bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular complexity of the cutaneous basement membrane zone
Authors: Christiano Angela M. , Uitto Jouni
Published: April 2006

Abstract Spectacular success has recently been made towards elucidation of the molecular basis of various forms of epidermolysis bullosa (EB), a group of heritable blistering skin diseases. The information derived from these studies has already had a profound impact in terms of precise diagnosis and classification, early prenatal prediction of the phenotype and genetic counseling in families at risk for recurrence. This review highlights recent progress made in defining the molecular basis of junctional and dystrophic forms of EB and the genotype/phenotype relationships established from these studies. Extensive molecular studies, such as the ones captured in this review, form a foundation for the rational design of gene therapies to counteract these conditions in the future.
All‐trans retinoic acid inhibits binding of 1,25‐dihydroxy‐vitamin D3 to the vitamin D receptor in cultured human keratinocytes
Authors: Fogh K. , Sølvsten H. , Jøhnke H. , Kragballe K.
Published: April 2006

Abstract Psoriasis is characterized by hyperproliferation and impared differentiation of epidermal keratinocytes (KCs). Psoriasis can be treated with derivatives of relinoic acid (RA) and vitamin D3. Analogues of vitamin D3 are able to inhibit proliferation and stimulate differentiation of KCs. In contrast, RA inhibits terminal differentiation of KCs. Interactions are known to occur between RA and vitamin D3 signalling pathways. The purpose of the present study was to determine the effect of all‐trans RA on the binding of 1,25‐dihydroxy‐vitamin D3 (1,25 (OH)2D3) to the vitamin D3, receptor (VDR) of cultured human KCs. Cultured KCs from normal adults were incubated with or without RA (10−9–10−7 M) for 4‐24 h. Cells were then harvested, homogenized and ultrasonicated. The extracted protein was incubated with 3H‐1,25 (OH)2D3 (0.015‐1.0 nM) with or without 250‐fold excess nonradioactive 1,25 (OH)2D3 for 24 h and specific binding was determined by use of the dextran coated charcoal binding assay. Western blot analysis utilizing the monoclonal antibody 9A7γ to VDR was performed on protein extracted from the KCs. The bands resulting from Western blot analysis were visualized by enhanced chemiluminescence. From Scatchard analysis it was found that KCs bind 1,25 (OH)2D3 with high affinity (Kd = 0.175 nM). This binding was dose and time dependently inhibited by RA (60% inhibition at 10−7 M after 24 h of incubation). By Western blot analysis, RA had no effect on the amount of protein extracted from KCs at any of the RA concentrations tested. In conclusion, these results show that binding of vitamin D3 to its receptor of human KCs can be inhibited markedly by RA without effecting the amount of protein. These results are in contrast to results with other cell types in which RA upregulates binding of 1,25 (OH)2D3 to the VDR. Because interaction between retinoids and vitamin D3 may occur at different levels during signal trans‐duction, it is not possible to predict from our results whether RA will inhibit the effects of vitamin D3, in vivo.
Lipoperoxidase activity of Pityrosporum: characterisation of by‐products and possible rôle in pityriasis versicolor
Authors: De Luca Chiara , Picardo Mauro , Brcathnach Aidon , Passi Siro
Published: April 2006

Abstract Modification of pigmentation and damage of melanocytes are characteristic features of skin colonisation by Pityrosporum orbiculare hyphae in pityriasis versicolor (PV). The yeast is lipophylic and lipid‐dependent, capable of oxidising unsaturated lipid components of skin surface, i.e. unsaturated fatty acids, cholesterol and squalene (SQ). The oxidation of unsaturated fatty acids gives rise to dicarboxylic acids (DA) which behave, in vitro, as competitive inhibitors of tyrosinase. In this work, we further investigate the oxidase activity of Pityrosporum in vitro, by evaluating (a) the generation of lipoperoxides in cultures supplemented with fatty acids at various degrees of unsaturation; (b) the mechanism of SQ oxidation; (c) the chemical characteristics of some by‐products of lipoperoxidation; (d) the formation of peroxisomes in fungal cells. In cultures supplemented with the saturated palmitic acid (C16:0) and monounsaturated oleic acid (C18:1 n‐9), low amounts of lipoperoxides were detected by a spectrophotometric test, whereas in cultures supplemented with di‐unsaturated linoleic acid (C 18:2 n‐6), significant concentrations were found. Gas chromatographymass spectrometry analyses showed the generation of linoleic acid hydroperoxides both in Pityrosporum cultures and following incubation of acetone powder of the fungus with the unsaturated fatty acid, indicating the presence of a lipoxygenase activity in the fungus. In cultures supplemented with linoleic acid plus SQ, and increase of lipoperoxide generation was observed and trans‐trans farnesal and squalene epoxides have been identified. Electron microscopic examinations have evidenced peroxisomes in cells grown in the presence of linoleic acid, whereas they were not delected in cultures supplemented with oleic acid and palmitic acid. The metabolic activities of peroxisomes, through the formation of hydrogen peroxide and the subsequent generation of hydroxyl radicals, may account for the peroxidation of SQ, which is not a substrate of lipoxygenase. Following these results, we propose a mechanism for DA generation by Pityrosporum metabolism and hypothesize that the lipoperoxidation process induced by lipoxygenase activity of the fungus may be the key to understanding the clinical appearance of skin manifestation of PV.
Simultaneously detected aberrant p53 tumor suppressor protein and HPV‐DNA localize mostly in separate keratinocytes in anogenital and common warts
Authors: Lassus Jan , Ranki Annamari
Published: April 2006

