bromochloroacetic-acid and Dermatitis--Atopic

The skin is the largest organ of the mammalian body. The outermost layer of mammalian skin, the stratum corneum (SC) of the epidermis, consists of piles of dead corneocytes that are the end-products of terminal differentiation of epidermal keratinocytes. The SC performs a crucial barrier function of epidermis. Langerhans cells, when activated, extend their dendrites through tight junctions just beneath the SC to capture external antigens. Recently, knowledge of the biology of corneocytes ('corneobiology') has progressed rapidly and many key factors that modulate its barrier function have been identified and characterized. In this review article on the SC, we summarize its evolution, formation, structure and function. Cornification is an important step of SC formation at the conversion of living epithelial cells to dead corneocytes, and consists of three major steps: formation of the intracellular keratin network, cornified envelopes and intercellular lipids. After cornification, the SC undergoes chemical reactions to form the mature SC with different functional layers. Finally, the SC is shed off at the surface ('desquamation'), mediated by a cascade of several proteases. This review will be helpful to understand our expanding knowledge of the biology of the SC, where immunity meets external antigens.

Topics: Animals; Cell Differentiation; Dermatitis, Atopic; Epidermis; Humans; Hypersensitivity; Immunity; Keratinocytes; Keratins; Langerhans Cells; Lipid Metabolism

Ceramides are epidermal lipids important for normal skin barrier function. Reduced Ceramide content is associated with atopic dermatitis (AD). House dust mite (HDM) has been localized in AD skin where it plays an exacerbator role. We set to examine the impact of HDM on skin integrity and the effect of three separate Ceramides (AD™, DS, Y30) on HDM-induced cutaneous damage. The effect was tested in vitro on primary human keratinocytes and ex vivo on skin explants. HDM (100 μg/mL) decreased the expression of adhesion protein E-cadherin, supra-basal (K1, K10) and basal (K5, K14) keratins and increased matrix metallopeptidase (MMP)-9 activity. The presence of Ceramide AD™ in topical cream inhibited HDM-induced E-cadherin and keratin destruction and dampened MMP-9 activity ex vivo which was not seen for the control cream or cream containing DS or Y30 Ceramides. The efficacy of Ceramide AD™ was tested in a clinical setting on moderate to very dry skin (as surrogate for environment-induced skin damage). When applied topically for 21 days, Ceramide AD™ significantly reduced transepidermal water loss (TEWL) in patients with very dry skin compared to their TEWL baseline data. Our study demonstrates Ceramide AD™ cream to be effective in restoring skin homeostasis and barrier function in damaged skin and warrants testing in larger clinical trials for possible treatment of AD and xerosis.

Topics: Animals; Ceramides; Dermatitis, Atopic; Dermatophagoides pteronyssinus; Emollients; Epidermis; Humans; Keratins; Pyroglyphidae; Skin

Atopic dermatitis (AD)/atopic eczema is a chronic relapsing inflammatory skin disease affecting nearly 14% of the adult population. An important pathogenetic pillar in AD is the disrupted skin barrier function (SBF). The atopic stratum corneum (SC) has been examined using several methods, including Raman microspectroscopy, yet so far, there is no depth-dependent analysis over the entire SC thickness. Therefore, we recruited 21 AD patients (9 female, 12 male) and compared the lesional (LAS) with non-lesional atopic skin (nLAS) in vivo with confocal Raman microspectroscopy. Our results demonstrated decreased total intercellular lipid and carotenoid concentrations, as well as a shift towards decreased orthorhombic lateral lipid organisation in LAS. Further, we observed a lower concentration of natural moisturising factor (NMF) and a trend towards increased strongly bound and decreased weakly bound water in LAS. Finally, LAS showed an altered secondary and tertiary keratin structure, demonstrating a more folded keratin state than nLAS. The obtained results are discussed in comparison with healthy skin and yield detailed insights into the atopic SC structure. LAS clearly shows molecular alterations at certain SC depths compared with nLAS which imply a reduced SBF. A thorough understanding of these alterations provides useful information on the aetiology of AD and for the development/control of targeted topical therapies.

Topics: Adult; Dermatitis, Atopic; Epidermis; Female; Humans; Keratins; Lipids; Male; Neoplasm Recurrence, Local; Skin

Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by skin barrier dysfunction. Sargassum horneri (S. horneri) is a brown alga that has been widely used in traditional medicine of eastern Asian countries. Recent studies proved that a brown alga S. horneri has anti-inflammatory activity. In this study, we investigated the effect of S. horneri ethanol extract (SHE) against AD in 2,4-dinitrobenzene (DNCB) induced AD in NC/Nga mice. We observed that SHE treatment decreased the epidermal thickness and epidermal hyperplasia that had been worsened through DNCB application. Moreover, SHE significantly inhibited the proliferation of mast cells and decreased the expression of IL-13 on CD4⁺ cells prompted by elevated thymic stromal lymphopoietin (TSLP) expression in DNCB-induced AD in mice. We also demonstrated that SHE directly inhibited the expression of keratinocyte-produced TSLP known to exacerbate skin barrier impairment. Especially, the decrease of filaggrin, an integral component of proper skin barrier function through a function in aggregating keratin filaments, observed in DNCB-induced AD mice was significantly improved when treated with SHE. More importantly, we proved that SHE was able to decrease the serum levels of IgG₁ and IgG₂ₐ, two crucial factors of AD, indicating the protective effect of SHE. Taken together, our findings suggest that SHE may protect NC/Nga mice against DNCB-induced AD via promoting skin barrier function.

Topics: Animals; Anti-Inflammatory Agents; Cytokines; Dermatitis, Atopic; Dinitrobenzenes; Immunoglobulin G; Interleukin-13; Keratins; Mice; Plant Extracts; Polyphenols; Sargassum; Skin; Skin Diseases

Dominant-negative mutations associated with signal transducer and activator of transcription 3 (STAT3) signaling, which controls epithelial proliferation in various tissues, lead to atopic dermatitis in hyper IgE syndrome. This dermatitis is thought to be attributed to defects in STAT3 signaling in type 17 helper T cell　specification. However, the role of STAT3 signaling in skin epithelial cells remains unclear. We found that STAT3 signaling in keratinocytes is required to maintain skin homeostasis by negatively controlling the expression of hair follicle-specific keratin genes. These expression patterns correlated with the onset of dermatitis, which was observed in specific pathogen-free conditions but not in germ-free conditions, suggesting the involvement of Toll-like receptor-mediated inflammatory responses. Thus, our study suggests that STAT3-dependent gene expression in keratinocytes plays a critical role in maintaining the homeostasis of skin, which is constantly exposed to microorganisms.

