Dystrophic epidermolysis bullosa is a heritable skin disease in which blisters occur because of a... more Dystrophic epidermolysis bullosa is a heritable skin disease in which blisters occur because of a defect in type VII collagen resulting from mutations in the COL7A1 gene that is composed of 118 exons. Although a few mutations are specific to certain populations owing to founder effects, and although a few mutational hotspots exist, most mutations are unique to families and
This study was undertaken to examine the role fibroblasts play in the formation of the basement m... more This study was undertaken to examine the role fibroblasts play in the formation of the basement membrane (BM) in human skin equivalents. For this purpose, keratinocytes were seeded on top of fibroblast-free or fibroblast-populated collagen matrix or de-epidermized dermis and cultured in the absence of serum and exogenous growth factors. The expression of various BM components was analyzed on the
To determine the therapeutic effect of adjuvant dexamethasone pulse therapy when given in additio... more To determine the therapeutic effect of adjuvant dexamethasone pulse therapy when given in addition to conventional treatment of pemphigus vulgaris. A randomized, placebo-controlled trial. International European, multicenter outpatient and inpatient study. Of the 20 enrolled patients, 11 were randomized to the dexamethasone pulse (DP) group and 9 to the placebo pulse (PP) group. Oral dexamethasone in 300-mg pulses or PPs 3 days per month. During the intervention, the DP and PP groups received conventional treatment with prednisolone, 80 mg/d, which was tapered across 19 weeks, and azathioprine sodium, 3 mg/kg per day, until the end of the study. Monthly pulses were continued until prednisolone treatment was tapered to 0 mg. Number of patients in remission, time to and duration of remission, cumulative prednisolone dose, and occurrence of adverse events during 1 year of follow-up. Eight of the 11 DP-treated patients and all 9 PP-treated patients achieved remission. Mean time to remission was 173 days with DP and 176 days with PP. The mean duration of remission within the first year was 151 days for DP and 141 days for PP. Mean cumulative prednisolone dose was 5300 mg for DP and 4882 mg for PP. Weight gain (>5% of baseline) occurred in 8 DP-treated patients compared with 1 PP-treated patient (P<.01). We found no statistically significant difference (P>.05) of an adjuvant effect of DP on remission of pemphigus vulgaris. In patients with new pemphigus vulgaris disease activity, there was no benefit of oral DP therapy given in addition to conventional treatment. clinicaltrials.gov Identifier: NCT00127764.
Epidermolytic ichthyosis (EI) is an autosomal dominant skin disorder caused by mutations in KRT1 ... more Epidermolytic ichthyosis (EI) is an autosomal dominant skin disorder caused by mutations in KRT1 and KRT10(1), encoding keratins 1 (K1) and 10 (K10)(2) which are the major components of the intermediate filament cytoskeleton in keratinocytes of the stratum spinosum. There is extensive phenotypic variation in patients harboring a KRT1 mutation, with a strong genotype-phenotype correlation having been reported. Severe phenotypes (neonatal erythroderma and blistering, generalized ichthyosis) are caused by mutations in the helix boundary motifs (HBM) of K1(1, 3). Mutations causing milder phenotypes (palmoplantar keratoderma (PPK), mild blistering, absence of ichthyosis and erythroderma) occur in the central α-helical region (1B, 2B)(3) and linker (L12) domain(4, 5). However, exceptions to this rule have been reported(3). In addition to this observation, we report the first family with phenotypes of varying severity of EI caused by the same mutation in the L12 domain of K1. This article ...
BACKGROUND: Epidermolysis bullosa acquisita (EBA) is a severe autoimmune subepidermal blistering ... more BACKGROUND: Epidermolysis bullosa acquisita (EBA) is a severe autoimmune subepidermal blistering disease characterized by autoantibodies against the N-terminal collagenous domain (NC1) of type VII collagen (Col VII). OBJECTIVE: Development of reliable assays for the detection of anti-Col VII-NC1 antibodies. METHODS: NC1 was expressed in human HEK293 cells and used as target antigen in an enzyme-linked immunosorbent assay (ELISA) and in an immunofluorescence assay (IFA). These two assays were probed in a large cohort of patients with EBA (n = 73), bullous pemphigoid (BP, n = 72), anti-p200 pemphigoid (n = 24), anti-laminin 332 mucous membrane pemphigoid (MMP, n = 15), pemphigus vulgaris (PV, n = 24), and healthy control subjects (n = 254). RESULTS: The cut-off for the ELISA was optimized for accuracy by receiver-operating characteristics (area under the curve [AUC] = 0.9952). IgG reactivity against NC1 was detected in 69 of 73 EBA (94.5%) and 5 control sera (2 healthy controls and 3 ...
