bluuebunny/crossref_metadata_2025 huggingface dataset:
Barrier function coordinately regulates epidermal IL‐1 and IL‐1 receptor antagonist mRNA levels
Authors: Wood Ladonna C. , Feingold Kenneth R. , Sequeira‐Martin Sandy M. , Elias Peter M. , Grunfeld Carl
Published: April 2006

Abstract Disruption of the cutaneous permeability barrier increases mRNA levels for TNF, GM‐CSF, FL‐1 alpha, and IL‐1 beta in the epidermis. We have hypothesized that the cylokines mediate the changes in lipid and DNA synthesis which occur following barrier disruption. To further characterize the cytokine response to barrier abrogation, we examined the levels of epidermal IL‐Ira mRNA in two acute models and one chronic model in the hairless mouse. IL‐Ira mRNA levels increased shortly after acute disruption of the barrier with acetone, reached a peak at 3–4 h after treatment, and returned to control levels by 8h. These changes in mRNA levels parallel those which occur for IL‐1 alpha and beta. Furthermore, IL‐Ira mRNA levels were elevated 5‐fold and 4‐fold, at 2.5 h and 4 h, respectively, following tape‐stripping, a second acute model of barrier disruption. Finally, IL‐Ira mRNA levels were elevated 2.5‐fold in the epidermis of EFAD mice, which have a chronic barrier defect. Thus, the cutaneous response to barrier disruption includes mechanisms which increase IL‐I and IL‐Ira mRNA levels in a coordinate manner. The net result provides a regulatory mechanism for controlling the biological effects of increased IL‐1 production.
Trans‐urocanic acid, a natural epidermal constituent, inhibits human natural killer cell activity in vitro
Authors: Uksila Jaakko , Laihia Jarmo K. , Jansén Christer T.
Published: April 2006

Abstract UV irradiation has been reported to influence NK cell function both in vitro and in vivo. Since urocanic acid may mediate UV‐induced immune modulation we tested the effect of trans‐ and cis‐urocanic acid (UCA) on the cylotoxic activity of human peripheral blood lymphocytes against the erythroleukemic target cell line K562 in vitro. Trans‐UCA was found to be a strong inhibitor of NK cell activity whereas cis‐UCA had no effect. Trans‐UCA also partially inhibited cytotoxic function of IL‐2‐activated NK cells and reduced IL‐2‐induced activation of NK cells. This is the first report describing trans‐UCA to be active, and cis‐UCA inactive, in regulating an immune function. In the skin, a decrease in epidermal trans‐urocanic acid concentration by UV radiation could produce a favorable milieu for NK cell activity, and thus counteract the impairment of antigen‐specific immune surveillance, induced by increased cis‐urocanic acid concentrations.
Human dermatophyte‐responsive T‐cell lines recognize cross‐reactive antigens associated with mannose‐rich glycoproteins
Authors: MacCarthy Kathleen G. , Blake John S. , Johnson Karyn L. , Dahl Mark V. , Kalish Richard S.
Published: April 2006

Abstract Resistance to dermatophyte infections has been shown to be mediated in part by T lymphocytes. The dermatophyle antigens recognized by human T lymphocytes and their degree of cross‐reactivity were analyzed. Dermatophyte‐responsive T‐cell lines were generated by in vitro sensitization to crude fungal extracts obtained from Trichophyton rubrum, Trichophyton tonsurans. Microsporum canis and Epidermophyton floccosum. Proliferation was measured by incorporation of 3H‐thymidine. The human T‐cell lines responded to fungal extracts derived from these various dermatophyte species, demonstrating the recognition of cross‐reactive antigens by human T cells. However, the T cells were dermato‐phyte‐specific as they did not respond to herpes antigen, nor did herpes‐specific T cells derived from the same donors respond to dermatophyte antigens. The mannose‐rich glycoprotein fraction (mannan) isolated from T. rubrum was able to induce proliferation of T‐cell lines generated by stimulation with various fungal extracts. Furthermore, a T‐cell line generated by stimulation with mannan derived from T. rubrum proliferated in response to extracts from various fungal species, indicating that a major cross‐reactive dermatophyte T‐cell antigen was present in the mannose‐rich glycoprotein fraction.
Interleukin‐4 and interferon‐γ interactions in the induction of intercellular adhesion molecule‐1 and major histocompatibility complex class II antigen expression of normal human keratinocytes
Authors: Viac J. , Gueniche A. , Gatto H. , Lizard G. , Schmitt D.
Published: April 2006

Abstract Interleukin‐4 (IL‐4) that may be produduced by T‐helper cells in atopic lesions has immunomodulatory activities on skin cells which are poorly known. Our study was aimed at determining whether the cytokine exerts some effects on keratinocyte activation and can either enhance or antagonize interferon‐γ (IFN‐γ)‐induced ICAM‐I or HLA‐DR antigen expression. Using normal keratinocytes cultured in defined medium and cytofluorography, we showed that treatments of the human cells with the cytokine IL‐4 alone had no effect on the induction of ICAM‐1 or HLA‐DR molecules. However, a transient, but significant enhanced expression of ICAM‐I was observed by the combination of IFN‐γ and IL‐4 after 24 h of stimulation, which was followed by a reduction at 48 and 72 h. Conversely, IL‐4, when added during the IFN‐γ activation stage, had no effect on MHC class II antigen expression of keratinocytes; however, the cytokine reduced the expression of these antigens when added 24 h before the stimulation by IFN‐γ. These results suggest that IFN‐γ and IL‐4 may interact to regulate ICAM‐1 and HLA‐DR expression on keratinocytes.
Expression of ɛBP, a β‐galactoside‐binding soluble lectin, in normal and neoplastic epidermis
Authors: Konstantinov K. N. , Shames B. , Izuno G. , Liu F.‐T.
Published: April 2006

