of these modifications, the unique pattern of Laboulbeniales infection in the new millipede species is obviously related to host sexual behavior. Rotational Scanning Electron Micrographs (rSEM) are used to create a 3D comprehensive model to examine the fungal-host interaction, a more advanced visualization of the ectoparasitic fungus on its host. Laboulbeniales diversity on millipedes is still understudied, and a consistent effort is needed to unveil and understand the extent and diversity of this biological interaction. Due to their minute size and apparently non-detrimental effect on their hosts, Laboulbeniales in general have been largely ignored by mycologists and neglected by generations of entomologists. As a result a significant component of global biodiversity has been strongly underestimated, and a wealth of new discoveries is still to be made both in the field and in existing museum collections.

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<![CDATA[The small molecule rhodomyrtone suppresses TNF-α and IL-17A-induced keratinocyte inflammatory responses: A potential new therapeutic for psoriasis]]> https://www.researchpad.co/article/5bce3d3840307c69b197f508

Psoriasis is a common skin disease pathogenically driven by TNF and IL-17A-induced epidermal hyperproliferation and inflammatory responses. The ongoing need for new therapeutic agents for psoriasis has highlighted medicinal plants as sources of phytochemicals useful for treating psoriatic disease. Rhodomyrtone, a bioactive phytochemical from Rhodomyrtus tomentosa, has well-established anti-proliferative activities. This study assessed the potential of rhodomyrtone for curtailing TNF/IL-17A-driven inflammation. Stimulating human skin organ cultures with TNF+IL-17A to model the skin inflammation in psoriasis, we found that rhodomyrtone significantly decreased inflammatory gene expression and the expression and secretion of inflammatory proteins, assessed by qRT-PCR, immunohistochemistry and ELISA assays respectively. RNA-seq analysis of monolayer primary keratinocytes treated with IL-17A/TNF showed that rhodomyrtone inhibited 724/1587 transcripts >2-fold altered by IL-17A/TNF (p<0.01), a number of which were confirmed at the mRNA and protein level. Suggesting that rhodomyrtone acts by modulating MAP kinase and NF-κB signaling pathways, rhodomyrtone inhibited TNF-induced ERK, JNK, p38, and NF-κBp65 phosphorylation. Finally, assessing the in vivo anti-inflammatory potential of rhodomyrtone, we examined its effects on imiquimod-induced skin inflammation in mice, finding rhodomyrtone reversed imiquimod-induced skin hyperplasia and epidermal thickening (p< 0.001). Taken together, these results suggest that rhodomyrtone may be useful in preventing or slowing the progression of inflammatory skin disease.

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<![CDATA[Low-dose ionizing radiation exposure represses the cell cycle and protein synthesis pathways in in vitro human primary keratinocytes and U937 cell lines]]> https://www.researchpad.co/article/5b498fb0463d7e0897c6e01f

The effects of the high-dose ionizing radiation used in radiotherapy have been thoroughly demonstrated in vitro and in vivo. However, the effects of low-dose ionizing radiation (LDIR) such as computed tomography-guided biopsies and X-ray fluoroscopy on skin cells remain controversial. This study investigated the molecular effects of LDIR on the human primary keratinocytes (HPKs) and U937 cells, monocytes-like cell lines. These cells were exposed to 0.1 Gray (Gy) X-ray as LDIR. The modulation of transcription was assessed using a cDNA array, and the protein expression after LDIR exposure was investigated using isobaric tags for relative and absolute quantification (iTRAQ) proteomic analysis at 24 hours. These effects were confirmed by immunoblotting analysis. The direct effects of LDIR on the U937 cells and HPKs and the bystander effects of irradiated HPKs on U937 cells were also investigated. LDIR downregulated c-Myc in both U937 cells and HPKs, and upregulated the p21WAF1/CIP1 protein expression in U937 cells along with the activation of TGFβ and protein phosphatase 2A (PP2A). In HPKs, LDIR downregulated the mTOR signaling with repression of S6 and 4EBP1 activation. Similar changes were observed as bystander effects of LDIR. Our findings suggest that LDIR inhibits protein synthesis and induces the cytokines activation associated with inflammation via direct and bystander effects, which might recapitulate the effects of LDIR in inflammated skin structures.