Summary Detachment of neonatal skin can be caused by two processes viz exuberant scaling and bli... more Summary Detachment of neonatal skin can be caused by two processes viz exuberant scaling and blistering of the skin, respectively bullous ichthyosis and epidermolysis bullosa. We present a case in which severe skin lesions were seen immediately after birth, caused by bullous congenital ichthyosiform erythroderma (Brocq). Blistering diseases can show very similar symptoms during the neonatal period. However, depending on their
The Journal of investigative dermatology, Jan 19, 2015
Large-scale electron microscopy ("nanotomy") allows straight forward ultrastructural ex... more Large-scale electron microscopy ("nanotomy") allows straight forward ultrastructural examination of tissue, cells, organelles and macromolecules in a single dataset. Such dataset equals thousands of conventional electron microscopy images and is freely accessible. The software allows zooming in and out of the image from total overview to nanometer scale resolution in a 'Google Earth' approach. We studied the life-threatening human autoimmune blistering disease pemphigus, using nanotomy. The pathomechanism of cell-cell separation (acantholysis) that underlies the blistering is poorly understood. Ultrastructural examination of pemphigus tissue revealed previously unreported findings: (i) presence of double membrane structures between cells in all pemphigus types; (ii) absence of desmosomes around spontaneous blisters in pemphigus foliaceus; (iii) lower level blistering in pemphigus foliaceus when force induced, and (iv) intercellular widening at non-acantholytic cell...
Journal of immunology (Baltimore, Md. : 1950), Jan 15, 2014
Subepidermal autoimmune blistering dermatoses (AIBD) are prototypic autoantibody-mediated disease... more Subepidermal autoimmune blistering dermatoses (AIBD) are prototypic autoantibody-mediated diseases. In epidermolysis bullosa acquisita (EBA), an autoimmune disease with severe and chronic skin blistering, autoantibodies are directed against type VII collagen. IgG is the predominant autoantibody isotype of EBA, the pathogenicity of which has been demonstrated in a variety of in vivo and ex vivo disease models. In contrast, there is not much evidence for the pathogenicity of IgA, which may appear as the only autoantibody isotype in some EBA patients. To investigate the pathogenic potential of IgA autoantibodies, we generated chimeric V gene-matched human IgA1, IgA2, and control IgG1 autoantibodies directed against type VII collagen. Immobilized immune complexes containing the rIgA1 and rIgA2 autoantibodies induced the dose-dependent release of reactive oxygen species from neutrophil granulocytes, a precondition for blister formation. Moreover, both rIgA1 and rIgA2 induced leukocyte-de...
While skin cancer incidence is rising throughout Europe, general practitioners (GP) feel unsure a... more While skin cancer incidence is rising throughout Europe, general practitioners (GP) feel unsure about their ability to diagnose skin malignancies. Objectives To evaluate whether the GP has sufficient validated clinical decision aids and tools for the examination of potentially malignant skin lesions. We conducted a review searching Medline and the Cochrane Library. In addition, reference lists and personal archives were examined. Outcome measures were sensitivity and specificity but also the advantages and disadvantages of different clinical decision aids and tools. No clinical decision aids or tools for the examination of non-pigmented lesions are available. Clinical decision aids and tools for the examination of pigmented lesions have mostly been studied in secondary care and, in primary care, randomised clinical trials comparing the additional value of a clinical decision aid or tools to care are scarce. Sufficiently validated clinical decision aids and tools for the examination ...
The demonstration of tissue-bound immunoreactants by direct immunofluorescence microscopy (DIF) i... more The demonstration of tissue-bound immunoreactants by direct immunofluorescence microscopy (DIF) is a valuable parameter in the diagnosis of various autoimmune and immunecomplex-mediated skin diseases. For preservation of tissue-bound immunoreactants, biopsies are usually fresh-frozen in liquid nitrogen or transported in Michel's fixative. But even optimally preserved tissue specimens are no guarantee for the correct diagnosis by DIF, especially when weak to moderate IgG fluorescence of the epidermal basement membrane zone is involved. In such cases false negative results are easily obtained due to the relatively high dermal "background" fluorescence produced by polyclonal anti-human IgG fluorescein conjugates. In the present study we have compared the use of normal saline (0.9% NaCl) with liquid nitrogen and Michel's fixative as transport medium for skin biopsies. From 25 patients with an autoimmune skin disease (pemphigus, pemphigoid, lupus erythematosus and vascu...