Abstract The study of animal lectins and glycoconjugates has become an important area of research in biomedical sciences, as these molecules are believed to play important roles in a variety of biological processes. This report describes a study of the expression of an animal lectin, IgE‐binding protein (ɛBP), also known as Mac‐2 and CBP35, in human skin. We have analyzed cultured human keratinocytes as well as normal human skin and a number of epidermal neoplasms, by immunoblotting. immuno‐fluorescence and immunohistochemistry. We showed that ɛBP is expressed in human keratinocytes, hair follicles, sebaceous and sweat glands. We found that cBP expression retains in various epidermal neoplasms, including basal cell carcinoma, squamous cell carcinoma and keratoacan‐thoma, although the level of expression appears to be reduced as compared to normal epidermis. The immunohistochemical analysis also suggests that the level of ɛBP expression appears to be dependent on the degree of cellular differentiation of keratinocytes.
Adhesion molecules in lesions of American cutaneous leishmaniasis
Authors: Tapia F. J. , Càceres‐Dittmar G. , Sànchez M. A. , Fernàndez C. T. , Rondòn A. J. , Convit J.
Published: April 2006

Abstract Accessory signals, which include adhesion molecules, MHC‐II molecules and cytokines. are necessary to foster the interaction between memory T cells and epidermal cells, that is required to promote cutaneous inflammatory responses. American cutaneous leishmaniasis (ACL) is characterized by a spectrum of immunological manifestations, and is a prototype disease for the study of regulatory mechanisms involved in immune protection against protozoal infection. In the present study, we show that diffuse cutaneous leishmaniasis (DCL) epidermis contains keratinocytes that do not express ICAM‐I and HLA‐DR molecules. Langerhans cells (LC) are within normal values or somewhat lower, and a very few cells expressing the HB15 molecule a new described member of the Ig superfamily are found in such lesions. Mucocutaneous leishmaniasis (MCL) epithelium shows an increased expression of ICAM‐1 and HLA‐DR molecules, few HBI5+ cells, and an absence of epithelial LC. Localized cutaneous leishmaniasis (LCL) epidermis displays ICAM‐+ keratinocytes organized in patches, a uniform expression of HLA‐DR, hyper‐plasia of LC, and numerous HBI5+ cells. In all forms of the disease, infiltrating T cells express more LFA‐1β than LFA‐1α, but LFA‐1β+ cells are more abundant in LCL granulomas. In contrast, there are more LFA‐lα+ cells in DCL and MCL than in LCL granulomas. LCL lesions also show the highest numbers of HB15+ cells within the granu‐loma. These results indicate the importance of adhesion molecules in ACL lesions, and open new possibilities for therapeutic schemes oriented towards the control of cell migration.
Topical glucocorticosteroids modulate the expression of CRABP I and II in human skin differently
Authors: Piletta P. , Jaconi S. , Siegenthaler G. , Didierjean L. , Saurat J. H.
Published: April 2006

Abstract Epidermal cells express two retinotic acid‐binding proteins (CRABP I and II). Because CRABP II protein is strongly induced by topical retinoic acid, the respective roles of the two proteins in the pharmacological activity and toxicity of topical retinoids deserve particular attention. Since topical steroids diminish the irritation induced by retinoic acid (RA), whereas retinoic acid may counteract the atrophogenic effects of steroids, the possible interplay of both compounds in the expression of CRABP I and II appeared worth studying. We have analyzed the effects of topical application of triamcinolone acetonide (TA) on the retinoic acid‐induced altered expression of CRABP I and II in normal human skin, at the protein and mRNA levels. We found that CRABP II protein and mRNA were strongly increased upon retinoic acid application: this induction was significantly inhibited by concomitant application of triam‐cinolone acetonide; a more potent steroid, dilluocortolone valerate, was also found to dimish normal endogenous expression of CRABP II. In contrast, CRABP I protein was decreased by topical retinoic acid, and the down modulating effect of retinoic acid was counteracted by triamcinolone acetonide.
Cell‐mediated immune responses to Malassezia furfur serovars A, B and C in patients with pityriasis versicolor, seborrheic dermatitis and controls
Authors: Ashbee H. R. , Ingham E. , Holland K. T. , Cunliffe W. J.
Published: April 2006

Abstract It has been postulated that patients with Malassezia furfur associated dermatoses have a deficient cell‐mediated immune response to M. furfur. This study examined the cell‐mediated immune responses to M. furfur serovars A, B and C of 10 patients with pityriasis versicolor and 10 age‐ and sex‐matched controls; and 10 patients with seborrheic dermatitis and 10 age‐ and sex‐matched controls. The responses to each serovar of M. furfur were assessed using the lymphocyte transformation assay and the leukocyte migration inhibition assay. The lymphocyte transformation responses of the patients with pityriasis versicolor to M. furfur serovars A, B and C (O/10, 6/10 and 5/10 respectively) were not significantly different from those of controls (0/10, 2/10 and 1/10). However, for patients with seborrheic dermatitis, significantly more patients lymphocytes responded to serovars B and C (6/10 and 6/10 respectively) than those of controls (1/10 and 1/10). No patient or control responded to serovar A. In the leukocyte migration inhibition assay, the leukocytes from a greater proportion of patients with pityriasis versicolor (5/7) responded to serovar B than controls (2/10); and the leukocytes from a greater proportion of patients with seborrheic dermatitis (4/10) responded to serovar C than controls (0/9). Thus, this data did not indicate the presence of any cell‐mediated immune deficiency to M. furfur in patients with pityriasis versicolor or seborrheic dermatitis, as measured by the lymphocyte transformation assay or the leukocyte migration inhibition assay. The greater responsiveness of T lymphocytes from patients may indicate that T lymphocytes might be involved in the pathogenesis of these diseases.
Progressive differentiation of human sebocytes in vitro is characterized by increasing cell size and altering antigen expression and is regulated by culture duration and retinoids
Authors: Zouboulis Christos C. , Krieter Axel , Gollnick Harald , Mischke Dietmar , Orfanos Constantin E.
Published: April 2006