Topics: Animals; Dermatitis, Atopic; Disease Models, Animal; Female; Gene Expression Regulation; Hair Follicle; Homeostasis; Humans; Keratinocytes; Keratins; Male; Mice; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Knockout; Signal Transduction; Skin; Skin Physiological Phenomena; STAT3 Transcription Factor; Th17 Cells

Hyperkeratotic hand eczema (HHE) is a typical clinical hand eczema subtype with a largely unknown pathophysiology.. To investigate histopathology, expression of keratins (K), epidermal barrier proteins, and adhesion molecules in HHE.. Palmar skin biopsies (lesional and perilesional) were obtained from seven HHE patients and two healthy controls. Moreover, 135 candidate genes associated with palmoplantar keratoderma were screened for mutations.. Immunofluorescence staining showed a significant reduction of K9 and K14 in lesional skin. Upregulation was found for K5, K6, K16, and K17 in lesional skin compared with perilesional and healthy palmar skin. Further, upregulation of involucrin and alternating loricrin staining, both in an extracellular staining pattern, was found. Filaggrin expression was similar in lesional, perilesional, and control skin. No monogenetic mutations were found.. Currently, the phenotype of HHE is included in the hand eczema classification system; however, it can be argued whether this is justified. The evident expression of filaggrin and involucrin in lesional skin does not support a pathogenesis of atopic eczema. The upregulation of K6, K16, and K17 and reduction of K9 and K14 might contribute to the underlying pathogenesis. Unfortunately, comparison with hand eczema studies is not possible yet, because similar protein expression studies are lacking.

Topics: Adult; Biomarkers; Dermatitis, Atopic; Female; Filaggrin Proteins; Humans; Hyperkeratosis, Epidermolytic; Immunohistochemistry; Keratins; Male; Up-Regulation

Skin barrier dysfunction has been reported in both atopic dermatitis (AD) and food allergy (FA). However, only one-third of patients with AD have FA. The purpose of this study was to use a minimally invasive skin tape strip sampling method and a multiomics approach to determine whether children with AD and FA (AD

Topics: Adolescent; Area Under Curve; Child; Child, Preschool; Dendritic Cells; Dermatitis, Atopic; Epidermis; Filaggrin Proteins; Food Hypersensitivity; Humans; Intermediate Filament Proteins; Keratins; Lipids; Microbiota; Skin; Surgical Tape; Transcriptome; Water Loss, Insensible

Atopic dermatitis (AD) is a common inflammatory skin disorder, characterized by skin barrier defects and enhanced allergen priming. Null mutations in the filaggrin gene (FLG) are strongly associated with moderate to severe AD, but the pathways linking barrier dysfunction and cutaneous inflammation are still largely unknown.. To assess alteration of endogenous cysteine protease activity in FLG-deficient keratinocytes, and to determine whether the alteration in cysteine protease activity affects epidermal barrier function and associated gene and protein expression.. We established a stable FLG knockdown cell line, and reconstructed epidermal equivalents in vitro. Barrier function of the reconstructed epidermis, the barrier-associated genes and proteins, and the activity of endogenous cysteine proteases were tested. Inhibitors of cysteine proteases were used to further evaluate the role of endogenous cysteine proteases in epidermal barrier function.. FLG knockdown induced impaired epidermal barrier function. Microarray, western blotting and fluorescence staining showed reduced expression of K10, ZO-1, E-cadherin, claudin-1 and occludin in FLG knockdown keratinocytes. Compared with cysteine protease activity in control cells, protease activity was dramatically enhanced in FLG knockdown keratinocytes. Furthermore, administration of cysteine protease inhibitors significantly recovered expression of K10 and tight junction proteins, and the barrier defect induced by FLG deficiency.. This is the first observation of elevated endogenous cysteine protease activity in FLG-deficient keratinocytes, which may play an important role in impaired barrier function in AD skin. Modulation of cysteine protease activity might be a novel therapeutic approach for AD treatment.

Topics: Cell Line; Cell Proliferation; Cysteine Proteases; Dermatitis, Atopic; Filaggrin Proteins; Humans; Intermediate Filament Proteins; Keratinocytes; Keratins; Skin Absorption

Deimination (also known as citrullination), the conversion of arginine in a protein to citrulline, is catalyzed by a family of enzymes called peptidylarginine deiminases (PADs). Three PADs are expressed in the epidermis, one of their targets being filaggrin. Filaggrin plays a central role in atopic dermatitis and is a key protein for the epidermal barrier. It aggregates keratins and is cross-linked to cornified envelopes. Following its deimination, it is totally degraded to release free amino acids, contributing to the natural moisturizing factor (NMF). The mechanisms controlling this multistep catabolism in human are unknown.. To test whether external humidity plays a role, and investigate the molecular mechanisms involved.. Specimens of reconstructed human epidermis (RHEs) produced in humid or dry conditions (>95% or 30-50% relative humidity) were compared.. RHEs produced in the dry condition presented structural changes, including a thicker stratum corneum and a larger amount of keratohyalin granules. The transepidermal water loss and the stratum corneum pH were decreased whereas the quantity of NMF was greater. This highly suggested that filaggrin proteolysis was up-regulated. The expression/activity of the proteases involved in filaggrin breakdown did not increase while PAD1 expression and the deimination rate of proteins, including filaggrin, were drastically enhanced. Partial inhibition of PADs with Cl-amidine reversed the effect of dryness on filaggrin breakdown.. These results demonstrate the importance of external humidity in the control of human filaggrin metabolism, and suggest that deimination plays a major role in this regulation.