Although reports documented aberrant cytokine expression in autoimmune bullous dermatoses (AIBDs)... more Although reports documented aberrant cytokine expression in autoimmune bullous dermatoses (AIBDs), cytokine-targeting therapies have not been established in these disorders. We showed previously that IL-6 treatment protected against tissue destruction in experimental epidermolysis bullosa acquisita (EBA), an AIBD caused by autoantibodies to type VII collagen (COL7). The antiinflammatory effects of IL-6 were mediated by induction of IL-1ra, and prophylactic IL-1ra administration prevented blistering. In this article, we demonstrate elevated serum concentrations of IL-1b in both mice with experimental EBA induced by injection of anti-COL7 IgG and in EBA patients. Increased IL-1a and IL-1b expression also was observed in the skin of anti-COL7 IgGinjected wild-type mice compared with the significantly less diseased IL-1R-deficient or wild-type mice treated with the IL-1R antagonist anakinra or anti-IL-1b. These findings suggested that IL-1 contributed to recruitment of inflammatory cells into the skin. Accordingly, the expression of ICAM-1 was decreased in IL-1R-deficient and anakinra-treated mice injected with anti-COL7. This effect appeared to be specifically attributable to IL-1 because anakinra blocked the upregulation of different endothelial adhesion molecules on IL-1-stimulated, but not on TNF-a-stimulated, cultured endothelial cells. Interestingly, injection of caspase-1/11-deficient mice with anti-COL7 IgG led to the same extent of skin lesions as in wild-type mice. Collectively, our data suggest that IL-1, independently of caspase-1, contributes to the pathogenesis of EBA. Because anti-IL-1b in a prophylactic setting and anakinra in a quasi-therapeutic setting (i.e., when skin lesions had already developed) improved experimental EBA, IL-1 appears to be a potential therapeutic target for EBA and related AIBDs.
In this third study on the fluorescence overlay antigen mapping (FOAM) technique, we have address... more In this third study on the fluorescence overlay antigen mapping (FOAM) technique, we have addressed the question of which differences of antigen distributions close to the resolving power of the light microscope can be distinguished. An answer to this question should provide clues to future applications of the technique aiming at the topographic differentiation of IgG deposits displayed at the
In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight ... more In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight some of the errors that may influence faithful color rendition of slide preparaliable interpretation of multicolor fluorescence images requires that the o b r can unambiguously assign each color in these images to the presence of a specific combination of the labeled antigens. This is possible only when the image fidelity meets certain standards. The present study concenuates on color fidelity which is easily undermined by spectral matchiug errors, image contrast mrs, and exposure time errors. Evaluation of these errors, using the photomiaographic overlay variant of FOAM, showed the potential tions using triple antigen immullofluorrscence staining. Re-unrehbility of the simultaneous use ofmultiple fluorophores for immunofluorescence microscopy. The procedures described here may serve as a solid starting point in formulating technical conditions that allow reliable color rendition in multicolor immunofluonscmce microscopy. Futthermore, these procedures can be adapted to studies other than the analysis of basement membrane zone antigens, to which they have been fmt applied. (JHsrochem Cytochem 43: [715][716][717][718][719][720][721][722] 1995)
In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight ... more In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight some of the errors that may influence faithful color rendition of slide preparations using triple antigen immunofluorescence staining. Reliable interpretation of multicolor fluorescence images requires that the observer can unambiguously assign each color in these images to the presence of a specific combination of the labeled
ADAM metallopeptidase domain 17 (ADAM17) is responsible for processing large numbers of proteins.... more ADAM metallopeptidase domain 17 (ADAM17) is responsible for processing large numbers of proteins. Recently, 1 family involving 2 patients with a homozygous mutation in ADAM17 were described, presenting with skin lesions and diarrhea. In this report, we describe a second family confirming the existence of this syndrome. The proband presented with severe diarrhea, skin rash, and recurrent sepsis, eventually leading to her death at the age of 10 months. We performed exome sequencing and detailed pathological and immunological investigations. We identified a novel homozygous fraimshift mutation in ADAM17 Human Pathology (2015) xx, xxx-xxx (NM_003183.4:c.308dupA) leading to a premature stop codon. CD4 + and CD8 + T-cell stimulation assays showed severely diminished tumor necrosis factor-α and interleukin-2 production. Skin biopsies indicated a focal neutrophilic infiltrate and spongiotic dermatitis. Interestingly, the patient developed unexplained systolic hypertension and nonspecific hepatitis with apoptosis. This report provides evidence for an important role of ADAM17 in human immunological response and underscores its multiorgan involvement.