Abstract The E6 oncoprotein of human papillomavirus (HPV) is known to inactivate the control function on cell cycle exerted by p53 tumor suppressor protein in vitro by binding to p53 and thus facilitating the degradation of p53. We have applied a simultaneous in situ demonstration method for detecting p53 protein and HPV‐DNA on formalin‐fixed tissue sections, and investigated the in vivo interrelationship of p53 protein and HPV‐DNA. Immunohistochemical staining for p53 protein with polyclonal and monoclonal antibodies, recognizing both wild‐type (wt) and mutated p53 protein, was performed first and in situ DNA hybridization (ISH) for HPV types 6/11 or 16/18 with digoxigenin‐labelled probes thereafter. 47% (25/53) of 48 histologically confirmed primary or recurrent condylomata acuminata (CA), 2 Bowenoid papulosis (BP) and 3 common wart (CW) biopsies, positive for HPV 6/11 or HPV 16/18 DNA, showed keratinocytes immunopositive for p53 protein. Of these. 11 lesions with abundant numbers of p53‐positive cells were further analyzed with the double method. Signals for abnormal p53 protein and HPV‐DNA were detected in separate cell nuclei in all biopsies and, additionally, in the same cell nuclei in 3 biopsies (1 BP, 1 CA, 1 CW). Usually the p53 positivity localized more basally in the epidermis than HPV‐DNA, although p53‐ and HPV‐positive keratinocytes were always located closely. The findings were similar for HPV‐types 6/11 and 16/18. Our finding of both p53 and HPV‐6/11 signals in the same cell nuclei may indicate complexing of p53 and low‐risk HPV's without degradation of p53. Our results show abnormal p53 expression in HPV‐infected skin lesions, and suggest that p53 protein is susceptible to aberrations even in the cells in the vicinity of productive HPV infection. However, it is not yet fully understood how HPV interferes with p53 protein in these cells.
Thrombin and melittin activate phospholipase C in human HaCaT keratinocytes
Authors: Haase I. , Czarnetzki B. M. , Rosenbach T.
Published: April 2006

Abstract Following the activation of specific receptors, phospholipase C has been shown to cleave the membrane phospholipid phosphatidylinositol bisphosphate into the 2nd messengers inositol 1,4,5‐trisphosphate and di‐acylgiycerol. Both 2nd messengers contribute to the regulation of cellular proliferation. The receptor for bradykinin is coupled to this pathway in keratinocytes, but knowledge about other activators of phospholipase C is limited. Additional mediators and agents were therefore examined regarding their ability to activate phospholipase C in HaCaT keratinocytes. Analysis for 3H‐inositol phosphates was performed by anion‐exchange HPLC. Thrombin and melittin induced a time‐ and dose‐dependent release of inositol 1,4,5‐trisphosphate. Several other mediators examined such as angiotension II, neurotensin, C3a, pituitary adenylate cyclase activating peptide, phenylephrin, and prostaglandin E2, did not induce the formation of inositol phosphates. In view of the mitogenic activity and the increased formation of thrombin after tissue injury, the coupling of the thrombin receptor to phospholipase C in HaCaT keratinocytes suggests a rôle of this protease in epidermal wound healing.
Heparin induces fibroblast proliferation, cell‐matrix interaction and epidermal growth inhibition
Authors: Imaizumi Takashi , Jean‐Louis Francette , Dubertret Mari‐Liesse , Dubertret Louis
Published: April 2006

Abstract We have examined the effect of heparin on fibroblasts cultivated in monolayer or in a 3‐dimensional culture system: the so‐called collagen lattices. Thereafter, we have investigated the effect of heparin on the kinetics of epidermal growth on the collagen lattices. In monolayer culture, heparin stimulated the fibroblast growth with an optimal response at 0.01 mg/ml. The volume of treated fibroblasts was smaller than that of untreated controls. In the collagen lattices, heparin stimulated the fibroblast growth with an optimal respone at 0.1 mg/ml. The volume of treated fibroblasts was greater than that of untreated controls, the opposite to the result observed in monolayer culture. The beginning of the contraction of the collagen lattices was inhibited by heparin. Heparin inhibited epidermal growth on the immersion as well as on the emersion collagen lattices. These effects of heparin should be the consequences of heparin‐induced modifications of cell‐matrix interactions.
Specific detection of anti‐cell surface antibodies in herpes gestationis sera
Authors: Hashimoto T. , Amagai M. , Murakami H. , Higashiyama M. , Hashimoto K. , Nishikawa T.
Published: April 2006

Abstract We examined 42 herpes gestationis sera with immunofluorescence of normal human skin sections, and found that anti‐keratinocyte cell surface antibodies were detected specifically in 10 herpes gestationis sera. The diagnosis of these herpes gestationis cases was confirmed by detecting antibodies against the 180 kD bullous pemphigoid antigen with immunoblotting of its fusion protein. The results of immunoadsorption assay using baculoproteins of both pemphigus vulgaris and pemphigus foliaceus antigens indicated that the herpes gestalionis sera did not recognize common pemphigus antigens. Immunoblotting of human epidermal extracts and immunofluorescence of various tissues also suggested that the sera did not recognize any other desmosomal components or paraneoplastic pemphigus antigens. The significance of this reactivity is unclear. However, because no control bullous pemphigoid sera showed this reactivity, it may suggest a different pathophysiology between herpes gestationis and bullous pemphigoid.
Interleukin‐7. Biology and implications for dermatology
Authors: Möller Peter , Böhm Markus , Czarnetszki Beate M. , Schadendorf Dirk
Published: April 2006