Topics: Adult; Arginine; Cell Differentiation; Citrulline; Climate; Cross-Linking Reagents; Dermatitis, Atopic; Epidermis; Female; Filaggrin Proteins; Fluorescent Dyes; Humans; Humidity; Hydrogen-Ion Concentration; Hydrolases; Intermediate Filament Proteins; Keratinocytes; Keratins; Microscopy, Electron, Transmission; Middle Aged; Protein-Arginine Deiminases; Skin; Transglutaminases

Topics: Animals; Blister; Cytokines; Cytoskeletal Proteins; Dermatitis, Atopic; Humans; Keratin-14; Keratin-5; Keratins; Mice; Mutation; Skin; Thymic Stromal Lymphopoietin

In this study, we found an effective and novel therapeutic approach to atopic dermatitis (AD) therapy via treatment with a canine adipose tissue stem cell (cATSC) extract. We determined that the therapeutic application of cATSC-derived interleukin 10 (IL10) and transforming growth factor β1 (TGFβ1) effectively modulated the overloaded immune response after the induction of AD. In addition, we investigated the molecular role of the cATSC extract during AD treatment. Dogs with naturally occurring AD that was treated at Seoul National University Veterinary Teaching Hospital was enrolled in this study. Owner consent was obtained for privately owned dogs before enrollment. We prepared a primary fat-derived cATSC extract that contained various functional factors, including IL10 and TGFβ1, as a treatment for AD. We found that the cATSC extract significantly ameliorated the pathological symptoms of canine AD. The cATSC extract secreted the immunomodulatory cytokines IL10 and TGFβ1, which modulated the overloaded immune response after the induction of AD. Moreover, these immunomodulatory cytokines modulated AD-induced inflammation and inactivated the pathological signals IL6, INFγ, iNOS, eNOS and Nox4. Additionally, these cytokines protected against apoptotic keratinocyte degeneration. This study demonstrated the novel therapeutic efficacy of the cATSC extract during successive AD treatments, which suggests a potential therapeutic use for human AD patients.

Topics: Adipose Tissue; Animals; Apoptosis; Dermatitis, Atopic; Dogs; Immunomodulation; Interleukin-10; JNK Mitogen-Activated Protein Kinases; Keratinocytes; Keratins; p38 Mitogen-Activated Protein Kinases; Reactive Oxygen Species; Signal Transduction; Stem Cells; T-Lymphocytes; Transforming Growth Factor beta1

Human skin mainly functions as an effective barrier against unwanted environmental influences. The barrier function strongly relies on the outermost layer of the skin, the stratum corneum (SC), which is composed of corneocytes embedded in an extracellular lipid matrix. The importance of a proper barrier function is shown in various skin disorders such as atopic dermatitis (AD), a complex human skin disorder strongly associated with filaggrin (FLG) null mutations, but their role in barrier function is yet unclear. To study the role of FLG in SC barrier properties in terms of SC lipid organization and lipid composition, we generated an N/TERT-based 3D-skin equivalent (NSE) after knock-down of FLG with shRNA. In these NSEs, we examined epidermal morphogenesis by evaluating the expression of differentiation markers keratin 10, FLG, loricrin and the proliferation marker ki67. Furthermore, the SC was extensively analysed for lipid organization, lipid composition and SC permeability. Our results demonstrate that FLG knock-down (FLG-KD) did not affect epidermal morphogenesis, SC lipid organization, lipid composition and SC permeability for a lipophilic compound in NSEs. Therefore, our findings indicate that FLG-KD alone does not necessarily affect the functionality of a proper barrier function.

Topics: Cell Proliferation; Dermatitis, Atopic; Epidermis; Fibroblasts; Filaggrin Proteins; Gene Knockdown Techniques; Heterozygote; Humans; Inflammation; Intermediate Filament Proteins; Keratin-10; Keratins; Ki-67 Antigen; Lipids; Membrane Proteins; Permeability; Phenotype; Skin; Skin Diseases

Combination therapy is often used in the treatment of atopic dermatitis (AD) to improve clinical efficacy or to spare the dose of each drug. Cyclosporine A (CsA) is a calcineurin inhibitor that was developed for the treatment of AD. Glucosamine (Glu) is a potent immunosuppressant that inhibits Th2-mediated immunity. We previously reported that Glu has an ameliorative effect on the development of the pathology in NC/Nga mice. The aims of our study were to investigate the therapeutic efficacy of combination of Glu and low-dose CsA in dermatophagoides farina (Df)-induced AD-like skin lesions in NC/Nga mice and to determine the underlying therapeutic mechanisms. The Df-induced NC/Nga mice with a clinical score of 7 were used for treatment with Glu (500mg/kg) alone, low-dose CsA (2, 5, and 10mg/kg) or in combination. The clinical scores were reduced significantly by the combination treatment with Glu and low-dose CsA. The suppression of dermatitis by combined therapy was accompanied by decrease in the plasma level of IgE and in the splenic level of IL-4, IL-5, IL-13, TARC and eotaxin. Histological analysis of the skin also revealed that combination treatment significantly reduced the inflammatory cellular infiltrate, including mast cells and eosinophils. Particularly, immunological evaluation reveals an increase of CD4(+)CD25(+) Treg cells in the combined treatment. The induction of TSLP, which leads to systemic Th2 response, was reduced in the skin on combination treatment. The protein expression of filaggrin and involucrin was recovered by combination treatment in the skin lesions, whereas the protein expression of keratin-10 and keratin-14 decreased in the combination treatment. Collectively, our findings suggest that combination treatment of Glu and low-dose CsA leads to the therapeutic effects in Df-induced AD-like skin lesion in NC/Nga mice through inhibition of IgE, inflammatory cellular infiltrate, and recovery of skin barrier function via a mechanism that may inhibition of Th2-mediated immune responses, in part, increment of CD4(+)CD25(+) Treg cells. These results suggest that this combined immunosuppressive treatment may provide important implications for the design of therapeutic strategies aimed at AD treatment.