Abstract Increasing cell size, lipid accumulation, and altered antigen expression are features of sebaceous differentiation in vivo. Enhanced lipid synthesis with progressive differentiation is also present in cultured human sebocytes. This study was conducted to investigate the evolution of cell size and antigen expression of human sebocyles with progressive differentiation in vitro. Subconlluent human sebocyte cultures were examined for sebocyte differentiation evaluated on cytocentrifuge preparations by light microscopy and classified in stages according to morphological criteria described for sebocytes in vivo. Rates of 5.1 ±2. 2% undifferentiated sebocytes. 29.2 ±4.9% early differentiated, 20.7 ±4.1% advanced differentiated, 37.6±6.4% fully differentiated, and 5.9± 1.9% mature sebocytes were calculated in secondary cultures. The size of cultured sebocytes measured by computer‐assisted planimetry significantly increased with progressive differentiation up to 4‐5.5 times. The low rates of mature sebocytes and the only moderate increase of their size with progressive differentiation indicate an incomplete terminal differentiation in vitro. Sebocytes were subsequently stained with a series of monoclonal antibodies (mAb) to determine antigen expression using the alkaline phosphatase anti‐alkaline phosphatase technique. The number of sebocytes labeled with the anti‐keratin mAb CK8.12 and KLI, and the mAb 34DII (82 kD protein) increased with progressive differentiation; significant differences were found after comparing early and advanced differentiated sebocytes. Sebocytes were positively stained with the anti‐keratin mAb 6BIO (K 4). RPNI162 (K 7), CK.I3 (K 13), RPNI165 (K 19). CK.8.60, and the mAb 115F5 (MAM‐6c). OM‐I (sebaceous gland antigen), and 24F10 (basic polypeptides) only at late‐stage differentiation. The expression of keratins 4, 7, 13, and 19 was confirmed by gel electrophoresis and immunoblotting. The data obtained were used to study the effects of the duration of cultivation and of the retinoids isotretinoin and tretinoin on sebocyte differentiation in vitro. Subcultivalion of sebocytes upregulaled, and treatment with isotretinoin but not with tretinoin downregulated labeling with mAb which recognize indicating progressive and late‐stage differentiation.
Predominant usage of distinct T‐cell receptor Vβ regions by epidermotropic T cells in psoriasis
Authors: Boehncke Wolf‐Henning , Küenzlen Christiane , Zollner Thomas Matthias , Mielke Volker , Sterry Wolfram
Published: April 2006

Abstract To investigate the role of T cells in psoriasis, we analyzed the T‐cell receptor (TCR) repertoire in peripheral blood lymphocytes (PBLs) and in skin lesions of 9 patients with chronic plaque‐stage psoriasis by means of immunohistochemistry. An extended panel of monoclonal antibodies detecting 11 different V(3 families was employed in this study. In the peripheral blood, no predominant TCR Vβ expression was detected and interindividual differences were small. Skin‐infiltrating T cells showed only marginal differences in the TCR Vβ usage when compared with PBLs. However, in 3 patients the intraepidermal infiltrate comprised up to 40% of T cells expressing a TCR Vβ region which was virtually absent in the peripheral blood and the dermal infiltrate. Thus, although the TCR Vβ usage of skin‐infiltrating T cells showed little difference compared to PBLs, several cases exhibited a significant increase of epider‐motrophie T cells using one particular Vβ region.
Contact sensitization induces proliferation of heterogeneous populations of hapten‐specific T cells1
Authors: Fehr Bettina S. , Takashima Akira , Matsue Hiroyuki , Gerometta John S. , Bergstresser Paul R. , Cruz Ponciano D.
Published: April 2006

Abstract To characterize the T cells that are activated during the induction of contact hypersensitivity (CH), two sets of studies were conducted: 1) dinitrophenol (DNP)‐specific proliferative responses of T cells in draining lymph nodes of BALB/c mice sensitized epicutaneously to dinitro‐fluorobenzene (DNFB) were examined, and 2) from these lymph node cells, DNP‐specific T cells were cloned by limiting dilution microculture and analyzed by FACS for surface markers, by RT‐PCR, HT2 bioassay and ELISA for cytokine expression at mRNA and protein levels respectively, and by proliferation assay for cytokine and antigen‐presenting cell (APC) requirements. Our results show that αβ TCR‐bearing T cells of both the CD4+ and CD8+ subtypes from lymph nodes of DNFB‐skin‐painted mice proliferate specifically to dinitrobenzene sulfonate (DNBS) in vitro. Four DNP‐specific, CD4+ T‐cell clones were characterized: clone 5S4 secreted IL‐4 and required 11‐4 for optimal growth; clone 5S10 secreted IL‐2 and required 1L‐2 for optimal growth; clone 5S2 secreted IL‐4 but required IL‐2 for optimal growth; and clone 5S8 secreted IL‐2 predominantly at 5 months, but switched to production of IL‐4 at 7 months. All four clones secreted IL‐10, and proliferated to DNBS when Langerhans cell (LC)‐enriched epidermal cells were used as APC. These findings indicate that heterogeneous populations of DNP‐specific T cells are activated in draining lymph nodes during the induction of CH to DNFB in BALB/c mice.
Humoral immunity to Malassezia furfur serovars A, B and C in patients with pityriasis versicolor, seborrheic dermatitis and controls
Authors: Ashbee H. R. , Fruin A. , Holland K. T. , Cunliffe W. J. , Ingham E.
Published: April 2006