Abstract In recent years, it has become apparent that IL‐7, originally characterized as a growth factor for pre‐B lymphocytes. also has important implications for the skin. Keratinocytes have been shown to produce IL‐7, which in turn can elicit a variety of biological responses on several cell types residing in the skin. IL‐7 has been demonstrated to augment the cytolytic activity of cytotoxic T cells (CTL) and natural killer (NK) cells against various neoplastic targets indcluding melanoma cells. Proliferation and long‐term survival of murine dendritic epidermal T lymphocytes (DETC) in vitro is supported by IL‐7. IL‐7 also induces secretion of inflammatory cytokines by monocytes/macrophages and renders these cells to become tumoricidal against melanoma cells. Normal and malignant melanocytes respond to IL‐7 with increased expression of intercellular adhesion molecule (ICAM‐I). In addition, IL‐7 has been shown to act as growth factor for Sczary cells, suggesting a röle of keratinocyte‐derived IL‐7 in the pathogenesis of cutaneous T cell lymphoma. Because of the potent in vitro immunomodulatory effects of IL‐7 which have been confirmed in mouse tumor models, IL‐7 may become a valuable additional agent to immunotherapeutical regimens currently studied in patients with advanced melanoma. This review summarizes our present knowledge about the molecular and immunological properties of IL7 with emphasis on the effects of that cytokine within the cutaneous compartment and the potentical clinical utility in dermatology.
In situ changes in the relative abundance of human epidermal cytokine messenger RNA levels following exposure to the poison ivy/oak contact allergen urushiol
Authors: Boehm Keith D. , Yun Jong K. , Strohl Kingman P. , Trefzer Uwe , Häffner Andreas , Elmets Craig A.
Published: April 2006

Abstract Epidermal keratinocytes in culture have been shown to produce many cytokines, and their proteins have been identified in skin tissue samples. It has therefore been assumed that these cytokines are transcribed in vivo by the epidermis in response to contact allergens. In this report, in situ hybridization was used to detect the messenger RNAs for interleukin‐1 alpha (IL‐1α), interleukin‐1 beta (IL‐1β) and tumour necrosis factor‐alpha (TNF‐α) in samples of human skin prior to and at various times after application of urushiol, the immunogenic component of poison ivy/oak. In sensitive subjects, IL‐1α and TNF‐α mRNAs showed a progressive increase in transcript levels that paralleled the clinical and histological features of the inflammatory process. The time–course of the IL‐1β response differed from that of IL‐1α and TNF‐α, in that there was an early (by 6 h after urushiol administration) elevation in IL‐1 β mRNA that occurred before there was evidence of inflammation and had returned to background levels by 72 h when the reaction had reached its peak. In contrast to urushiol‐sensitive subjects, urushiol‐allergic individuals did not exhibit an increase in IL‐1α. IL‐1β or TNF‐α mRNA levels. The data provide evidence for an in vivo role for epidermal IL‐1α, IL‐1β and TNF‐α transcription in the regulation of IL‐lβ and TNF‐α polypeptide levels in the epidermis in response to this common contact allergen.
UVA‐induced oxidative damage in fibroblasts cultured in a 3‐dimensional collagen matrix
Authors: Coulomb B. , Lebreton C. , Mathieu N. , Morlière P.
Published: April 2006

Abstract The exposure of dermal equivalent, i.e., fibroblasts cultured in a 3‐dimensional collagen matrix to realistic doses of UVA (≤ 200 kJ/m2), results in a lipid peroxidation process as evidenced by the release of thiobarbituric acid‐reactive substances in the supernatant. This peroxidalive process is shown to be associated with the presence of libroblasts and is inhibited by pre‐incubation with vitamin E. the well‐known chain‐breaking antioxidant. Moreover, the UVA irradiation triggers cyclotoxic effects which can also be reversed by preincubation with vitamin E. While the peroxidation extent is similar for fibroblasts cultured in monolayers or in dermal equivalent, the cytotoxic response to UVA is more pronounced in dermal equivalent.
Rôle of DNA damage in local suppression of contact hypersensitivity in mice by UV radiation
Authors: Kripke Margaret L. , Cox Patricia A. , Bucana Corazon , Vink Arie A. , Alas Lori , Yarosh Daniel B.
Published: April 2006

Abstract Exposure of mice to UVB radiation down‐regulates the induction of contact hypersensitivity (CHS) responses to haptens applied to the site of irradiation. Concomittantly, the activity of antigen‐presenting cells (APC) in the draining lymph nodes is decreased, and T lymphocytes that suppress the induction of CHS are induced. We assessed the röle of DNA damage in modulation of the CHS response by UV irradiation by applying liposomes containing T4 endonuclease V (T4N5) to the UV‐irradiated skin. Liposomal T4N5. which increases the rate of repair of cyclobutyl pyrimidine dimers (CPD) in DNA. prevented the reduction in the CHS response, the impairment in APC function, and the induction of transferrable immune suppression. Liposomes containing heat‐inactivated T4N5 did not restore immune responsiveness. In this model, hapten‐bearing APC from unirradiated mice also fail to induce CHS upon injection into UV‐irradiated recipients. This systemic effect of UV irradiation on APC function was also prevented by application of liposomes containing active, but not inactive, T4N5. These studies support the hypothesis that DNA damage is an essential initiator of one or more steps leading to impaired immune responsiveness after UV irradiation. They further imply that the release of cytokines that modulate APC function after UV irradiation is triggered by DNA damage.
Merkel cells and Merkel cell carcinoma express the BCL‐2 proto‐oncogene
Authors: Plettenberg Andreas , Pammer Johannes , Tschachler Enwin
Published: April 2006