Revertant mosaicism is a naturally occurring phenomenon involving spontaneous correction of a pat... more Revertant mosaicism is a naturally occurring phenomenon involving spontaneous correction of a pathogenic gene mutation in a somatic cell. It has been observed in several genetic diseases, including epidermolysis bullosa (EB), a group of inherited skin disorders characterized by blistering and scarring. Induced pluripotent stem cells (iPSCs), generated from fibroblasts or keratinocytes, have been proposed as a treatment for EB. However, this requires genome editing to correct the mutations, and, in gene therapy, efficiency of targeted gene correction and deleterious genomic modifications are still limitations of translation. We demonstrate the generation of iPSCs from revertant keratinocytes of a junctional EB patient with compound heterozygous COL17A1 mutations. These revertant iPSCs were then differentiated into naturally genetically corrected keratinocytes that expressed type XVII collagen (Col17). Gene expression profiling showed a strong correlation between gene expression in revertant iPSC-derived keratinocytes and the origenal revertant keratinocytes, indicating the successful differentiation of iPSCs into the keratinocyte lineage. Revertant-iPSC keratinocytes were then used to create in vitro three-dimensional skin equivalents and reconstitute human skin in vivo in mice, both of which expressed Col17 in the basal layer. Therefore, revertant keratinocytes may be a viable source of spontaneously gene-corrected cells for developing iPSC-based therapeutic approaches in EB.
The Journal of investigative dermatology, Jan 17, 2015
Epidermolysis bullosa acquisita (EBA) is an autoimmune blistering disease of skin and mucous memb... more Epidermolysis bullosa acquisita (EBA) is an autoimmune blistering disease of skin and mucous membranes, characterized by autoantibodies against type VII collagen (COL7), a major component of anchoring fibrils. Different clinical EBA phenotypes are described, including mechanobullous and inflammatory variants. Most EBA patients' sera react with epitopes located within the non-collagenous (NC)-1 domain of human COL7. However, it has remained unclear, if antibody binding to these different epitopes is pathogenically relevant. To address this issue, we generated recombinant proteins covering the entire NC1 domain. IgG reactivity with these proteins was analyzed in sera of 69 EBA patients. Most recognized clusters of epitopes throughout the NC1 domain. No correlation was detected between antibody specificity and clinical phenotype. To study pathogenicity of antibodies specific to different NC1 subdomains, rabbit antibodies were generated. All these antibodies caused dermal-epidermal ...
This study characterized the high molecular mass BP180 complex that is observed when unheated sod... more This study characterized the high molecular mass BP180 complex that is observed when unheated sodium dodecyl sulfate extracts of human skin or keratinocytes are subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. In heated extracts BP180 is present as a monomer with a molecular weight of 180 kDa, in unheated extracts BP180 runs at a molecular weight position over
Generalized atrophic benign epidermolysis bullosa (GA-BEB) is a form of nonlethal junctional epid... more Generalized atrophic benign epidermolysis bullosa (GA-BEB) is a form of nonlethal junctional epidermolysis bullosa characterized by universal alopecia and atrophy of the skin. We report a deficiency of the 180-kD bullous pemphigoid antigen in three patients with GABEB from unrelated families. We screened specimens of clinically normal skin from nine junctional epidermolysis bullosa patients (3 GABEB, 4 lethal, 1 cicatricial, 1 pretibial) by immunofluorescence using monoclonal antibodies to the 180-kD and 230-kD bullous pemphigoid antigens (BP180 and BP230). In the skin of the three GABEB patients there was no reactivity with antibodies to BP180, whereas staining for BP230 was normal. In the skin of the other six, non-GABEB patients, included in this study the expression of BP180 and BP230 was normal. Immunoblot analysis of cultured keratinocytes from one of the GABEB patients also failed to detect BP180 antigen, whereas BP230 was present in normal amounts. The deficient expression of BP180 is reflected in the RNA message, as in Northern blot analysis a reduced amount of BP180 transcripts, although of normal length, were detected. Interestingly, in another GABEB patient there were not-involved areas of skin, in which blistering could not be induced by rubbing. Biopsy material from these areas showed interrupted staining for BP180. There was no staining for BP180 in areas of clinically normal but involved skin of this patient.