Abstract This study examined the humoral immune responses to Malassezia furfur serovars A, B and C of 10 patients with pityriasis versicolor, 10 patients with seborrheic dermatitis and 20 age‐ and sex‐matched controls. A transferable solid‐phase ELISA was used to determine titres of total Igs, IgM, IgA and IgG specific to M. furfur serovars A, B and C. The results demonstrated that patients with seborrheic dermatitis had a significantly higher litre of total Igs to serovar A than patients with pityriasis versicolor; and that patients with seborrheic dermatitis had a significantly higher litre of IgA to serovar C than patients with pityriasis versicolor. The litres of total Igs for controls and patienls with seborrheic dermatitis were significantly lower to serovar B than to serovar C. A modified TSP ELISA was used to determine the titres of the IgG subclasses. Titres of IgG 1,3,4 to serovar B were significantly higher in seborrheic dermatitis patients than pityriasis versicolor patients and litres of IgG3, to serovar A were significantly higher in seborrheic dermatitis patients than pityriasis versicolor patients. However, despite the differences between the patient groups, none of these results was significantly different to those of controls. Thus, this study did not demonstrate any differences in humoral immunity of patients suffering from Malassezia‐associated dermatoses when compared to normal controls. These results may suggest that the humoral immune response to M. furfur is not related to the pathogenesis of Malassezia‐associated dermatoses, but simply to the carriage of M. furfur on the skin.
Skin changes in patients claiming to suffer from “screen dermatitis”: a two‐case open‐field provocation study
Authors: Johansson Olle , Hilliges Marita , Björnhagcn Viveca , Hall Kerstin
Published: April 2006

Abstract An open‐field provocation, in front of an ordinary TV set, of 2 patients regarding themselves as suffering from skin problems due to work at video display terminals (VDTs) is presented. Using immunohistochemistry, in combination with a wide range of antisera directed towards cellular and neurochemical markers, we were able to show a high‐to‐very high number of somatostatin‐immunoreactive dendritic cells as well as histamine‐positive mast cells in skin biopsies from the anterior neck taken before the start of the provocation. At the end of the provocation the number of mast cells was unchanged; however, the somatostatin‐positive cells had seemingly disappeared. The reason for this latter findings is discussed in terms of loss of immunoreactivity, increase of breakdown, etc. The high number of mast cells present may explain the clinical symptoms of itch, pain, edema and erythema. Naturally, in view of the present public debate, the observed results are highly provocative and, we believe, have to be taken seriously.
Effects of immunological responsiveness on Langerhans cell behavior in contact sensitization
Authors: Kolde Gerhard
Published: April 2006

Abstract The epicutaneous application of haptens results in a functional activation of the antigen‐presenting Langerhans cells (LCs) which is necessary for the induction of contact sensitivity. In this ultrastructural study, We investigated the effects of the immune response on these cellular properties of the LCs by using 2 strains of guinea pigs with genetically determined high and non responsiveness, respectively, to the strong sensitizer 2,4‐dinitrochlorobenzene (DNCB). After skin painting, both strains showed a similar cellular and endocytotic activation of the LCs and a similar intraepidermal localization of DNCB on immunoelectron microscopical visualization. There were however few LC‐lymphoid cell interactions in the non responders, in contrast to numerous such appositions in the other strain. Intravenous tolerization with 2,4‐dinitrobenzene‐l‐sulfonic acid, which is known to block the DNCB receptor of T cells, hampered the lymphoid cell interactions in the DNCB treated high responders, but it did not affect the LC activation. These in vivo observations demonstrate that the hapten‐induced changes of the LC properties is the initial, T‐cell independent event in contact sensitization.
Analysis of mast cell subpopulations (MCT, MCTC) in cutaneous inflammation using novel enzyme‐histochemical staining techniques
Authors: Algermissen B. , Bauer F. , Schadendorf D. , Kropp J.‐D. , Czarnetzki B. M.
Published: April 2006

Abstract In order to gain insights into the dynamics of mast cell subpopulations in normal and diseased skin, a novel enzyme‐histochemical double and triple staining method was employed that allowed the detection of metachromasia (toluidine blue) and the mast cell proteases tryp‐tase and chymase within the same cell. Cryostat sections were used of skin biopsies from the following specimens: normal skin (N=4), psoriasis (N=13), atopic eczema (N=7), lichen planus (N=6), interferon α2a injection sites (N=l) of a leukemic infiltrate and corresponding normal skin of the same patient before and after treatment. (i) Equal numbers of tryptase‐and chymase‐positive mast cells (MCTC) were obtained in all normal and diseased specimens in papillary and reticular dermis, with threefold increases around appendages, (ii) Tryptase‐positive mast cells (MCT) were absent in normal skin, but were markedly increased in a disease‐specific pattern within the papillary dermis, the inflammatory infiltrate and around appendages, (iii) Marked increases of MCT were also noted at interferon injection sites within the leukemic infiltrate, but not in the normal skin of the same patient. These data suggest that disease‐dependent mast cell dynamics involve only MCT in cutaneous inflammation and that MCT numbers are controlled by distinct, disease‐specific local tissue factors.
Regulation of monocyte and keratinocyte interleukin 6 production by transforming growth factor β
Authors: Möller A. , Schwarz A. , Neuner P. , Schwarz T. , Luger T. A.
Published: April 2006