Abstract The bcl‐2 proto‐oncogene, which is involved in the regulation of apoptosis, is expressed in a wide varity of fetal and adult tissues. We and others have demonstrated recently that in the human skin melanocytes, nevus cells and melanoma cells express bcl‐2 constitutively. In the present study, we have analysed the expression of bcl‐2 in Merkel cells and in Merkel cell carcinomas. In 2 colour immunofluorescence staining, normal human Merkel cells as identified by the expression of cytokeratins 8, 18 and 20, were also anti‐bcl‐2 positive. Staining of paraffin sections of Merkel cell carcinomas with an anti‐bcl‐2 monoclonal antibody revealed strong bcl‐2 protein immunoreactivity in all 5 tumors tested. Serial sections of Merkel cell carcinomas stained with the monoclonal antibodies CK 20. CAM 5.2, anti‐neuron‐specific enolase and anti‐bcl‐2 showed that the anti‐bcl‐2 reactive cells were indeed tumor cells. Our data demonstrate for the first lime, that normal human Merkel cells and Merkel cell carcinomas express hcl‐2 constitutively. Considering the biological function of the bcl‐2 proto‐oncogene, i.e., its anti‐apoptotic effect, it is conceivable that in the near future, modulations of the expression of this protein may offer a new strategy in the therapy of bcl‐2 expressing tumors such as Merkel cell carcinoma
High‐titer interferon‐α antibodies in a patient with pemphigus foliaceus
Authors: Prümmer Otto , Zillikens Detlef , Porzsolt Franz
Published: April 2006

Abstract Among 13 patients with pemphigus or bullous pemphigoid, high titers of anti‐interferon‐α (IFN‐α) antibodies were present in all serum samples of one patient suffering from pemphigus foliaceus. This patient was characterized by a relatively benign course of the disease. The IFN antibodies were of oligoclonal or polyclonal origin, predominantly of the IgG subtype, and displayed a broad spectrum of specificity including various natural and recombinant IFN‐α subtypes as well as recombinant IFN‐ωl. In vitro, these antibodies neutralized both the antiviral and antiproliferative activities of the respective IFN types. Recognition of the patient's endogenous IFN‐α demonstrated their autoantibody nature. The IFN antibodies were present at diagnosis and resistant to continued immunosuppressive treatment. Despite clinical remission, the IFN antibodies persisted, suggesting that they were not pathogenically related to the skin manifestations of the pemphigus. There were no signs of immune complex‐mediated organ damage. IFN antibodies constitute a new class of autoantibodies that may occur in conjunction with pemphigus and likely interfere with the endogenous IFN system.
Differential regulation of IL‐1 and IL‐1 receptor antagonist in HaCaT keratinocytes by tumor necrosis factor‐α and transforming growth factor‐β1
Authors: Kang Kefei , Hammerberg Craig , Cooper Kevin D.
Published: April 2006

Abstract Cytokines such as TNFα and TGFβ1 have potent effects on keratinocyte differentiation and have been implicated in cutaneous injury, immunologic reactions, and wound healing. To determine whether such conditions might alter the balance of epidermal keratinocyte IL‐1 and the IL‐1 receptor antagonist (IL‐1ra), TNFα and TGFβ1 were added to HaCaT cells, a human adult keratinocyte cell line. mRNA levels of IL‐1α, IL‐1β, and IL‐IRa were detected by polymerase chain reaction (PCR) on reverse transcribed RNA extracts, followed by Southern blot of the PCR products, 35S‐labeled probe hybridization, and quantification against standard curves. TNFα (100 ng/ml) at the 3‐h time point significantly induced increases in mRNA expression of Iliα (9.2±2.9 fold increase) and IL‐1β (2.5±0.7 fold increase) (n=7) which were concordant with increases in IL‐1α protein (7.1±1.3 fold increase) and II‐β protein (4.4±1.0 fold increase) measured by ELISA 24 h after stimulation. By contrast, icIL‐IRa mRNA and protein levels were not affected by TNFα. TGFβl induced a mild increase in IL‐lα mRNA (3.8±1.8 fold) and protein (3.5±1.2 fold). TGFβl did not affect IL‐1α mRNA levels but caused variable increases in IL‐1β protein levels. TGFβ1 did not alter icIL‐1Ra mRNA or protein levels. Inhibition of RNA synthesis with actinomycin D demonstrated that the rate of degradation of IL‐1β mRNA was reduced by treatment with TNFα. This stabilization of IL‐1β mRNA was specific, because TGFP I did not stabilize IL‐1β mRNA, and TGFβ1 and TNFα did not increase the stability of II‐1α mRNA. icIL‐l Ra mRNA was fairly stable over a 20 hour period and its slow degradation was not affected by treatment with either TNFα or TGFβ1, indicating a higher steady state stability of icIL‐1ra mRNA relative to IL‐1 mRNA's. Given the high rate of degradation of IL‐1α and IL‐1β mRNA, levels of these mRNAs may rapidly decrease while the icIL‐1ra mRNA levels remain constant, thus allowing for rapid dampening of IL‐1 activity soon after the stimuli provoking an inflammatory or reparative response have abated. In conclusion, TNFα and TGFβl, cytokines with potent effects on inflammation and differentiation, both induce keratinocyte IL‐1α mRNA and protein levels, but differentially regulate IL‐1β mRNA. They both exert little effect on IL‐1 Ra levels, which were constitutively highly stable. Such differential regulation provides mechanisms for separately controlling the relative activity of these cytokines under normal and disordered conditions.
Promoter region polymorphism in the human TNF‐α gene is not associated with lichen sclerosus
Authors: Clay F. E. , Cork M. J. , Wilson A. G. , Crane A. M. , Lewis F. , Harrington C. I. , Duff G. W.
Published: April 2006