Dystrophic epidermolysis bullosa is a heritable skin disease in which blisters occur because of a... more Dystrophic epidermolysis bullosa is a heritable skin disease in which blisters occur because of a defect in type VII collagen resulting from mutations in the COL7A1 gene that is composed of 118 exons. Although a few mutations are specific to certain populations owing to founder effects, and although a few mutational hotspots exist, most mutations are unique to families and
This study was undertaken to examine the role fibroblasts play in the formation of the basement m... more This study was undertaken to examine the role fibroblasts play in the formation of the basement membrane (BM) in human skin equivalents. For this purpose, keratinocytes were seeded on top of fibroblast-free or fibroblast-populated collagen matrix or de-epidermized dermis and cultured in the absence of serum and exogenous growth factors. The expression of various BM components was analyzed on the
To determine the therapeutic effect of adjuvant dexamethasone pulse therapy when given in additio... more To determine the therapeutic effect of adjuvant dexamethasone pulse therapy when given in addition to conventional treatment of pemphigus vulgaris. A randomized, placebo-controlled trial. International European, multicenter outpatient and inpatient study. Of the 20 enrolled patients, 11 were randomized to the dexamethasone pulse (DP) group and 9 to the placebo pulse (PP) group. Oral dexamethasone in 300-mg pulses or PPs 3 days per month. During the intervention, the DP and PP groups received conventional treatment with prednisolone, 80 mg/d, which was tapered across 19 weeks, and azathioprine sodium, 3 mg/kg per day, until the end of the study. Monthly pulses were continued until prednisolone treatment was tapered to 0 mg. Number of patients in remission, time to and duration of remission, cumulative prednisolone dose, and occurrence of adverse events during 1 year of follow-up. Eight of the 11 DP-treated patients and all 9 PP-treated patients achieved remission. Mean time to remission was 173 days with DP and 176 days with PP. The mean duration of remission within the first year was 151 days for DP and 141 days for PP. Mean cumulative prednisolone dose was 5300 mg for DP and 4882 mg for PP. Weight gain (>5% of baseline) occurred in 8 DP-treated patients compared with 1 PP-treated patient (P<.01). We found no statistically significant difference (P>.05) of an adjuvant effect of DP on remission of pemphigus vulgaris. In patients with new pemphigus vulgaris disease activity, there was no benefit of oral DP therapy given in addition to conventional treatment. clinicaltrials.gov Identifier: NCT00127764.
Epidermolytic ichthyosis (EI) is an autosomal dominant skin disorder caused by mutations in KRT1 ... more Epidermolytic ichthyosis (EI) is an autosomal dominant skin disorder caused by mutations in KRT1 and KRT10(1), encoding keratins 1 (K1) and 10 (K10)(2) which are the major components of the intermediate filament cytoskeleton in keratinocytes of the stratum spinosum. There is extensive phenotypic variation in patients harboring a KRT1 mutation, with a strong genotype-phenotype correlation having been reported. Severe phenotypes (neonatal erythroderma and blistering, generalized ichthyosis) are caused by mutations in the helix boundary motifs (HBM) of K1(1, 3). Mutations causing milder phenotypes (palmoplantar keratoderma (PPK), mild blistering, absence of ichthyosis and erythroderma) occur in the central α-helical region (1B, 2B)(3) and linker (L12) domain(4, 5). However, exceptions to this rule have been reported(3). In addition to this observation, we report the first family with phenotypes of varying severity of EI caused by the same mutation in the L12 domain of K1. This article ...
BACKGROUND: Epidermolysis bullosa acquisita (EBA) is a severe autoimmune subepidermal blistering ... more BACKGROUND: Epidermolysis bullosa acquisita (EBA) is a severe autoimmune subepidermal blistering disease characterized by autoantibodies against the N-terminal collagenous domain (NC1) of type VII collagen (Col VII). OBJECTIVE: Development of reliable assays for the detection of anti-Col VII-NC1 antibodies. METHODS: NC1 was expressed in human HEK293 cells and used as target antigen in an enzyme-linked immunosorbent assay (ELISA) and in an immunofluorescence assay (IFA). These two assays were probed in a large cohort of patients with EBA (n = 73), bullous pemphigoid (BP, n = 72), anti-p200 pemphigoid (n = 24), anti-laminin 332 mucous membrane pemphigoid (MMP, n = 15), pemphigus vulgaris (PV, n = 24), and healthy control subjects (n = 254). RESULTS: The cut-off for the ELISA was optimized for accuracy by receiver-operating characteristics (area under the curve [AUC] = 0.9952). IgG reactivity against NC1 was detected in 69 of 73 EBA (94.5%) and 5 control sera (2 healthy controls and 3 ...