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<![CDATA[IL-1 Coordinates the Neutrophil Response to C. albicans in the Oral Mucosa]]> https://www.researchpad.co/article/5989d9ebab0ee8fa60b6c884

Mucosal infections with Candida albicans belong to the most frequent forms of fungal diseases. Host protection is conferred by cellular immunity; however, the induction of antifungal immunity is not well understood. Using a mouse model of oropharyngeal candidiasis (OPC) we show that interleukin-1 receptor (IL-1R) signaling is critical for fungal control at the onset of infection through its impact on neutrophils at two levels. We demonstrate that both the recruitment of circulating neutrophils to the site of infection and the mobilization of newly generated neutrophils from the bone marrow depended on IL-1R. Consistently, IL-1R-deficient mice displayed impaired chemokine production at the site of infection and defective secretion of granulocyte colony-stimulating factor (G-CSF) in the circulation in response to C. albicans. Strikingly, endothelial cells were identified as the primary cellular source of G-CSF during OPC, which responded to IL-1α that was released from keratinocytes in the infected tissue. The IL-1-dependent crosstalk between two different cellular subsets of the nonhematopoietic compartment was confirmed in vitro using a novel murine tongue-derived keratinocyte cell line and an established endothelial cell line. These data establish a new link between IL-1 and granulopoiesis in the context of fungal infection. Together, we identified two complementary mechanisms coordinating the neutrophil response in the oral mucosa, which is critical for preventing fungal growth and dissemination, and thus protects the host from disease.

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<![CDATA[Effects of Plectin Depletion on Keratin Network Dynamics and Organization]]> https://www.researchpad.co/article/5989dadcab0ee8fa60bba058

The keratin intermediate filament cytoskeleton protects epithelial cells against various types of stress and is involved in fundamental cellular processes such as signaling, differentiation and organelle trafficking. These functions rely on the cell type-specific arrangement and plasticity of the keratin system. It has been suggested that these properties are regulated by a complex cycle of assembly and disassembly. The exact mechanisms responsible for the underlying molecular processes, however, have not been clarified. Accumulating evidence implicates the cytolinker plectin in various aspects of the keratin cycle, i.e., by acting as a stabilizing anchor at hemidesmosomal adhesion sites and the nucleus, by affecting keratin bundling and branching and by linkage of keratins to actin filament and microtubule dynamics. In the present study we tested these hypotheses. To this end, plectin was downregulated by shRNA in vulvar carcinoma-derived A431 cells. As expected, integrin β4- and BPAG-1-positive hemidesmosomal structures were strongly reduced and cytosolic actin stress fibers were increased. In addition, integrins α3 and β1 were reduced. The experiments furthermore showed that loss of plectin led to a reduction in keratin filament branch length but did not alter overall mechanical properties as assessed by indentation analyses using atomic force microscopy and by displacement analyses of cytoplasmic superparamagnetic beads using magnetic tweezers. An increase in keratin movement was observed in plectin-depleted cells as was the case in control cells lacking hemidesmosome-like structures. Yet, keratin turnover was not significantly affected. We conclude that plectin alone is not needed for keratin assembly and disassembly and that other mechanisms exist to guarantee proper keratin cycling under steady state conditions in cultured single cells.

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<![CDATA[Three-dimensional cell shapes and arrangements in human sweat glands as revealed by whole-mount immunostaining]]> https://www.researchpad.co/article/5989db5fab0ee8fa60be116e

Because sweat secretion is facilitated by mechanical contraction of sweat gland structures, understanding their structure-function relationship could lead to more effective treatments for patients with sweat gland disorders such as heat stroke. Conventional histological studies have shown that sweat glands are three-dimensionally coiled tubular structures consisting of ducts and secretory portions, although their detailed structural anatomy remains unclear. To better understand the details of the three-dimensional (3D) coiled structures of sweat glands, a whole-mount staining method was employed to visualize 3D coiled gland structures with sweat gland markers for ductal luminal, ductal basal, secretory luminal, and myoepithelial cells. Imaging the 3D coiled gland structures demonstrated that the ducts and secretory portions were comprised of distinct tubular structures. Ductal tubules were occasionally bent, while secretory tubules were frequently bent and formed a self-entangled coiled structure. Whole-mount staining of complex coiled gland structures also revealed the detailed 3D cellular arrangements in the individual sweat gland compartments. Ducts were composed of regularly arranged cuboidal shaped cells, while secretory portions were surrounded by myoepithelial cells longitudinally elongated along entangled secretory tubules. Whole-mount staining was also used to visualize the spatial arrangement of blood vessels and nerve fibers, both of which facilitate sweat secretion. The blood vessels ran longitudinally parallel to the sweat gland tubules, while nerve fibers wrapped around secretory tubules, but not ductal tubules. Taken together, whole-mount staining of sweat glands revealed the 3D cell shapes and arrangements of complex coiled gland structures and provides insights into the mechanical contraction of coiled gland structures during sweat secretion.