Abstract The pathogenesis of lichen sclerosus remains unknown. However, it has been frequently associated clinically with autoimmunity. The MHC haplo‐type A1, B8, DR3 is associated with many autoimmune conditions and has also been associated with the uncommon allele of the tumour necrosis factor (TNF‐α) promoter polymorphism. This allele is also associated with higher production of TNF in vivo and in vitro, and thus it has been speculated that it is the TNF‐α gene which underlies the genetic association of many diseases with the autoimmune haplotype. There have been many reports of HLA associations with lichen scleroses, but these have not been concordant. We therefore decided to analyse the TNF‐oc polymorphism in patients with lichen scleroses to determine if TNF‐α was likely to play a role in susceptibility or severity of lichen scleroses. No association between alleles of the TNF‐α polymorphism and lichen scleroses was found.
Proinflammatory cytokines and their corresponding receptor proteins in eccrine sweat glands in normal and cutaneous leishmaniasis human skin
Authors: Ahmed Ahmed Abdelaziz , Nordlind Klas , Schultzberg Marianne , Lidén Sture
Published: April 2006

Abstract Paraformaldehyde‐fixed biopsy specimens of normal and chronic cutaneous leishmaniasis human skin were investigated for the presence and cellular distribution of interleukin‐1α, interleukin‐1β, interleukin‐6 and tumour necrosis factor‐α and the corresponding receptors in eccrine sweat glands, using an indirect immunoperoxidase technique. There was cytoplasmic staining for all 4 cytokines as well as their receptor proteins in the clear cells of the eccrine sweat glands of both normal and inflamed skin specimens. No staining could be seen in the dark cells or the myoepithelial cells, neither in normal nor in inflamed skin. However, a difference between normal and inflamed skin was observed in the ductal syslem. Thus, cell layers of the dermal ducts in leishmaniasis skin were stained for all 4 cytokines, with more intense labelling in the basal cell layer of the coiled ducts, while in the normal skin, an intense staining was more evident in the inner luminal layer, with variable and less intense labelling of the basal layer. The immunolabelling for the cytokine receptors within the dermal ducts exhibited similar staining intensity in both luminal and basal cell layers, except in the case of the IL‐6 receptor, which showed a moderate to intense signal in the basal cell layer but a weak staining of the luminal cell layer. Infiltrating inflammatory cells around the sweat gland apparatus in leishmaniasis skin exhibited immunoreactivilies for all cylokines and their corresponding receptors.
Metabolism of topical retinaldehyde and retinol by mouse skin in vivo: predominant formation of retinyl esters and identification of 14‐hydroxy‐4, 14‐retro‐retinol
Authors: Sass J. O. , Didierjean L. , Carraux P. , Plum C. , Nau H , Saurat J.‐H.
Published: April 2006

Abstract We have previously shown that retinaldehyde (RAL), a natural metabolite of (β‐carotene and retinol (ROL), can be used topically in human skin and exerts biological activity: it may be a convenient way to deliver multipotential vitamin A activity in epidermis. RAL can be converted enzymatically into 2 pathways: one leads to ROL (and then retinyl esters), the other to retin‐oic acid (RA). The aim of the present study was 2‐fold: (i) to see if RAL is metabolised in vivo when topically applied on mouse skin, and (ii) if so. to an‐alyse the occurrence and relative importance of the 2 metabolic pathways as compared to ROL. We studied by HPLC the metabolites detectable in mouse tail skin upon topical application of RAL and ROL. As compared to vehicle‐treated controls, RAL‐treated mouse skin contained low amounts of all‐trans RA and 13‐cis‐RA, whereas ROL content increased 10‐fold and retinyl esters 30‐fold after RAL application. As compared to RAL. ROL‐treated mouse skin showed no detectable RA, slightly less retinyl esters but a significant amount of 14‐hydroxy‐4, 14‐retro‐ROL (14‐HRR). a metabolite not previously reported in the skin. 14‐HRR was the predominant polar metabolite of ROL. These data indicate that keratinocytes metabolise topical RAL. thus confirming the concept of using RAL as a precursor. Both pathways are used but in significantly different proportions. Thus, only a low proportion of RAL is metabolised into all‐trans‐RA, which may explain the low irritancy profile of topical RAL and supports the concept of a controlled delivery of ligands. That keratinocytes predominantly channel RAL into storage forms indicates that RAL, should also be considered as a convenient way to load the epidermis with vitamin A. The detection of 14‐HRR. a metabolite not previously reported in skin, that promotes growth of B lymphocytes and activation of T lymphocytes, suggests distinct potentials of topical ROL and RAL.
Expression, but lack of calcium mobilization by high‐affinity IgE Fcε receptor I on human epidermal and dermal Langerhans cells
Authors: Shibaki A. , Ohkawara A. , Shimada S. , Ra C. , Aiba S. , Cooper K. D.
Published: April 2006