Topics: Animals; Antigens, Dermatophagoides; CD4 Antigens; Cells, Cultured; Cyclosporine; Cytokines; Dermatitis, Atopic; Dermatophagoides farinae; Drug Dosage Calculations; Drug Therapy, Combination; Eosinophils; Filaggrin Proteins; Glucosamine; Humans; Immunoglobulin E; Immunosuppressive Agents; Interleukin-2 Receptor alpha Subunit; Intermediate Filament Proteins; Keratins; Male; Mast Cells; Mice; Mice, Inbred Strains; Skin; T-Lymphocytes, Regulatory; Th1-Th2 Balance

Canine atopic dermatitis (CAD) is a common allergic skin disease in dogs, associated with a defective epidermal barrier. In this study we investigated the alterations in skin keratinocyte proliferation and differentiation in CAD by quantitative reverse transcription-polymerase chain reaction. Gene expression of keratin (KRT) markers of proliferative and differentiated keratinocytes, together with that of cornified envelope proteins, involucrin (IVL) and filaggrin (FLG), were evaluated. An upregulation of KRT5 and KRT17 in both lesional and non-lesional AD skin was observed (p<0.05) whereas KRT2e, KRT14, IVL and FLG expression were significantly increased only in lesional AD skin (p<0.05). Additionally, the expression levels of KRT5, KRT14, KRT17 and IVL in CAD were strongly correlated. In conclusion, the expression of the majority of the studied keratins, as well as IVL and FLG is increased in CAD with close correlation between the proliferative keratins. This is the first report of a correlation of KRT and IVL genes with CAD.

Topics: Animals; Dermatitis, Atopic; Dog Diseases; Dogs; Filaggrin Proteins; Gene Expression Regulation; Intermediate Filament Proteins; Keratins; Protein Precursors; Skin; Transcriptome

Discovery of the significant impact of filaggrin (FLG) mutations on the genetic predisposition to atopic dermatitis (AD) focused attention on the 1q21 locus, where not only FLG but also other epidermal genes are located. In the present study, we compared 1q21 gene expression in lesional versus nonlesional AD skin.. A real-time quantitative PCR analysis of 10 1q21 genes, selected on the basis of a previous microarray study, was performed in skin biopsies from 33 individuals with AD. Three alternative pathway keratins were also evaluated.. In chronic AD skin lesions, we observed an increase in RNA encoding involucrin, S100 calcium-binding proteins A2 and A7-A9 and small proline-rich region (SPRR) proteins 1A and 2C, with fold changes ranging from 2.0 for S100A2 to 15.4 for S100A8 (p < 0.001, Bonferroni corrected), in parallel to the overexpression of the alternative pathway keratins 6A, 6B and 16. The loricrin (LOR) expression level was significantly decreased in lesional AD skin (fold change 0.5; p < 0.01). The expression of the majority of 1q21 genes and alternative keratins was closely correlated; however, for SPRR1A (and SPRR2C) in lesional skin, the correlation with other genes was lost.. We hypothesize that the deregulated increase in SPRR1A expression in chronic atopic skin lesions reflects an insufficient rise in SPRR transcripts, unable to compensate for the lack of LOR and thus contributing to the persistence of chronic AD skin lesions. Turning off the stress response in the skin may be regarded as a goal in the treatment of AD skin lesions, and SPRR genes might be targets for such an approach.

Topics: Adult; Chronic Disease; Cornified Envelope Proline-Rich Proteins; Dermatitis, Atopic; Female; Filaggrin Proteins; Gene Expression Regulation; Humans; Immunoglobulin E; Intermediate Filament Proteins; Keratins; Male; Membrane Proteins; Middle Aged; Protein Precursors; S100 Proteins; Skin

Hematoxylin-stainability of keratohyalin granules (KHG) using biochemical and immunohistochemical techniques is due to the presence of a fibrinogen γ-chain protein. A protein with a molecular weight of 100 kDa was stained with anti-Ted-H-1 monoclonal antibody and hematoxylin solution (hematoxylin-stainable protein). Since the amino acid sequence of the hematoxylin-stainable protein was to that of fibrinogen γ-chain protein, a peptide was synthesized and an antibody against the peptide was produced. This antibody reacted with the hematoxylin-stainable protein and fibrinogen γ-chain protein on immunoblot analysis and with KHG on immunohistochemical examination. Furthermore, a commercial anti-fibrinogen γ-chain protein antibody (Ab) also reacted with the hematoxylin-stainable protein as well as fibrinogen. In contrast, anti-fibrinogen β-chain protein Ab did not react with the hematoxylin-stainable protein. The fibrinogen γ-chain protein also stained with hematoxylin. These findings suggested that fibrinogen γ-chain protein may be a novel component protein of KHG and may induce the hematoxylin-stainability of KHG.

Topics: Antibodies, Monoclonal; Biomarkers; Cytoplasmic Granules; Dermatitis, Atopic; Dimerization; Fibrinogen; Fluorescent Antibody Technique, Indirect; Hematoxylin; Humans; Keratins; Peptide Fragments; Protein Binding; Sequence Analysis, Protein; Skin

Epidermal keratinocytes have the potential to produce inflammatory mediators that are considered to play an important role in skin diseases such as atopic dermatitis (AD). Thus, cell lines of canine epidermal keratinocytes are useful for studying the biological reactivity of keratinocytes in vitro. However, there has been no report on properly analyzing the phenotype of canine keratinocyte cell lines. In this work, we performed phenotypic analysis of CPEK, which was derived from the epidermis of an adult dog in order to examine the phenotypic similarity with epidermal keratinocytes. The present findings indicated that CPEK cells expressed markers for epidermal keratinocytes including cytokeratin 14, alpha6 integrin and PCNA. Our findings demonstrated that CPEK could be a useful cell line for investigating the central role of epidermal keratinocytes in the pathogenesis of AD in vitro.

Topics: Animals; Cell Line; Dermatitis, Atopic; Dog Diseases; Dogs; Epidermal Cells; Epidermis; Flow Cytometry; Immunohistochemistry; Integrin alpha6; Keratinocytes; Keratins; Phenotype

Lens epithelium-derived growth factor/dense fine speckles 70 kDa protein (LEDGF/DFS70) is a transcriptional cofactor, a transcriptional activator, survival factor, and HIV-1 transporter. It is also a major autoantigen in patients with atopic dermatitis (AD), because autoantibodies to this protein are found in approximately 30% of AD patients. To better understand the role of autoantibodies and autoantigens in the pathogenesis of AD, we examined the distribution of LEDGF/DFS70 in the epidermis of normal human skin by light and electron microscopic immunocytochemistry. Increased amounts of LEDGF/DFS70 were located in the nuclei of cells in the basal layer, whereas the cytoplasm of cells in the granular layer stained for LEDGF/DFS70 by light microscopy. Using immunoelectron microscopy, we observed the accumulation of LEDGF/DFS70 in keratohyalin granules (KGs) in the cytoplasm of cells in the granular layer. In addition, Ig heavy chain-binding protein/glucose-regulated protein, 78-kDa (Bip/GRP78), a stress sensing protein in the endoplasmic reticulum, colocalized with LEDGF/DFS70 in the KGs. These results suggest that LEDGF/DFS70 is predominantly located in the nucleus of the basal epidermal cells and translocates into the cytoplasm during differentiation. Once in the cytoplasm, LEDGF/DFS70 accumulates in the KGs in the granular layer. Finally, LEDGF/DFS70, a "nuclear" autoantigen in AD, may play a functional role in the KGs.