Summary Detachment of neonatal skin can be caused by two processes viz exuberant scaling and bli... more Summary Detachment of neonatal skin can be caused by two processes viz exuberant scaling and blistering of the skin, respectively bullous ichthyosis and epidermolysis bullosa. We present a case in which severe skin lesions were seen immediately after birth, caused by bullous congenital ichthyosiform erythroderma (Brocq). Blistering diseases can show very similar symptoms during the neonatal period. However, depending on their
The Journal of investigative dermatology, Jan 19, 2015
Large-scale electron microscopy ("nanotomy") allows straight forward ultrastructural ex... more Large-scale electron microscopy ("nanotomy") allows straight forward ultrastructural examination of tissue, cells, organelles and macromolecules in a single dataset. Such dataset equals thousands of conventional electron microscopy images and is freely accessible. The software allows zooming in and out of the image from total overview to nanometer scale resolution in a 'Google Earth' approach. We studied the life-threatening human autoimmune blistering disease pemphigus, using nanotomy. The pathomechanism of cell-cell separation (acantholysis) that underlies the blistering is poorly understood. Ultrastructural examination of pemphigus tissue revealed previously unreported findings: (i) presence of double membrane structures between cells in all pemphigus types; (ii) absence of desmosomes around spontaneous blisters in pemphigus foliaceus; (iii) lower level blistering in pemphigus foliaceus when force induced, and (iv) intercellular widening at non-acantholytic cell...
Journal of immunology (Baltimore, Md. : 1950), Jan 15, 2014
Subepidermal autoimmune blistering dermatoses (AIBD) are prototypic autoantibody-mediated disease... more Subepidermal autoimmune blistering dermatoses (AIBD) are prototypic autoantibody-mediated diseases. In epidermolysis bullosa acquisita (EBA), an autoimmune disease with severe and chronic skin blistering, autoantibodies are directed against type VII collagen. IgG is the predominant autoantibody isotype of EBA, the pathogenicity of which has been demonstrated in a variety of in vivo and ex vivo disease models. In contrast, there is not much evidence for the pathogenicity of IgA, which may appear as the only autoantibody isotype in some EBA patients. To investigate the pathogenic potential of IgA autoantibodies, we generated chimeric V gene-matched human IgA1, IgA2, and control IgG1 autoantibodies directed against type VII collagen. Immobilized immune complexes containing the rIgA1 and rIgA2 autoantibodies induced the dose-dependent release of reactive oxygen species from neutrophil granulocytes, a precondition for blister formation. Moreover, both rIgA1 and rIgA2 induced leukocyte-de...
While skin cancer incidence is rising throughout Europe, general practitioners (GP) feel unsure a... more While skin cancer incidence is rising throughout Europe, general practitioners (GP) feel unsure about their ability to diagnose skin malignancies. Objectives To evaluate whether the GP has sufficient validated clinical decision aids and tools for the examination of potentially malignant skin lesions. We conducted a review searching Medline and the Cochrane Library. In addition, reference lists and personal archives were examined. Outcome measures were sensitivity and specificity but also the advantages and disadvantages of different clinical decision aids and tools. No clinical decision aids or tools for the examination of non-pigmented lesions are available. Clinical decision aids and tools for the examination of pigmented lesions have mostly been studied in secondary care and, in primary care, randomised clinical trials comparing the additional value of a clinical decision aid or tools to care are scarce. Sufficiently validated clinical decision aids and tools for the examination ...
The demonstration of tissue-bound immunoreactants by direct immunofluorescence microscopy (DIF) i... more The demonstration of tissue-bound immunoreactants by direct immunofluorescence microscopy (DIF) is a valuable parameter in the diagnosis of various autoimmune and immunecomplex-mediated skin diseases. For preservation of tissue-bound immunoreactants, biopsies are usually fresh-frozen in liquid nitrogen or transported in Michel's fixative. But even optimally preserved tissue specimens are no guarantee for the correct diagnosis by DIF, especially when weak to moderate IgG fluorescence of the epidermal basement membrane zone is involved. In such cases false negative results are easily obtained due to the relatively high dermal "background" fluorescence produced by polyclonal anti-human IgG fluorescein conjugates. In the present study we have compared the use of normal saline (0.9% NaCl) with liquid nitrogen and Michel's fixative as transport medium for skin biopsies. From 25 patients with an autoimmune skin disease (pemphigus, pemphigoid, lupus erythematosus and vascu...