Abstract In atopic dermatitis (AD) patients, IgE molecules are demonstrated on the surface of Langerhans cells (LC). FcεRI molecules, which are present on the surface of LC in AD patients as well as normal individuals, are responsible for this binding. In this study, we have investigated phenotypic and functional characteristics of FcεRI on epidermal and dermal cell populations. Epidermal and dermal cell suspensions were prepared enzymatically with dispase followed by either trypsin or collagenase treatment, respectively. Peripheral blood basophils were negatively selected by excluding other leukocytes with surface marker staining. Consistent with previous reports, both peripheral blood basophils and epidermal LC were positively stained with anti FcεRI monoclonal antibody. In addition, an FcεRI positive population was demon‐strated among dermal HLA‐DR positive cells. These cells express significant amounts of HLA‐DR molecules (DRHi) and co‐express CD la molecules, which identifies them as LC‐like dendritie APC of the dermis. No other FcεRI positive population was found in the other dermal DRMid or DR populations, except for a minor DRlo population, presumably mast cells. To analyze whether these FcεRI molecules are signal transducing for LC, intracellular calcium mobilization after crosslinking of FcεRI was measured with How cytometry. Following crosslinking, peripheral blood basophils clearly increased intracellular calcium. On the other hand, neither normal epidermal LC nor dermal DRHiCD Ia+ cells changed their intracellular calcium level after FcεRI crosslinking. These data indicate that normal epidermal and dermal LC, but not basophils, are resistant to calcium flux following FcεRI engagement.
The chemotactic activity of T‐lymphocytes in response to interleukin 8 is significantly decreased in patients with psoriasis and atopic dermatitis
Authors: Zheng M. , Sun G. , Mrowietz U.
Published: April 2006

Abstract Involvement of T‐lymphocytes in the pathogenesis of psoriasis and atopic dermatitis is well established. The question arises as to whether not only tissue infiltrating but also circulating T‐lymphocytes are involved in the disease process. Therefore we sought to determine whether T‐lymphocytes from patients with psoriasis and atopic dermatitis show abnormal biological behavior to the proinflammatory chemokine interleukin 8 (IL‐8) in vitro as studied by their chemotactic activity. In addition, the expression of T‐cell activation markers such as HLA‐DR and interleukin 2 receptor (IL‐2R) were analysed with FACS‐technique. In all, 25 patients with psoriasis (13 patients with severe psoriasis and 12 patients with mild psoriasis) and 11 patients with atopic dermatitis were investigated. For comparison, T‐lymphocytes from 14 healthy controls were tested equally. The results show that T‐cell chemotactic responses to IL‐8 were significantly decreased in patients with severe psoriasis as compared to healthy controls. T‐cells from patients with atopic dermatitis demonstrated an even more pronounced decrease in chemotactic response as compared to T‐cells from psoriasis patients or healthy controls. In contrast, increased expression of activation markers HLA‐DR and IL‐2R were demonstrated in circulating T‐cells from patients with severe psoriasis and atopic dermatitis in comparison to healthy controls. It can be concluded that circulating T‐cells in patients with severe psoriasis and atopic dermatitis show a decreased in vitro chemotactic response to IL‐8. Furthermore, the in vivo phenotypic activation state of T‐lymphocytes in these patients seemed to be associated with their decreased in vitro functional capacity.
Pentoxifylline reduces the formation of sunburn cells
Authors: Schwarz Agatha , Mahnke Karsten , Luger Thomas A. , Schwarz Thomas
Published: April 2006

Abstract Irradiation with ultraviolet (UV) B light results in the formation of apoptotic keratinocyles called sunburn cells (SC). Although generation of SC appears to be one of the most characteristic features of UV‐induced skin damage and has been a well‐known phenomenon for a long time, the mechanisms involved are not quite clear. Recently, it was demonstrated that tumor necrosis factor α (TNFα) appears to be involved in the formation of SC since neutralization of TNFα both in vitro and in vivo reduced UVB‐induced apoptosis of keratinocytes. Pentoxifylline is a methylxanthine derivative suppressing the release of TNFα. Therefore, we studied whether PTX is able to prevent the formation of SC. Addition of PTX to UVB‐exposed HaCaT cells reduced DNA‐fragmentation as examined by nick translation evaluated by floweytometry. To prove whether PTX also reduces UVB‐induced apoptosis in vivo. BALB/c mice were exposed to UVB on their abdomens, skin biopsies performed 24 h later and SC counted. A single dose of 2000 J/m2 caused a significant induction of SC which were remarkably reduced when PTX was injected intraperitoneally 3 h before and 12 h after UVB exposure. In summary, the data demonstrate that PTX can reduce the formation of SC both in vitro and in vivo and thus further support that TNFα is involved in UVB‐induced apoptosis of keratinocytes.
Human epidermal melanocyte and keratinocyte melanotropin receptors: visualization by melanotropic peptide conjugated macrospheres (polyamide beads)
Authors: Jiang J. , Sharma S. D. , Hruby V. J. , Fink J. L. , Hadley M. E.
Published: April 2006

Abstract The objectives of this research were to determine whether melanotropin receptors are characteristic membrane markers of human epidermal melanocytes. Methodologies were developed to visualize these receptors by light microscopy. Multiple copies (up to a thousand) of [Nle4,D‐Phe7]α‐MSH, a superpotent analog of α‐melanocyte stimulating hormone (α‐MSH), were conjugated to a macromolecular carrier, large polyamide beads (macrospheres). Incubation in (the presence of the I conjugated macrospheres resulted in binding of human epidermal melanocytes to the macrospheres. Specificity of the binding of melanocytes of the melanotro‐pin‐conjugated macrospheres was demonstrated by several studies: (i) Binding of melanocytes to the conjugate was specific since it could be blocked by prior incubation of the cells in the presence of (the unconjugated hormone analog: (ii) The macrospheres after removal of the bound ligand did not bind to the melanocytes: (iii) Another peptide hormone ligand (e.g., a substance‐P analog) attached to the macrospheres failed to bind to the melanocytes: (iv) BI6/F10 mouse melanoma cells known lo express melanotropin receptors bound to the macrospheres; (v) Cells of nomnelanocyte origin (e.g., mammary cancer cells, lung cancer cells, fibroblasts) did not bind lo the macrospheres. One exception was that human epidermal keratinocytes also expressed melanotropin receptors as determined by all the criteria established for epidermal melanocyles. Thus, cell specific melanotropin receptors appear to be characteristic cell surface markers of epidermal melanocytes and keratinocytes.
The changes of stratum corneum interstices and calcium distribution of follicular epithelium of experimentally induced comedones (EIC) by oleic acid*
Authors: Choi Eung Ho , Ahn Sung Ku , Lee Seung Hun
Published: April 2006