Abstract There is strong evidence for a complex network‐like interaction between cytokines, growth factors and other mediators being responsible for cell growth and differentiation as well as for the outcome of an inflammatory reaction. Therefore, the regulation of the production of the ubiquitous proinflammatory cytokine interleukin 6 (IL‐6) by transforming growth factor β (TGFP) was investigated. Human peripheral blood mononuclear cells (PBMC), human normal keralinocytes (HNK), and an epidermoid carcinoma cell line (KB) were treated with TGFβ or TGFβ2 and subsequently IL‐6 secretion was evaluated. Addition of TGFβ1 as well as TGFβ2 to PBMC, HNK and KB cells resulted in a significantly increased release of IL‐6 activity. The inducing effect of TGFβ was dose dependent and maximal when supernatants were harvested 48 h after stimulation. In addition, upon Western blot analysis using a monoclonal IL‐6 antibody significantly increased amounts of IL‐6 protein were detected in KB cell supernatants following stimulation with TGFβ 1. These results were further confirmed at the transcriptional level using a cDNA probe specific for IL‐6 and Northern blot analysis. Accordingly, an increased IL‐6 inRNA expression in PBMC or KB cells was detected following TGFβl treatment. These findings indicate that TGFβ in contrast to its antiinflammatory capacities also may stimulate IL‐6 production in PBMC and keratinocytes. This further supports the possibly important immunoregulatory role of growth factors such as TGFβ.
The renin‐angiotensin‐system in the skin
Authors: Steckelings U. M. , Czarnelzki B. M.
Published: April 2006

Abstract Among the several hormonal systems regulating body functions, the renin‐angiotensin‐system has long been considered a classical endocrine system with angiotensin II, its effector hormone, being synthesized in and subsequently distributed by the circulation to act on its numerous, mainly renal and cardiovascular target organs throughout the body. Angiotensin II has long been regarded to be primarily responsible for the regulation of blood‐pressure and of volume‐ and electrolyte‐homeostasis. Recent evidence suggests that it also affects cellular proliferation and differentiation via the so‐called local or tissue‐renin‐angiotensin‐systems. Such trophic actions have already been observed in tissues not belonging to the renal or cardiovascular systems such as cultured cells of neuronal origin. Evidence for a role of angiotensin II in the skin is so far scanty and mainly based on the demonstration of angiotensin receptors on cultured human keratinocytes and in subcutaneous tissue of rats. Although almost every single component of the renin‐angio‐tensin‐system has already been identified in skin of one or another species, comprehensive data regarding the skin renin‐angiotensin‐system as a whole within one particular species, especially in man, are still lacking. The present manuscript reviews novel recent data regarding the renin‐angiotensin‐system particularly in skin, and it discusses a possible functional rôle of the cutaneous renin‐angiotensin‐system on the basis of these findings.
Messenger RNAs for the multifunctional cytokines interleukin‐1α, interleukin‐1β and tumor necrosis factor‐α are present in adnexal tissues and in dermis of normal human skin
Authors: Boehm Keith D. , Yun Jong K. , StrohI Kingman P. , Elmets Craig A.
Published: April 2006

Abstract Interleukin‐1α (IL‐1α), interleukin‐1 β (IL‐1β) and tumor necrosis factor‐α (TNF‐α) are 3 cytokines that play a key rôle in cutaneous homeostasis and in the immunopathogenesis of a number of dermatologic diseases. Most studies have focussed on their production by keratino‐cytes and Langerhans cells. To determine whether there are non‐epidermal sites of cytokine transcription, biopsy specimens of normal human skin were probed for IL‐lα, IL‐lβ and TNF‐α messenger RNAs using the method of in situ hybridization. The results demonstrate that each cytokine mRNA is present at multiple sites within the skin, including epidermal appendages and adnexal structures (hair follicles, sebaceous glands), the dermal microvasculature, arrectores pilorum smooth muscle, and the dermal connective tissue. These data provide evidence that in vivo there are multiple sites other than the epidermis of constitutive IL‐lα, 1L‐Iβ, and TNF‐α gene transcription in normal human skin.
Cytokine and protease levels in healing and non‐healing chronic venous leg ulcers
Authors: Harris I. R. , Yee K. C. , Walters C. E. , Cunliffe W J. , Kearney J. N. , Wood E. J. , Ingham E.
Published: April 2006

Abstract Leg ulcers present a common and recurring problem in older people creating discomfort and distress for the patient and a great cost to the health care services. Cultured keratinocyte grafts have been used by many investigators to stimulate healing of chronic venous ulcers. It has been proposed that they may do this by producing cytokines which modulate the healing process. However, the types and levels of cytokines in the leg ulcer fluid before and during healing arc not known. Wound fluid was collected from venous leg ulcers in 18 patients beneath occlusive Tega‐derm™ dressing for 4 to 6 h. The leg ulcers were divided on clinical criteria into ‘healing’ and ‘non‐healing’. PDGF‐AB, GM‐CSE IL‐1α, IL‐1β. IL‐6 and bFGF were measured by ELISA and the levels of IL‐1α, IL‐1β. and IL‐6 were also measured using biological assays. The effect of leg ulcer wound fluid on fibroblast and keratinocyte proliferation was measured indirectly by 3H‐thymidine incorporation and MTT assay. Total protein, albumin levels, fibronectin degrading activity and collagenase activity, both active and latent were measured. No statistically significant differences in the levels of cytokines or collagenase were identified between healing and non‐healing leg ulcers in the sample of leg ulcers studied. However, this study does give valuable information concerning the levels of cytokines and collagenase in chronic leg ulcer wound fluid.
Bullous pemphigoid: serum antibody titre and antigen specificity
Authors: Pas Hendri H. , de Jong Marcelus C. J. M. , Jonkman Marcel E , Heeres Klaas , Slijper‐Pal Ida J. , van der Meer Jan B.
Published: April 2006