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<![CDATA[Forsythia suspensa Suppresses House Dust Mite Extract-Induced Atopic Dermatitis in NC/Nga Mice]]> https://www.researchpad.co/article/5989db32ab0ee8fa60bd21af

Forsythia suspensa (F. suspensa) is a traditional medicine for treatment of inflammation. In this study, we evaluated the therapeutic effects of an ethanol extract from F. suspensa fruits on atopic dermatitis both in vivo and in vitro. We investigated the inhibitory effects of F. suspensa extract on the development of atopic dermatitis-like skin lesions in an NC/Nga mouse model exposed to Dermatophagoides farinae crude extract. Topical application of F. suspensa extract to the mice attenuated the atopic dermatitis symptoms, including increased dermatitis severity score, ear thickness, infiltration of inflammatory cells in the skin lesions, serum levels of IgE, TNF-α, and histamine, and expression of chemokines, cytokines, and adhesion molecules in ear tissue. In addition, F. suspensa extract inhibited the production of chemokines in TNF-α/IFN-γ-activated human keratinocytes. High-performance liquid chromatography analysis of FSE revealed the presence of four chemical constituents (forsythiaside, phillyrin, pinoresinol, and phylligenin). These compounds inhibited the production of chemokines in TNF-α/IFN-γ-activated human keratinocytes. These results suggest that the F. suspensa might be a useful candidate for treating allergic skin inflammatory disorders.

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<![CDATA[Cutaneous HPV8 and MmuPV1 E6 Proteins Target the NOTCH and TGF-β Tumor Suppressors to Inhibit Differentiation and Sustain Keratinocyte Proliferation]]> https://www.researchpad.co/article/5989db53ab0ee8fa60bdcee7

Cutaneous beta-papillomaviruses are associated with non-melanoma skin cancers that arise in patients who suffer from a rare genetic disorder, Epidermodysplasia verruciformis (EV) or after immunosuppression following organ transplantation. Recent studies have shown that the E6 proteins of the cancer associated beta human papillomavirus (HPV) 5 and HPV8 inhibit NOTCH and TGF-β signaling. However, it is unclear whether disruption of these pathways may contribute to cutaneous HPV pathogenesis and carcinogenesis. A recently identified papillomavirus, MmuPV1, infects laboratory mouse strains and causes cutaneous skin warts that can progress to squamous cell carcinoma. To determine whether MmuPV1 may be an appropriate model to mechanistically dissect the molecular contributions of cutaneous HPV infections to skin carcinogenesis, we investigated whether MmuPV1 E6 shares biological and biochemical activities with HPV8 E6. We report that the HPV8 and MmuPV1 E6 proteins share the ability to bind to the MAML1 and SMAD2/SMAD3 transcriptional cofactors of NOTCH and TGF-beta signaling, respectively. Moreover, we demonstrate that these cutaneous papillomavirus E6 proteins inhibit these two tumor suppressor pathways and that this ability is linked to delayed differentiation and sustained proliferation of differentiating keratinocytes. Furthermore, we demonstrate that the ability of MmuPV1 E6 to bind MAML1 is necessary for papilloma formation in experimentally infected mice. Our results, therefore, suggest that experimental MmuPV1 infection in mice will be a robust and useful experimental system to model key aspects of cutaneous HPV infection, pathogenesis and carcinogenesis.