Abstract Abnormal follicular keralinization is important for comedo formation in acne, but the precise mechanism is not known. A recent report about acne vulgaris suggested that an impaired water barrier function may be responsible for comedo formation, since the barrier dysfunction is accompanied by hyperkeratosis of the follicular epithelium. Furthermore, the integrity of the water barrier is crucial for the maintenance of the epidermal calcium gradient. Yet stratum corneum intercellular lipid structures, a major factor of the skin barrier function, and calcium distribution in the follicular epithelium of comedones, were not reported. To see SC intercellular lipid and calcium distribution of the follicular epithelium of comedo by electron microscopy, we applied oleic acid on the inner surface of the ear of New Zealand white rabbits to induce comedones, and then we obtained specimens and performed osmium and ruthenium tetroxide postfixation and calcium ion‐capture cytochemical procedure. We found incomplete lipid bilayer structures, prominent dilatation of lacunar domains and the loss of follicular epidermal calcium gradient in experimentally induced comedones. From our results, we suggest that the permeability barrier disruption in oleic‐acid‐applied follicular epithelium can be induced by the changes of SC intercellular membrane structures and lacunar dilatation, and the calcium gradient is lost, so follicular epithelial proliferation and hyperkeratosis can be induced and then comedo formation occurs.
Calcium and potassium inhibit barrier recovery after disruption, independent of the type of insult in hairless mice
Authors: Mao‐Qiang Man , Mauro Theodora , Bench Graham , Warren Raphael , Elias Peter M. , Feingold Kenneth R.
Published: April 2006

Abstract Disruption of the cutaneous permeability barrier induces metabolic responses in the epidermis which result in barrier recovery. Barrier disruption by either solvent treatment or tape stripping results in the loss of the epidermal calcium gradient. Previous studies in acetone treated hairless mice have shown that maintaining this calcium gradient inhibits barrier repair, suggesting that alterations in the epidermal calcium concentration may be an important signal for barrier homeostasis. In the present study, we show that in hairless mice disruption of the barrier by treatment with the detergent. SDS, also results in the loss of the calcium gradient, as demonstrated both semi‐quantitatively with ultrastructural cytochemical localization and quantitatively using proton induced X‐ray emission (PIXE). Additionally, immersion in calcium containing solutions delays barrier repair after either detergent (SDS treatment) or mechanical (tape stripping) disruption of the barrier, as reported previously for acetone treated skin. These results indicate that barrier disruption, regardless of the insult, induces changes in the epidermal calcium gradient which may play an important role in signaling the metabolic changes required for barrier homeostasis.
Spectrofluorimetric determination of 5‐methoxypsoralen pharmacokinetic in patients' serum
Authors: Saïd A. , Makki S. , Muret P. , Toubin G. , Humbert Ph. , Millet J.
Published: April 2006

Abstract In medicine, psoriasis and vitiligo are most often treated with PUVA therapy (psoralen plus ultraviolet A). The determination of psoralen in patients' blood is necessary, as it is admitted that the therapecutic efficiency depends on drug concentration in patients' serum. The amount of UVA to administer is inversely proportional to serum peak concentration. High performance liquid chromatography (HPLC) and gas chromatography are the most employed methods for determining psoralens in patients' serum. The 2 techniques are precise and very sensitive, but time consuming. The aim of this paper is to propose a suitable method which is rapid and simple. It is a spectrofluorimetric technique for 5‐methoxypsoralen (5‐MOP) determination in the serum of patients treated with PUVA therapy. 5‐MOP extraction was carried out with an heptane/dichloromethane mixture (4/1; v/v), according to the Stolk method (1980). A calibration curve (CC) was plotted from 5‐MOP concentrations (range 50, 100, 200, 300, 400, 500 ng/ml). The CC was linear with a good coefficient of correlation: r= 0.9971, and a suitable coefficient of variation (CV) of 7.0%. The recovery of the method ranged from 85.3±4.2 to 108±4.1%/. The assay precision gave a CV ranging from 0.10 to 6.90%, with an error inferior to ± 10%. The method did not reveal any interference from serum components on the 5‐MOP emission wavelength. The limit of detection of 5‐MOP was 15 ng/ml. The proposed procedure was proved to be appropriate for a rapid determination of 5‐MOP in patients' serum. This technique could also be employed for other proralens used in PUVA therapy (e.g., 8‐methoxypsoralen.)
Expression of GD3 disialoganglioside antigen on peripheral T‐lymphocytes in patients with disseminated malignant melanoma
Authors: Welte Birgitta , Handgretinger Rupert , Rassner Gernot , Fierlbeck Gerhard
Published: April 2006