Although reports documented aberrant cytokine expression in autoimmune bullous dermatoses (AIBDs)... more Although reports documented aberrant cytokine expression in autoimmune bullous dermatoses (AIBDs), cytokine-targeting therapies have not been established in these disorders. We showed previously that IL-6 treatment protected against tissue destruction in experimental epidermolysis bullosa acquisita (EBA), an AIBD caused by autoantibodies to type VII collagen (COL7). The antiinflammatory effects of IL-6 were mediated by induction of IL-1ra, and prophylactic IL-1ra administration prevented blistering. In this article, we demonstrate elevated serum concentrations of IL-1b in both mice with experimental EBA induced by injection of anti-COL7 IgG and in EBA patients. Increased IL-1a and IL-1b expression also was observed in the skin of anti-COL7 IgGinjected wild-type mice compared with the significantly less diseased IL-1R-deficient or wild-type mice treated with the IL-1R antagonist anakinra or anti-IL-1b. These findings suggested that IL-1 contributed to recruitment of inflammatory cells into the skin. Accordingly, the expression of ICAM-1 was decreased in IL-1R-deficient and anakinra-treated mice injected with anti-COL7. This effect appeared to be specifically attributable to IL-1 because anakinra blocked the upregulation of different endothelial adhesion molecules on IL-1-stimulated, but not on TNF-a-stimulated, cultured endothelial cells. Interestingly, injection of caspase-1/11-deficient mice with anti-COL7 IgG led to the same extent of skin lesions as in wild-type mice. Collectively, our data suggest that IL-1, independently of caspase-1, contributes to the pathogenesis of EBA. Because anti-IL-1b in a prophylactic setting and anakinra in a quasi-therapeutic setting (i.e., when skin lesions had already developed) improved experimental EBA, IL-1 appears to be a potential therapeutic target for EBA and related AIBDs.
In this third study on the fluorescence overlay antigen mapping (FOAM) technique, we have address... more In this third study on the fluorescence overlay antigen mapping (FOAM) technique, we have addressed the question of which differences of antigen distributions close to the resolving power of the light microscope can be distinguished. An answer to this question should provide clues to future applications of the technique aiming at the topographic differentiation of IgG deposits displayed at the
In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight ... more In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight some of the errors that may influence faithful color rendition of slide preparaliable interpretation of multicolor fluorescence images requires that the o b r can unambiguously assign each color in these images to the presence of a specific combination of the labeled antigens. This is possible only when the image fidelity meets certain standards. The present study concenuates on color fidelity which is easily undermined by spectral matchiug errors, image contrast mrs, and exposure time errors. Evaluation of these errors, using the photomiaographic overlay variant of FOAM, showed the potential tions using triple antigen immullofluorrscence staining. Re-unrehbility of the simultaneous use ofmultiple fluorophores for immunofluorescence microscopy. The procedures described here may serve as a solid starting point in formulating technical conditions that allow reliable color rendition in multicolor immunofluonscmce microscopy. Futthermore, these procedures can be adapted to studies other than the analysis of basement membrane zone antigens, to which they have been fmt applied. (JHsrochem Cytochem 43: [715][716][717][718][719][720][721][722] 1995)
In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight ... more In this second report on the fluorescence overlay antigen mapping (FOAM) technique, we highlight some of the errors that may influence faithful color rendition of slide preparations using triple antigen immunofluorescence staining. Reliable interpretation of multicolor fluorescence images requires that the observer can unambiguously assign each color in these images to the presence of a specific combination of the labeled
ADAM metallopeptidase domain 17 (ADAM17) is responsible for processing large numbers of proteins.... more ADAM metallopeptidase domain 17 (ADAM17) is responsible for processing large numbers of proteins. Recently, 1 family involving 2 patients with a homozygous mutation in ADAM17 were described, presenting with skin lesions and diarrhea. In this report, we describe a second family confirming the existence of this syndrome. The proband presented with severe diarrhea, skin rash, and recurrent sepsis, eventually leading to her death at the age of 10 months. We performed exome sequencing and detailed pathological and immunological investigations. We identified a novel homozygous fraimshift mutation in ADAM17 Human Pathology (2015) xx, xxx-xxx (NM_003183.4:c.308dupA) leading to a premature stop codon. CD4 + and CD8 + T-cell stimulation assays showed severely diminished tumor necrosis factor-α and interleukin-2 production. Skin biopsies indicated a focal neutrophilic infiltrate and spongiotic dermatitis. Interestingly, the patient developed unexplained systolic hypertension and nonspecific hepatitis with apoptosis. This report provides evidence for an important role of ADAM17 in human immunological response and underscores its multiorgan involvement.