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<![CDATA[Forkhead Box C1 Regulates Human Primary Keratinocyte Terminal Differentiation]]> https://www.researchpad.co/article/5989daefab0ee8fa60bc0bfa

The epidermis serves as a critical protective barrier between the internal and external environment of the human body. Its remarkable barrier function is established through the keratinocyte (KC) terminal differentiation program. The transcription factors specifically regulating terminal differentiation remain largely unknown. Using a RNA-sequencing (RNA-seq) profiling approach, we found that forkhead box c 1 (FOXC1) was significantly up-regulated in human normal primary KC during the course of differentiation. This observation was validated in human normal primary KC from several different donors and human skin biopsies. Silencing FOXC1 in human normal primary KC undergoing differentiation led to significant down-regulation of late terminal differentiation genes markers including epidermal differentiation complex genes, keratinization genes, sphingolipid/ceramide metabolic process genes and epidermal specific cell-cell adhesion genes. We further demonstrated that FOXC1 works down-stream of ZNF750 and KLF4, and upstream of GRHL3. Thus, this study defines FOXC1 as a regulator specific for KC terminal differentiation and establishes its potential position in the genetic regulatory network.

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<![CDATA[Additive Effects of Millimeter Waves and 2-Deoxyglucose Co-Exposure on the Human Keratinocyte Transcriptome]]> https://www.researchpad.co/article/5989db2dab0ee8fa60bd1bff

Millimeter Waves (MMW) will be used in the next-generation of high-speed wireless technologies, especially in future Ultra-Broadband small cells in 5G cellular networks. Therefore, their biocompatibilities must be evaluated prior to their massive deployment. Using a microarray-based approach, we analyzed modifications to the whole genome of a human keratinocyte model that was exposed at 60.4 GHz-MMW at an incident power density (IPD) of 20 mW/cm2 for 3 hours in athermic conditions. No keratinocyte transcriptome modifications were observed. We tested the effects of MMWs on cell metabolism by co-treating MMW-exposed cells with a glycolysis inhibitor, 2-deoxyglucose (2dG, 20 mM for 3 hours), and whole genome expression was evaluated along with the ATP content. We found that the 2dG treatment decreased the cellular ATP content and induced a high modification in the transcriptome (632 coding genes). The affected genes were associated with transcriptional repression, cellular communication and endoplasmic reticulum homeostasis. The MMW/2dG co-treatment did not alter the keratinocyte ATP content, but it did slightly alter the transcriptome, which reflected the capacity of MMW to interfere with the bioenergetic stress response. The RT-PCR-based validation confirmed 6 MMW-sensitive genes (SOCS3, SPRY2, TRIB1, FAM46A, CSRNP1 and PPP1R15A) during the 2dG treatment. These 6 genes encoded transcription factors or inhibitors of cytokine pathways, which raised questions regarding the potential impact of long-term or chronic MMW exposure on metabolically stressed cells.

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<![CDATA[Esophageal Epithelial-Derived IL-33 Is Upregulated in Patients with Heartburn]]> https://www.researchpad.co/article/5989da70ab0ee8fa60b9476e

Background

Interleukin-33 (IL-33) is a tissue-derived cytokine that is constitutively expressed in epithelial cells of tissues exposed to the environment and plays a role in sensing damage caused by inflammatory diseases. IL-33 acts as both a traditional cytokine and as a chromatin-associated nuclear factor in both innate and adaptive immunity. We recently showed that IL-33 in esophageal mucosa is upregulated in reflux esophagitis. However, IL-33 expression in patients with heartburn without mucosal injury and its relationship with intercellular space (ICS) have never been examined. We therefore examined the expression of cytokines and ICS in patients with heartburn.

Methods

The expression of IL-33 in the middle and distal esophageal mucosa of patients with heartburn without mucosal break and control samples was examined using real-time RT-PCR and immunofluorescence. The mRNA expression of IL-6, IL-8, MCP-1, and RANTES, and ICS was also analyzed.

Results

IL-33 expression and the mean ICS were significantly increased in the mucosa of patients with heartburn compared to that of the control. IL-33 and ICS were not different between the patients who were taking a PPI and those who were not. The upregulated IL-33 expression in the heartburn group was located in the nuclei of the basal cell layer. Although IL-6, IL-8, MCP-1 and RANTES levels were not different between control and patients with heartburn samples, IL-33 mRNA levels were still significantly correlated with IL-6, IL-8, or MCP-1 mRNA levels.

Conclusion

Nuclear IL-33 is upregulated in patients with heartburn. Upregulated IL-33 in heartburn patients is related to the symptoms.