Abstract 2 antigens have been identified as possible targets for autoantibody depositions in bullous pemphigoid: a 230‐kD protein (BP230) and a 180‐kD protein (BP180). We studied the relationship of these 2 antigens with the immunofluorescence determined serum antibody titre. 2 groups of bullous pemphigoid patients were selected on the basis of immunoblot‐determined antibody specificity. One group (13 patients) had antibody specificity for BP230 and not for BP180, while the other group (9 patients) had antibody specificity for BP180 and not for BP230. The immunofluorescence titres of the circulating antibodies determined on monkey oesophagus substrate displayed, for the BP230‐specific group, a mean of 1:1102. The maximal observed titre was 1:5120. The mean titre in the BP180‐specific group was only 1:29, with a highest titre of only 1:160. This result suggests that in routine indirect immunofluorescence of bullous pemphigoid sera, the contribution of the BP180‐specific antibodies to the total anti‐epidermal basement membrane zone antibody titre is relatively much lower than that of the BP230‐specilic antibodies. Thus, at high dilutions, only the BP230‐speeific antibodies contribute to the overall indirect immunfluorescence titre.
Detection of point mutations in human tyrosinase gene by improved allele‐specific amplification
Authors: Matsunaga Jun , Tomita Yasushi , Tagami Hachiro
Published: April 2006

Abstract Allele‐specific amplification (ASA) is a simple and non‐radioactive technique for detecting known point mutations that produce genetic diseases. Although this technique is based on the specific amplification of the target allele by a polymerase chain reaction (PCR) with allele‐specific primers, the specificity of the amplification may depend on various PCR conditions. To avoid non‐specific amplification which leads to false‐positive results in ASA, we modified both the normal and mutant allele‐specific primers so that they would have one constant base mismatch, located at the penultimate 3′ position. We confirmed that our modification could inhibit such unfavorable amplification by using as templates genomic DNAs of patients affected with tyrosinase‐negative oculocutaneous albinism (OCA). We then analyzed new patients affected with tyrosinase‐negative OCA, and based the diagnosis on both the results of a clinical examination and those of a hair bulb test using ASA with the modified allele‐specific primers. The results indicated that more than 3 alleles of the tyrosinase gene with a pathological mutation existed in Japanese patients.
Thy‐1 + dendritic cells express truncated form of POMC mRNA
Authors: Farooqui Jamal Z. , Medrano Estela E. , Boissy Raymond E. , Tigelaar Robert E. , Nordlund James J.
Published: April 2006

Abstract The present study was designed to examine the expression of proopiomelanocortin (POMC) and its related derivative peptide adreno‐corticotropic hormone (ACTH) in marine derived Thy‐1 + dendritic cells. Immunostaining using a polyclonal antibody specific to ACTH and parent POMC molecule indicated the presence of POMC and its derivative peptide, ACTH, in cultures of Thy‐1 + dendritic cells. To explore whether the POMC peptide is present as a reservoir or synthesized de novo in Thy‐1 + dendritic cells, Northern blot analysis using 30‐mer oligonucleotide probe for α‐MSH/ACTH precursor POMC was carried out in total RNA from these cells. Northern blot analysis revealed the presence of POMC like mRNA transcript. However, the observed size of transcript was smaller (∼0.9 kb) than that expressed by murine AtT20 cells (∼1.2 kb), an anterior pituitary tumor cell line used as a positive control. These observations suggest that the epidermal Thy‐14‐ lymphocytes, like thymic lymphocytes, might serve the epidermis as one source for the synthesis of POMC. The synthesis and presence of POMC in the epidermis may be related to some of the pigmentary anomalies observed in many mucocutaneous disorders.
Prostaglandin E1 inhibits TNFα‐induced T‐cell adhesion to endothelial cells by selective down‐modulation of ICAM‐1 expression on endothelial cells*
Authors: Weiss J. M. , Pilarski K. A. , Wcyl A. , Peschen M. , Schöpf E. , Vestweber D. , Vanscheidt W , Simon J. C.
Published: April 2006

Abstract Prostaglandins have been shown to be involved in the suppression of contact hypersensitivity (CHS) by so‐far ill understood mechanisms. T‐cell migration across the lining of cytokine‐activated endo‐thelial cells (EC) is thought to be a central step in the initiation of CHS. The aim of our investigation was therefore to examine whether prosta‐glandin E1 (PGE1) influences cytokine‐induced TK‐1 mouse T‐cell lymphoma adhesion to eEnd.2 mouse endothelioma cells. Here, we report that PGE1 (10−12− 10−8 M) dose‐dependently reduced TNFα‐induccd T‐cell binding, while TNFα‐unstimulated adhesion was not affected. To test whether PGE, acted primarily on T‐cells or on EC, they were separately pretreated with PGE, prior to the adhesion assay. Selective PGE1 pretreatment of eEnd.2, but not of TK‐1 dose‐dependenlly inhibited TNFα, stimulated T‐cell adhesion. Since binding of TK‐1 to TNFα‐treated eEnd.2 is mediated by the interaction of ICAM‐1 and VCAM‐1 (on EC) with their receptors LFA‐1 and VLA‐4 (on T‐cells), we further investigated whether PGE1 would modulate the expression of these molecules. FACS‐analysis revealed PGE1 to inhibit TNFα‐induced upregulation of ICAM‐1, but not of VCAM‐I on EC. Furthermore, constitutive LFA‐1 and VLA‐4 expression on T‐cells was not affected by PGE1. We conclude that PGE1 supresses T‐cell adhesion to EC by selectively inhibiting TNFα‐induced upregulation of ICAM‐1 on EC. This may be one mechanism by which prostaglandins suppress immune responses requiring T‐cell EC interactions such as contact hypersensitivity in skin.
Prevention and reversal of superantigen‐induced anergy by contact allergen exposure
Authors: Saloga J. , Enk A. H. , Becker D. , Spieles S. , Bellinghausen I. , Knop J.
Published: April 2006