Topics: Adaptor Proteins, Signal Transducing; Adult; Aged; Antibody Specificity; Autoantigens; Dermatitis, Atopic; Endoplasmic Reticulum Chaperone BiP; Epidermis; Humans; Keratins; Male; Protein Transport; Transcription Factors

During pregnancy, maternal cells may enter the fetal circulation and persist until adulthood. The fate of these cells remains unknown. As unexplained T-cell-mediated conditions such as pityriasis lichenoides (PL) may occur in children, we aimed at identifying maternal cells in lesional skin of PL and controls. Archived skin biopsy specimens from young males with PL, atopic dermatitis, or normal skin were scanned for the presence of female (presumably maternal) cells using fluorescence in situ hybridization (FISH) with X and Y chromosome-specific probes. Phenotyping of maternal cells relied on FISH combined with anti-CD45, anti-CD1a, or anti-cytokeratin labelling, identifying leukocytes, Langerhans cells, and keratinocytes, respectively. Maternal cells were found in PL (11/12) and controls (4/7), but their average frequency was higher in PL: 99 per million cells as compared to 5 per million cells in controls (P = 0.005). In the epidermis, the maternal microchimeric cells were labelled by anti-cytokeratin in all cases. We identified maternally derived keratinocytes in the skin of male children with inflammatory skin disorders. These cells may either help repair the damaged skin or home initially in the skin and trigger a host (child) versus graft (mother) disease.

Topics: Antigens, CD1; Child; Child, Preschool; Chimera; Chimerism; Dermatitis, Atopic; Female; Humans; In Situ Hybridization, Fluorescence; Keratinocytes; Keratins; Leukocyte Common Antigens; Male; Maternal-Fetal Exchange; Pityriasis Lichenoides; Pregnancy

The keratin-14 IL-4 transgenic (Tg) mouse model of atopic dermatitis (AD) is characterized by skin infiltration of T cells, early up-regulation of T(h)2 cytokines and late surge of T(h)1 cytokines. In the present study, we investigated the role of CCL27, a T cell skin-homing chemokine known to be elevated in sera of human AD patients, in disease development in our animal model of AD. The results showed that the mRNA and protein levels of CCL27 in the skin and serum were significantly increased in IL-4 Tg mice. The percentage of T cells expressing CCR10 in skin draining lymph nodes of IL-4 Tg mice was increased, consistent with the findings of >80% of skin-infiltrating T cells in Tg mice expressing CCR10. Chemotaxis transmigration assay demonstrated that CCL27 promotes a greater degree of migration of T cells in diseased Tg mice. Subcutaneous injection of neutralizing anti-CCL27 to IL-4 Tg mice with early skin lesions resulted in reduced clinical progression of inflammation, accompanied with decreased T cell and mast cell infiltration in the skin, and down-regulation of inflammatory cytokines. In conclusion, CCL27 and CCR10 interaction is important for the development of skin inflammation in our AD model.

Topics: Animals; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Movement; Chemokine CCL27; Chemokines, CC; Dermatitis, Atopic; Interleukin-4; Keratin-14; Keratinocytes; Keratins; Lymphocyte Activation; Mice; Mice, Transgenic; Receptors, CCR10; Receptors, Chemokine; Skin; Up-Regulation

Atopic dermatitis (AD)-specific genes have not yet been clarified. Objectives To identify gene expression specific to active atopic skin lesions.. We analysed 23,000 genes in skin biopsy samples from 17 patients with AD and four normal controls using Affymetrix oligonucleotide arrays.. Four of the 10 genes with the greatest differences in expression between patients and controls, S100A8 and S100A7 (upregulated), and loricrin and filaggrin (downregulated), were epidermal differentiation genes located on 1q21, a locus previously reported to have a genetic linkage with AD.. Our results, showing downregulation of the cornified envelope genes and upregulation of the alternative keratinization pathway, are the first to suggest abnormal epidermal differentiation and defective defences as key abnormalities in AD.

Topics: Adult; Cell Differentiation; Chromosomes, Human, Pair 1; Dermatitis, Atopic; Epidermis; Female; Filaggrin Proteins; Gene Expression Profiling; Gene Expression Regulation; Genetic Predisposition to Disease; Humans; Keratinocytes; Keratins; Male; Multigene Family; Oligonucleotide Array Sequence Analysis

Angiogenesis plays an important role in psoriasis, but its role in atopic dermatitis is unknown. The authors examined the dermal microvasculature of an IL-4 transgenic mouse model of atopic dermatitis to determine whether angiogenesis was present.. Transmission and scanning electron microscopy and confocal microscopy studies were performed.. Transmission electron microscopy showed sprouting, transcapillary pillars of intussusception, thickened endothelial cells with large nuclei, and increased interendothelial junctional cleft number and length. Compared to nontransgenic littermates, there was a significant increase in the lengths and numbers of the interendothelial junctional clefts, along with a decrease in the length ratios of tight junction to interendothelial junctional clefts in both the early and late disease stages. In the early and late skin lesions, scanning electron microscopy of vascular corrosion casts showed disorganization of the capillary network hierarchy with increased density of capillary sprouts. Confocal microscopy of the animals with early and late skin lesions showed significant reduction in tight junction protein claudin-5.. Angiogenesis is the major pathologic feature in this model of atopic dermatitis. The chronic skin inflammation is intertwined with and may cause the angiogenesis, but the angiogenesis itself is likely to be important in this disease process.