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<![CDATA[Improved epidermal barrier formation in human skin models by chitosan modulated dermal matrices]]> https://www.researchpad.co/article/5989db50ab0ee8fa60bdc085

Full thickness human skin models (FTMs) contain an epidermal and a dermal equivalent. The latter is composed of a collagen dermal matrix which harbours fibroblasts. Current epidermal barrier properties of FTMs do not fully resemble that of native human skin (NHS), which makes these human skin models less suitable for barrier related studies. To further enhance the resemblance of NHS for epidermal morphogenesis and barrier formation, we modulated the collagen dermal matrix with the biocompatible polymer chitosan. Herein, we report that these collagen-chitosan FTMs (CC-FTMs) possess a well-organized epidermis and maintain both the early and late differentiation programs as in FTMs. Distinctively, the epidermal cell activation is reduced in CC-FTMs to levels observed in NHS. Dermal-epidermal interactions are functional in both FTM types, based on the formation of the basement membrane. Evaluation of the barrier structure by the organization of the extracellular lipid matrix of the stratum corneum revealed an elongated repeat distance of the long periodicity phase. The ceramide composition exhibited a higher resemblance of the NHS, based on the carbon chain-length distribution and subclass profile. The inside-out barrier functionality indicated by the transepidermal water loss is significantly improved in the CC-FTMs. The expression of epidermal barrier lipid processing enzymes is marginally affected, although more restricted to a single granular layer. The novel CC-FTM resembles the NHS more closely, which makes them a promising tool for epidermal barrier related studies.

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<![CDATA[Th17 micro-milieu regulates NLRP1-dependent caspase-5 activity in skin autoinflammation]]> https://www.researchpad.co/article/5989db52ab0ee8fa60bdc912

IL-1β is a potent player in cutaneous inflammation and central for the development of a Th17 micro-milieu in autoinflammatory diseases including psoriasis. Its production is controlled at the transcriptional level and by subsequent posttranslational processing via inflammatory caspases. In this study, we detected inflammatory caspase-5 active in epidermal keratinocytes and in psoriatic skin lesions. Further, interferon-γ and interleukin-17A synergistically induced caspase-5 expression in cultured keratinocytes, which was dependent on the antimicrobial peptide psoriasin (S100A7). However, diseases-relevant triggers for caspase-5 activity and IL-1β production remain unknown. Recently, extranuclear DNA has been identified as danger-signals abundant in the psoriatic epidermis. Here, we could demonstrate that cytosolic double-stranded (ds) DNA transfected into keratinocytes triggered the activation of caspase-5 and the release of IL-1β. Further, interleukin-17A promoted caspase-5 function via facilitation of the NLRP1-inflammasome. Anti-inflammatory vitamin D interfered with the IL-1β release and suppressed caspase-5 in keratinocytes and in psoriatic skin lesions. Our data link the disease-intrinsic danger signals psoriasin (S100A7) and dsDNA for NLPR1-dependent caspase-5 activity in psoriasis providing potential therapeutic targets in Th17-mediated skin autoinflammation.

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<![CDATA[The CXCL12/CXCR4 Signaling Pathway: A New Susceptibility Factor in Human Papillomavirus Pathogenesis]]> https://www.researchpad.co/article/5989da71ab0ee8fa60b9500f

The productive human papillomavirus (HPV) life cycle is tightly linked to the differentiation and cycling of keratinocytes. Deregulation of these processes and stimulation of cell proliferation by the action of viral oncoproteins and host cell factors underlies HPV-mediated carcinogenesis. Severe HPV infections characterize the wart, hypogammaglobulinemia, infection, and myelokathexis (WHIM) immunodeficiency syndrome, which is caused by gain-of-function mutations in the CXCR4 receptor for the CXCL12 chemokine, one of which is CXCR41013. We investigated whether CXCR41013 interferes in the HPV18 life cycle in epithelial organotypic cultures. Expression of CXCR41013 promoted stabilization of HPV oncoproteins, thus disturbing cell cycle progression and proliferation at the expense of the ordered expression of the viral genes required for virus production. Conversely, blocking CXCR41013 function restored virus production and limited HPV-induced carcinogenesis. Thus, CXCR4 and its potential activation by genetic alterations in the course of the carcinogenic process can be considered as an important host factor for HPV carcinogenesis.

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