Abstract The superantigen Staphylococcal enterotoxin B (SEB) and the contact allergen 2,4‐dinitrofluorbenzene (DNFB) both react with Vβ8+ T‐cells delivering distinct signals. Pre‐treatment with DNFB painted onto the same skin site where SEB was to be injected, prevented the induction of anergy in Vβ8+ T‐cells that was otherwise induced after SEB had been injected intradermally over a period of 2 weeks. Application of the irritant sodium dodecyl sulfate (SDS) instead of DNFB did not exert this effect. Application of DNFB at a site distant from the site where SEB was injected resulted in a much weaker inhibitory influence on the induction of anergy by SEB. Established anergy of Vβ8+ T‐cells (proliferative non‐responsiveness to SEB in vitro that could be overcome by addition of exogenous interleukin 2 (IL‐2)) could be largely reversed by repeated cutaneous exposure to DNFB painted to the site where SEB had been injected before. The moderate decrease of Vβ8 + T‐cells normally induced by SEB‐treatment was also partially prevented by DNFB pre‐treatment. The data indicate the importance of the sequence of signals delivered to T cells and the plasticity of the responsiveness of this cell type.
Differential promoter activity in benign and malignant human cells of skin origin*
Authors: Artuc Metin , Nürnberg Wolf , Platzer Matthias , Czarnetzki Beate M. , Schadendorf Dirk
Published: April 2006

Abstract In order to develop systems to express mammalian proteins in human skin‐derived cells, we tested 6 different viral and 1 eukaryotic promoter (pCMV, pRSV, pSV, pMMTV, pPoly E, pPoly L, pHMT) for their ability to drive the expression of the chloramphenicol acetyltrans‐ferase (CAT) enzyme in different human skin‐derived cells. DNA was transfected in human keratinocytes derived from normal foreskin and cervix, in the HPV‐negative cervical cancer line HT‐3 and in malignant melanoma cell lines (SK‐Mcl 23, SK‐Mcl 37) using a liposome‐based technique or calcium precipitation. Transaction efficacy was controlled by cotrans‐fection of a p‐galactosidase gene construct. The enzymatic activity of the CAT‐gene expression was determined by incubation of the cell extract prepared from the transfected cells with 14 C‐labeled chloramphenicol. The CMV‐promoter was highly active in all skin‐ or mucosal‐derived cells. In contrast to the strong CMV‐promoter, the RSV‐, SV‐, and HMT‐promoter were less active and varied in dependence of the cell type. The pattern of the promoter activity differed between benign and transformed genital keratinocytes. Only the SV‐promoter showed a comparable strong basal activity, which was restricted to the SK‐Mel 37 cells. In conclusion, the promoter activity has to be tested for each cell type depending on the aims of the gene expression.
Cell cycle regulators in the keratinocyte (cyclin‐cdk)
Authors: Inohara S. , Kitano Y. , Kitagawa K.
Published: April 2006

Abstract It has recently become clear that cyclin‐dependent kinase (cdk) complex regulates the cell cycle by phosphorylating Rb protein, a tumor suppressor protein. It is likely that this complex is a target of various growth factors and anti‐growth factors (UV TGF‐β etc.) in keratinocyte (KC). It has also been suggested that abnormalities in the cell cycle regulating mechanism such as increased activity of cyclin‐cdk due to mutation of p53, a tumor suppressor gene, and overexpression of cyclin D may be concerned with carcinogenesis of KC. Thus, recent studies indicate that the cyclin‐cdk complex is a common target of proliferation and carcinogenesis in KC.
Interleukin‐6 expression in the skin of patients with lupus erythematosus
Authors: Nürnberg W , Haas N. , Schadendorf D. , Czarnetzki B. M.
Published: April 2006

Abstract It has been proposed that interleukin‐6 may play a role in the pathogenesis of autoimmune diseases like lupus erythematosus. We have therefore investigated the immunoreactivity of IL‐6 in 32 skin biopsies of 23 patients suffering from chronic discoid lupus erythematosus (n=16), subacute cutaneous lupus erythematosus (n=5) and systemic lupus erythematosus (n = 5) as well as in uninvolved skin (n = 6) and in normal skin from healthy volunteers (n = 3). Increased immunohistochemical staining was detectable in 14 of 26 biopsies from lesional skin. The remaining biopsies from lesional, non‐lesional and normal skin displayed only minimal or no reactivity, but 8 out of 12 lupus erythematosus patients had been pretreated with local or systemic antiinflammatory drugs. Irrespective of the LE subtype, immunolabelling was generally most intense in the basal layer of the epidermis, with additional intense suprabasal staining in sections from 2 of 5 SLE patients. Preferential production of IL‐6 in the lower parts of the epidermis was confirmed by RNA in situ hybridization. No correlation was found between the deposition of immuno‐globulins and complement at the dermo‐epidermal junction and IL‐6 expression in keratinocytcs. These data suggest that IL‐6 may be involved in LE although its exact role in the pathogenesis of the disease needs to be further elucidated.
Tyrphostins suppress the growth of psoriatic keratinocytes
Authors: Ben‐Bassat Hannah , Vardi Daniel V. , Gazit Aviv , Klaus Sidney N. , Chaouat Malka , Hartzstark Zipora , Levitzki Alexander
Published: April 2006

Abstract Tyrosine kinase inhibitors of the tyrphostin family which block EGF receptor kinase are reported to arrest the growth of psoriatic keratinocytes in vitro. Three tyrphostins with the potency ratio AG555 >> AG18 >> AG814 were found to arrest growth with no adverse cytotoxic effects. The potency ratio to inhibit keratinocyte proliferation follows their potency to inhibit EGF receptor kinase activity in vitro. These compounds represent novel leads for the therapy of psoriasis.
Separation of epidermis from dermis in the Rhesus monkey
Authors: Frank John D. , Manson Jeanne M. , Cartwright Mark E.
Published: April 2006