Abstract Disialoganglioside antigens GD2 and GD3 are expressed on most melanoma cells. On melanoma surrounding T‐cells in immunohostological sections, disialogangliosides can also be found, as well as in a small % of T‐lymphocytes in peripheral blood from healthy persons. In order to find out if there is a difference in ganglioside expression on peripheral T‐lymphocytes between melanoma patients and healthy persons, we examined the expression of CD3 as T‐lymphocytic antigen and GD2 or GD3 antigens, respectively, by flow cytometry. We used peripheral mononuclear blood cells of 12 patients with advanced disseminated malignant melanoma and of 12 healthy control donors. For immunostaining, murine monoclonal antibodies Leu‐4, 14G2a and MB3.6 were used, recognizing CD3, GD2 and GD3. GD2 expression was found on only a low proportion of T‐lymphocytes in patients and healthy persons (pat.: mean = 1.2%± 0.7%, co.: mean = 0.4%± 0.4%). Disialoganglioside antigen GD3, however, could be demonstrated on an average of 8.4%± 4.6% of patients' and on 4.0%± 2.1% of healthy persons' T‐cells. There is a statistically significant difference (P < 0.01) between the data of patients' and control group. We conclude that there is a correlation between advanced malignant melanoma and expression of GD3 antigen on patients' peripheral T‐lymphocytes. The immunological relevance of our findings is discussed.
Identification of the LAMB3 hotspot mutation R635X in a Hungarian case of Herlitz junctional epidermolysis bullosa
Authors: Cserhalmi P. B. , Horvath A. , Boros V. , Sapi Z. , Kormendi M. , Christiano A. M. , Karpati S.
Published: April 2006

Abstract The Herlitz type of junctional epidermolysis bullosa (H‐JEB) is a serve blistering disease affecting the skin and mucous membranes, which is usually lethal within the first year of life. The laminin 5 genes have been implicated as candidate genes for most patients with H‐JEB. Recently two hotspot mutations were delineated in the LAMB3 gene, known as R42X and R635X, and have been noted in over 50% of mutant LAMB3 alleles. Here, we present a case of H‐JEB of Hungarian origin with a neonatal lethal outcome. Monoclonal antibody staining showed a lack of expression of the laminin 5β3 chain, as a possible result of a mutation in one of the laminin 5 genes. Screening of the family identified the previously described mutation R635X in exon 14 of LAMB3 in each of the parents and one healthy sibling in the heterozygous form, while proband was homozygous for R635X, and the other sibling proved to be genotypically normal. These results underscore the widespread prevalence of R635X in H‐JEB cases form around the world.
Profile of cytokine mRNA expression in spontaneous and UV‐induced skin lesions from actinic prurigo patients
Authors: Santos‐Martínez L. , Llorente L. , Baranda L. , Richaud‐Patin Y. , Torres‐Alvarez B. , Moncada B. , González‐Amaro R.
Published: April 2006

Abstract Background– Actinic prurigo (AP) appears to be an immunemediated disease triggered by exposure to ultraviolet light (UV). Objective– To assess the profile of cytokine production in skin lesions from AP patients. Methods– The cytokine production (IL‐1β 1L‐2, IL‐4. IL‐6. IL‐I0. 1L‐I3, TNF‐α, IFN‐τ, and TGF‐β) in skin biopsies from 12 AP lesions was determined by a semiquantitative coupled reverse transcription‐poly‐merase chain reaction. Result– We found expression of TGF‐β and 1L‐13 genes in most AP skin lesions; IL‐lβ. IL‐6. TNF‐α. IFN‐τ. and IL.‐10 were detected in some of these specimens. However, the levels of expression of all cytokines studied were not significantly different in AP skin lesions compared to nonlesional skin. Conclusions TGF‐β and IL‐13 might have a key role in both the inflammatory phenomenon and absence of significant expression of most cytokines in AP skin. The cytokine production in AP skin resembles that observed in rheumatoid synovium. a paucity in cytokine expression despite the presence of infiltrating activated mononuclcar cells.
Modulation of TGF‐β1 production from human keratinocytes by UVB
Authors: Lee Hyung‐Suk T. , Kooshesh Fatemeh , Saunder Daniel N. , Kondo Seiji
Published: April 2006

Abstract Transforming growth factor‐β (TGF‐β) plays an important role not only in cell growthasontrol but also in inflammation and immunoregulation. There are at least five different isoforms of TGF‐β. TGF‐β1 has a large variety of biological functions including the modulation of inflammation and the immune system and has most extensively been studied in skin. Since ultraviolet B (UVB) is known to induce skin erythema and immunosuppression. we sought to examine whether UVB would alter the expression and production of TGF‐β1 in normal human keralinocytes. Using reverse transcription‐polymerase chain reaction (RT‐PCR). constitutive expression of TGF‐β1 mRNA was detected in keralinocytes and the level of TGF‐β1 mRNA was increased 4 and 8 h after 300 J/m2 UVB irradiation. Production of TGF‐β1 protein in culture supernatants assayed by ELISA was also increased at 24 h after irradiation. Cycloheximide treatment blocked this TGF‐β1 protein induction indicating de novo protein synthesis of TGF‐βl from keratinocytes induced by UVB. These results suggest a possible role for TGF‐β1 in UVB‐induced skin inflammation and immunosuppression.
Epidermal localization and protective effects of topically applied superoxide dismutase
Authors: Filipe P. , Emerit I. , Vassy J. , Rigaut J. P. , Martin E. , Freitas J. , Fernandes A.
Published: April 2006

Abstract Data from the literature, as well as our previous work, indicate a protective effect of superoxide dismutase (SOD) in topical application against UV‐induced cutaneous damage. In the present article we show that pre‐treatment of the skin with SOD protects against PUVA‐induced inflammatory reactions not only in murine, but also in human skin. Using fluorescently labelled Cu, Zn SOD, epifluorescence microscopy and digital image processing, we demonstrate that the FITC fluorescence localizes in the stratum corneum and upper granulosa, as well as in the epidermal cell layer surrounding the lumina of the hair follicles. These findings were similar for murine and human skin. Since autofluorescence was eliminated by a special filter, it can be ascertained that the fluorescence observed in the tissues was due to FITC‐labelled SOD.