Topics: Animals; Dermatitis, Atopic; Dermis; Disease Models, Animal; Interleukin-4; Keratin-14; Keratins; Mice; Mice, Transgenic; Microscopy, Confocal; Microscopy, Electron; Neovascularization, Pathologic

We have previously characterized the keratin 14 interleukin-4-transgenic (IL-4-Tg) mouse model of atopic dermatitis as a chronic pruritic inflammatory skin disease typified by skin infiltration of inflammatory cells and early up-regulation of Th2 cytokines and late surge of Th1 cytokines. In the present study, we examined the involvement of B cells. Systematic examinations of the following immunological parameters on B cells were carried out in non-Tg control mice and in IL-4-Tg mice at before disease onset and early and late disease stages so that we could determine the immunological sequence of events leading to the disease development: surface expressions of IA/IE, activation and costimulatory molecules, proliferation under LPS or IgM stimulation, quantification of cell surface and serum IgE, IgG1, and IgG2a. Our results showed that as the disease progresses from before onset to early disease and to late disease, there is a parallel increase in surface markers of B cell activation (IA/IE, CD44, CD69, CD80 and CD86), in B cell proliferation, and in cell surface and serum IgE. Significant increases of Th2-driven serum IgG1 and IgE in early disease was followed by significant increase of Th1-driven IgG2a in late disease. Importantly the significant increases of activation molecule (IA/IE), proliferation (to LPS), and surface IgE on B cells of the IL-4-Tg mice precedes the up-regulation of serum IgE and disease onset. These data suggest that activated B cells may play a role in atopic dermatitis disease development by up-regulating serum IgE concentration, which serves as a marker of disease onset.

Topics: Animals; B-Lymphocytes; Biomarkers; Cell Division; Dermatitis, Atopic; Disease Models, Animal; Disease Progression; Immunoglobulin E; Immunoglobulin G; Interleukin-4; Keratin-14; Keratinocytes; Keratins; Lymph Nodes; Lymphocyte Activation; Lymphocyte Count; Mice; Mice, Transgenic; RNA, Messenger; Spleen; Up-Regulation

A defective permeability barrier leads to the penetration of environmental allergens into the skin and initiates immunological reactions and inflammation crucially involved in the pathogenesis of atopic dermatitis (AD). Decreased stratum corneum ceramide content may cause the defect in permeability barrier function consistently found in AD. Acid and neutral sphingomyelinase (A- and N-SMase) generate ceramides with structural and signal transduction functions in epidermal proliferation and differentiation. We determined epidermal SMase activities, DNA synthesis, involucrin, loricrin, filaggrin, and keratin expression in lesional and non-lesional skin of AD patients. We found decreased epidermal A-SMase activity in lesional and non-lesional skin, correlating with reduced stratum corneum ceramide content and disturbed barrier function. N-SMase activity was reduced in non-lesional skin and more significantly reduced in lesional skin, correlating with impaired expression of cornified envelope proteins and keratins, important for skin barrier function. Changes in involucrin, loricrin, filaggrin, keratin K 5 (basal) and K 16 (proliferation associated) were noticed in non-lesional and lesional skin, whereas changes in K 10 (suprabasal), K 6 (proliferation associated), and K 17 (inflammation associated) were found only in lesional skin. In summary, reduction in SMase-generating ceramides and impaired differentiation are involved in the defective barrier function found in AD.

Topics: Biomarkers; Blotting, Western; Cell Differentiation; Cell Division; Ceramides; Dermatitis, Atopic; Epidermis; Female; Filaggrin Proteins; Humans; Intermediate Filament Proteins; Keratins; Male; Membrane Proteins; Protein Precursors; Signal Transduction; Sphingomyelin Phosphodiesterase; Substrate Specificity; Water

MAIL (molecule-possessing ankyrin repeats induced by lipopolysaccharide) is a nuclear IkappaB protein that is also termed interleukin-1-inducible nuclear ankyrin repeat protein or inhibitor of nuclear factor kappaB (IkappaB) zeta. In this study, we generated Mail-/- mice to investigate the roles of MAIL in whole organisms. Mail-/- mice grew normally until 4-8 weeks after birth, when they began to develop lesions in the skin of the periocular region, face, and neck. MAIL mRNA and protein were constitutively expressed in the skin of wild type controls, especially in the keratinocytes. Serum IgE was higher in Mail-/- mice than in normal. Histopathological analysis indicated that the Mail-/- skin lesions appeared to be atopic dermatitis (AD) eczema with inflammatory cell infiltration. In addition, markedly elevated expression of some chemokines such as thymus and activation-regulated chemokine was detected in the Mail-/- skin lesions, similar to that observed in the skin of patients with AD. In Mail-/- mice, MAIL-deficient keratinocytes might be activated to produce chemokines and induce intraepidermal filtration of inflammatory cells, resulting in the onset of the AD-like disease. These findings suggest that MAIL is an essential molecule for homeostatic regulation of skin immunity. The Mail-/- mouse is a valuable new animal model for research on AD.

Topics: Adaptor Proteins, Signal Transducing; Alleles; Animals; Ankyrins; Chemokines; Dermatitis, Atopic; Disease Models, Animal; Genetic Vectors; Genome; I-kappa B Proteins; Immunoglobulin E; Immunohistochemistry; Inflammation; Islets of Langerhans; Keratinocytes; Keratins; Mice; Mice, Inbred C57BL; Mice, Transgenic; Models, Genetic; Nuclear Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transgenes

Atopic dermatitis (AD) is a pruritic inflammatory skin disease. Because IL-18 directly stimulates T cells and mast cells to release AD-associated molecules, Th2 cytokines, and histamine, we investigated the capacity of IL-18 to induce AD-like inflammatory skin disease by analyzing KIL-18Tg and KCASP1Tg, which skin-specifically overexpress IL-18 and caspase-1, respectively. They spontaneously developed relapsing dermatitis with mastocytosis and Th2 cytokine accumulation accompanied by systemic elevation of IgE and histamine. Stat6-deficient KCASP1Tg displayed undetectable levels of IgE but manifested the same degree of cutaneous changes, whereas IL-18-deficient KCASP1Tg evaded the dermatitis, suggesting that IL-18 causes the skin changes in the absence of IgE/stat6. KIL-18Tg and IL-1-deficient KCASP1Tg took longer to display the lesion than KCASP1Tg. Thus, AD-like inflammation is initiated by overrelease of IL-18 and accelerated by IL-1. Our present study might provide insight into understanding the pathogenesis of and establishing therapeutics for chronic inflammatory skin diseases including AD.