Abstract Effective methods exist for separating epidermis from dermis for many species; however, a simple and effective skin separation method for non‐human primates is not available. This investigation describes an easy and reliable method for separating epidermis from dermis in Rhesus monkeys. Skin was shaved and washed prior to necropsy. Skin samples were placed on cardboard and then in Whirl‐Pak bags, frozen on dry ice and stored at −70°C. Just prior to the separation procedure, Whirl‐Pak bags were returned to dry ice storage. Immediately after removal from dry ice, each closed Whirl‐Pak bag was placed into a waterbath maintained between 60 and 67°C. After 2 minutes, the Whirl‐Pak bag was removed from the waterbath, opened and the skin surface of the application site was gently scraped with a scalpel blade to remove the epidermis. Effectiveness of removal was verified by histologic examination of the remaining dermal samples.
Regulation of CRABP II mRNA expression in human keratinocytes
Authors: Eller Mark S. , Muz Peter , Gilchrest Barbara A.
Published: April 2006

Abstract Cultured human neonatal keratinocytes were used to study the mechanisms and factors involved in the regulation of CRABP II gene expression. Post‐confluent, relatively differentiated keratinocyte cultures had higher levels of CRABP II mRNA, but nuclear run‐on experiments detected no sustained increase in CRABP II gene transcription rate between pre‐confluent and post‐confluent cells. Also, our studies could detect no change in the long half‐life <32 hours of this message in pre‐ and post‐confluent cultures. Hydrocortisone was found to reduce the confluency‐related increase in CRABP II mRNA in keratinocyte cultures. Because corticosteroids are known to reduce the effect of various cytokines, a series of epidermal cytokines were examined for a modulating effect on CRABP II mRNA content in cultured keratinocytes. IL1 α produced the greatest increase and IL6 the strongest reduction in the level of this message in cells grown in serum‐free, defined medium. These data support a role for CRABP II in the proliferation and differentiation of human keratinocytes and suggest that epidermal cytokines may at least in part regulate the expression of the CRABP II gene at the mRNA level.
Epiligrin is decreased in papulonodular basal cell carcinoma tumor nest basement membranes and the extracellular matrix of transformed human epithelial cells
Authors: Lazarova Zelmira , Domloge‐Hultsch Nouha , Yancey Kim B.
Published: April 2006

Abstract Patients with anti‐epiligrin cicatricial pemphigoid have anti‐basement membrane autoantibodies that immunoprecipitate a set of disulfide‐linked human keratinocyte polypeptides that co‐migrate in sodium dodecyl sulfate polyacrylamide gel electrophoresis with the same complex identified by monoclonal anti‐epiligrin (P1E1) and monoclonal anti‐nicein/kalinin (GB3) antibodies. In an attempt to further compare the reactivity of patient autoantibodies, P1E1 and GB3, these reagents were tested against the tumor nest basement membranes of 7 papulonodular basal cell carcinomas. These studies found that all of these reagents showed markedly decreased or no reactivity against this substrate. Though their concordant lack of reactivity failed to distinguish these antibodies, these studies did identify a significant defect in papulonodular basal cell carcinoma tumor nest basement membranes. Similarly, integrin subunits α6, β4, α3. and α2 as well as bullous pemphigoid antigens 1 and 2 (all potential receptors for the extracellular matrix ligands epiligrin and nicein/kalinin) were also reduced in these tumor nest basement membranes. These findings signify an extensive impairment in the lamina lucida of this neoplasm's basement membrane. Related comparative studies of normal human keratinocytes and transformed human epithelial cell lines (specifically, A‐431 and HaCat cells) showed that epiligrin production is markedly decreased in the latter. Decreased expression of epiligrin and nicein/ kalinin in papulonodular basal cell carcinoma tumor nest basement membranes in vivo and transformed epithelial cells in vitro indicate that this complex is a transformation‐sensitive cell adhesion ligand.
Divalent cations (Mg2+, Ca2+) differentially influence the β1 integrin‐mediated migration of human fibroblasts and keratinocytes to different extracellular matrix proteins
Authors: Lange Thilo Sascha , Kirchberg Katja , Bielinsky Anja Katrin , Leuker Andreas , Bank Ilan , Ruzicka Thomas , Scharffetter‐Kochanek Karin
Published: April 2006

Abstract Directed migration of keratinocytes and fibroblasts is a fundamental prerequisite in wound healing. Cation‐dependent affinity changes of integrins are responsible for cell adhesion to and deadhesion from extracellular matrix proteins and have been implicated in driving cell migration. The specific requirements for divalent cations in the integrin‐dependent migration of human dermal fibroblasts and human epidermal keratinocytes to various extracellular matrix proteins have been studied in vitro using blindwell Boyden chambers. The migration of the tested cells to collagen type I was mediated by the α2β2 integrins, to fibronectin by the combined action of the α3β2 and the α5β1 integrin, and the migration of fibroblasts to laminin dependent both on the α2β1 and the α6β1 integrins. No migration of keratinocytes to laminin was detected. Mg2+ alone induced cell migration with an optimum at 2 mM for fibroblasts and at 10 mM for keratinocytes. Ca2+ alone at 2 mM only marginally enhanced fibroblast and keratinocyte migration. At higher concentrations Ca2+ suppressed the stimulatory Mg2+ effect. 2 mM Ca2+ combined with 2 mM Mg2+ showed an additive stimulatory effect on the migration of fibroblasts to fibronectin. These data suggest that extracellular divalent cations differentially influence the integrin‐mediated cell migration. A concentration gradient of Mg2+/Ca2+, as reported in tissue injury, thus may play a regulatory role in cell migration required for tissue remodelling.