Topics: Animals; Dermatitis, Atopic; Humans; Immunoglobulin E; Inflammation; Interleukin-18; Keratins; Mice; Mice, Transgenic; Promoter Regions, Genetic; Pruritus; Rabbits; Signal Transduction; Specific Pathogen-Free Organisms; STAT6 Transcription Factor; T-Lymphocytes; Th2 Cells; Trans-Activators

Topics: Dermatitis, Atopic; Humans; Intermediate Filaments; Keratins; Skin Diseases; Skin Physiological Phenomena

Mice with a targeted disruption of the Rel / NF-kappaB family member RelB develop a complex inflammatory phenotype and hematopoietic abnormalities. RelB-deficient (relB(- / -)) mice were clinically normal until 4 - 10 weeks after birth when thickening of the skin and hair loss developed. Histological and immunohistochemical evaluation of relB(- / -) skin lesions revealed hyperkeratosis and marked epidermal hyperplasia. Many CD4(+) T cells and eosinophils mixed with lesser numbers of CD8(+) T cells and neutrophils were present in the dermis. There was a moderate increase of MHC class II-positive dermal dendritic cells and dermal mast cells. Increased expression of Th2 cytokines correlated with increased mRNA levels of eotaxin and CCR3 in relB(- / -) skin. The dermatitis did not develop in the offspring of relB(- / -) mice crossed with transgenic mice that lack peripheral T cells, demonstrating that the skin lesions were T cell dependent. The dermatitis observed in RelB-deficient mice had many similarities with atopic dermatitis in human patients including infiltrating CD4(+) T cells and eosinophils in the skin, increased number of eosinophils in the blood and increased serum IgE. Thus, the relB(- / -) mouse should be a useful model to study the pathogenesis of this common allergic human disease.

Topics: Animals; Chemokines; Cytokines; Dermatitis, Atopic; Histocompatibility Antigens Class II; Humans; Keratins; Mast Cells; Mice; Mice, Inbred C57BL; Neutrophils; Proto-Oncogene Proteins; Skin; T-Lymphocytes; Transcription Factor RelB; Transcription Factors

Keratin K17, the myoepithelial keratin, is expressed in psoriasis but is not present in healthy skin. Psoriasis is associated with production of gamma interferon (IFN gamma), which induces the expression of keratin K17 by activating transcription factor STAT1. Our hypothesis states that the induction of K17 is specific for the inflammatory reactions associated with high levels of IFN gamma and activation of STAT1. One of the corollaries of the hypothesis is that the STAT1-activating cytokines should induce the expression of keratin K17, whereas those cytokines that work through other mechanisms should not. Furthermore, because the STAT activation pathway is dependent upon protein phosphorylation events, phosphorylation inhibitors should attenuate the induction of keratin K17, whereas protein phosphatase inhibitors should augment it. To test this hypothesis, we analyzed lesional samples of inflammatory diseases using immunofluorescence, transfected keratinocytes with K17 gene promoter DNAs in the presence of various cytokines, and followed nuclear translocation of STAT1 in keratinocytes using specific antibodies. Confirming the hypothesis, we found that K17 is induced in psoriasis and dermatitis caused by delayed type hypersensitivity, which are associated with high levels of IFN gamma, but not in samples of atopic dermatitis, which is not. Two cytokines, interleukin-6 and leukemia inhibitory factor, which can induce phosphorylation of STAT1, can also induce K17 expression, whereas interleukin-3, interleukin-4, interleukin-10, and granulocyte macrophage colony stimulating factor have no effect on K17 expression. As expected, staurosporine and genistein inhibited, whereas okadaic acid augmented, the induction of K17 by IFN gamma. Our data indicate that in inflammatory skin diseases, lymphocytes, through the cytokines they produce, differently regulate not only each other, but also keratin gene expression in epidermis one of their target tissues.

Topics: Cytokines; Dermatitis; Dermatitis, Atopic; DNA-Binding Proteins; Enzyme Inhibitors; Epidermis; Growth Inhibitors; Humans; Interferon-gamma; Interleukin-10; Interleukin-4; Interleukin-6; Keratins; Leukemia Inhibitory Factor; Lymphokines; Phosphoprotein Phosphatases; Promoter Regions, Genetic; Protein Kinase Inhibitors; Psoriasis; STAT1 Transcription Factor; Trans-Activators; Transcription, Genetic

Keratinocyte expression of the monocyte/macrophage surface antigens defined by OKM1 and OKM5 antibodies (Ortho Diagnostics) was examined using the peroxidase anti-peroxidase immunohistochemical technique. A range of inflammatory cutaneous disorders were investigated, including lichen planus, psoriasis and atopic dermatitis. Positive suprabasal keratinocyte expression of OKM5 antigen was observed in all disorders, while keratinocyte staining with OKMI antibody was consistently negative. These results provide further evidence that keratinocytes may play an important role in cutaneous immune responses. Furthermore, they are consistent with the recent observation that HLA-DR positive keratinocytes may modulate cutaneous immunological reactions by inducing T-cell unresponsiveness.

Topics: Adult; Antigens, Differentiation; CD36 Antigens; Dermatitis, Atopic; Epidermis; Humans; Keratins; Lichen Planus; Lupus Erythematosus, Discoid; Lymphoma; Monocytes; Psoriasis; Skin Diseases; Skin Neoplasms

In 30% to 40% of cases atopic dermatitis (AD) is believed to be associated with autosomal dominant ichthyosis vulgaris (ADI). The diagnosis of ADI can be proved by the ultrastructural demonstration of a defective keratohyalin (KH) synthesis, resulting in minute granules of crumbly appearance in only one layer of granular cells. To investigate the suggested frequent association of ADI with AD, ultrastructural examination of dry skin of 49 AD patients was performed. Only in 2 patients abnormal KH was demonstrated by electron microscopy. 17 patients, including the 2 patients with abnormal KH, showed hyperlinear palms. The present study shows that hyperlinear palms and dry skin are in most cases a phenotypic marker of AD alone and not a sign of concomitant ADI. A histologically one-layered or absent stratum granulosum may occur in the dry skin of patients with only AD and does not indicate a manifestation of concomitant ADI in all cases.

Topics: Adolescent; Adult; Dermatitis, Atopic; Female; Foot Dermatoses; Hand Dermatoses; Humans; Hyalin; Ichthyosis; Keratins; Male; Skin; Skin Diseases

Epidermal patterns of class II human lymphocyte antigen (HLA) expression in atopic and allergic contact dermatitis have been compared, using monoclonal antibodies recognizing each subregion. Expression of class II human lymphocyte antigens on keratinocytes has been confirmed in allergic contact dermatitis, while we have found them to be absent in atopic dermatitis. This finding argues that cell-mediated immune responses, possibly to epicutaneous contact with allergen, are not implicated in the pathogenesis of atopic dermatitis.

Topics: Adult; Antibodies, Monoclonal; Dermatitis, Atopic; Dermatitis, Contact; Epidermal Cells; HLA-D Antigens; Humans; Keratins

Topics: Adolescent; Adult; Dermatitis, Atopic; Diagnosis, Differential; Epidermis; Genes, Dominant; Humans; Ichthyosis; Keratins

In recent years much interest has been focused on the functions of the membrane-coating granules (MCGs). These granules seem to play an essential role in the formation of the barrier of the stratum corneum by extruding their lipid-rich content into the extracellular space of the corneocytes. The dry non-eczematous skin in atopic dermatitis has been reported to have defective barrier function. In the present study a quantitative electron microscopic analysis was made of the volume of MCGs in the transition zone between the stratum granulosum and stratum corneum in dry skin of patients with atopic dermatitis. The relative volume of MCGs was significantly greater than that in normal skin. This finding may indicate a disturbance of the "maturation" of the MCGs, leading to a defect in the barrier function in atopic dermatitis.

Topics: Adult; Dermatitis, Atopic; Epidermal Cells; Epidermis; Humans; Keratins; Microscopy, Electron

Staphylococcus aureus has a peculiar ability to colonize the skin of patients with atopic dermatitis. We examined the possibility that this might be due to a specific ability of this pathogenic staphylococcus to adhere to atopic stratum corneum. We used an in vitro model to show that S aureus does have an unusual ability to adhere to atopic corneocytes when compared with corneocytes obtained from patients with other cutaneous diseases, including psoriasis. Protein A--a component of the staphylococcal cell wall--may be responsible in part for this adherence phenomenon. This trait did not extend to the other gram-positive bacteria tested.

Topics: Cell Division; Dermatitis, Atopic; Epidermal Cells; Epidermis; Humans; Hydrogen-Ion Concentration; Keratins; Lipopolysaccharides; Phosphatidic Acids; Skin; Staphylococcal Protein A; Staphylococcus aureus; Teichoic Acids; Temperature

The effect of cellular pathology and keratinization of skin and nasal cells upon binding of Staphylococcus aureus were examined. Adherence with epithelial cells obtained from either the skin or nasal mucosa of patients with atopic dermatitis was greater than that observed with normal cells (p less than 0.001); the difference in adherence between psoriatic and normal cells was not statistically significant. Tested nasal cells were microscopically differentiated into 4 general types based on stage or layer of keratinization: spinous, low granular, high granular, and keratin. The degree of adherence was related to the progress of keratinization. Data indicated the existence of 2 types of receptors for S. aureus on nasal cells: One, present upon both granular and fully keratinized cells, is not blocked by teichoic acid and appears responsible for the higher bacterial counts on atopic cells; the second is found on keratinized cells only and is susceptible to teichoic acid.

Topics: Adhesiveness; Bacterial Physiological Phenomena; Dermatitis, Atopic; Epithelium; Humans; Keratins; Nasal Mucosa; Psoriasis; Skin; Skin Physiological Phenomena; Staphylococcus aureus

Keratin proteins were extracted from scales of normal skin, clinically active psoriatic lesions, and atopic dermatitis. Filaments prepared by in vitro assembly upon dialysis of the proteins against a low ionic strength buffer were comparatively characterized by electron microscopy, SDS gel electrophoresis, and amino acid analysis. Filaments formed using keratin obtained from the skin of normal individuals were thin and wavy, whereas those formed using keratin isolated from the scales of psoriatic patients were straight and showed a tendency to assemble side by side. Filaments of atopic dermatitis were indistinguishable from those of normal individuals. Filaments of both normal and atopic dermatitis contained the protein band of 67,000 daltons, which was absent in filaments of psoriasis. In contrast, 2 protein bands of 54,000 and 57,000 daltons were only detectable in psoriasis. Amino acid analysis of these filaments further demonstrated that filaments of psoriasis differ from those of normal individuals in that they have a glycine content that is 60% of normal.

Topics: Amino Acids; Dermatitis, Atopic; Humans; Keratins; Molecular Weight; Proteins; Psoriasis; Skin

Topics: Blister; Dermatitis, Atopic; Dermatitis, Exfoliative; Extremities; Facial Dermatoses; Foot Dermatoses; Genes, Dominant; Genes, Recessive; Hand Dermatoses; Humans; Ichthyosis; Keratins; Keratosis; Microscopy, Electron; Ribosomes; Scalp Dermatoses; Sex Chromosomes; Skin; Thorax

Topics: Catecholamines; Citric Acid Cycle; Dermatitis, Atopic; Glucose; Glycogen; Humans; Hydrocortisone; Infant; Keratins; Lysosomes; Metabolism; Oxidoreductases; Skin; Transferases

Topics: Animals; Antigen-Antibody Reactions; Caseins; Cattle; Cyanides; Dermatitis; Dermatitis, Allergic Contact; Dermatitis, Atopic; Erythema; gamma-Globulins; Gelatin; Hydrogen-Ion Concentration; Keratins; Nickel; Proteins; Research; Serum Albumin; Serum Albumin, Bovine; Sulfides; Toxicology

Topics: Alopecia; Alopecia Areata; Dermatitis; Dermatitis, Atopic; Dermatitis, Seborrheic; Dermatology; Humans; Keratins; Mercury; Nails; Ointments; Pigmentation; Psoriasis