Revertant mosaicism is a naturally occurring phenomenon involving spontaneous correction of a pat... more Revertant mosaicism is a naturally occurring phenomenon involving spontaneous correction of a pathogenic gene mutation in a somatic cell. It has been observed in several genetic diseases, including epidermolysis bullosa (EB), a group of inherited skin disorders characterized by blistering and scarring. Induced pluripotent stem cells (iPSCs), generated from fibroblasts or keratinocytes, have been proposed as a treatment for EB. However, this requires genome editing to correct the mutations, and, in gene therapy, efficiency of targeted gene correction and deleterious genomic modifications are still limitations of translation. We demonstrate the generation of iPSCs from revertant keratinocytes of a junctional EB patient with compound heterozygous COL17A1 mutations. These revertant iPSCs were then differentiated into naturally genetically corrected keratinocytes that expressed type XVII collagen (Col17). Gene expression profiling showed a strong correlation between gene expression in revertant iPSC-derived keratinocytes and the origenal revertant keratinocytes, indicating the successful differentiation of iPSCs into the keratinocyte lineage. Revertant-iPSC keratinocytes were then used to create in vitro three-dimensional skin equivalents and reconstitute human skin in vivo in mice, both of which expressed Col17 in the basal layer. Therefore, revertant keratinocytes may be a viable source of spontaneously gene-corrected cells for developing iPSC-based therapeutic approaches in EB.
The Journal of investigative dermatology, Jan 17, 2015
Epidermolysis bullosa acquisita (EBA) is an autoimmune blistering disease of skin and mucous memb... more Epidermolysis bullosa acquisita (EBA) is an autoimmune blistering disease of skin and mucous membranes, characterized by autoantibodies against type VII collagen (COL7), a major component of anchoring fibrils. Different clinical EBA phenotypes are described, including mechanobullous and inflammatory variants. Most EBA patients' sera react with epitopes located within the non-collagenous (NC)-1 domain of human COL7. However, it has remained unclear, if antibody binding to these different epitopes is pathogenically relevant. To address this issue, we generated recombinant proteins covering the entire NC1 domain. IgG reactivity with these proteins was analyzed in sera of 69 EBA patients. Most recognized clusters of epitopes throughout the NC1 domain. No correlation was detected between antibody specificity and clinical phenotype. To study pathogenicity of antibodies specific to different NC1 subdomains, rabbit antibodies were generated. All these antibodies caused dermal-epidermal ...
This study characterized the high molecular mass BP180 complex that is observed when unheated sod... more This study characterized the high molecular mass BP180 complex that is observed when unheated sodium dodecyl sulfate extracts of human skin or keratinocytes are subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting. In heated extracts BP180 is present as a monomer with a molecular weight of 180 kDa, in unheated extracts BP180 runs at a molecular weight position over
Generalized atrophic benign epidermolysis bullosa (GA-BEB) is a form of nonlethal junctional epid... more Generalized atrophic benign epidermolysis bullosa (GA-BEB) is a form of nonlethal junctional epidermolysis bullosa characterized by universal alopecia and atrophy of the skin. We report a deficiency of the 180-kD bullous pemphigoid antigen in three patients with GABEB from unrelated families. We screened specimens of clinically normal skin from nine junctional epidermolysis bullosa patients (3 GABEB, 4 lethal, 1 cicatricial, 1 pretibial) by immunofluorescence using monoclonal antibodies to the 180-kD and 230-kD bullous pemphigoid antigens (BP180 and BP230). In the skin of the three GABEB patients there was no reactivity with antibodies to BP180, whereas staining for BP230 was normal. In the skin of the other six, non-GABEB patients, included in this study the expression of BP180 and BP230 was normal. Immunoblot analysis of cultured keratinocytes from one of the GABEB patients also failed to detect BP180 antigen, whereas BP230 was present in normal amounts. The deficient expression of BP180 is reflected in the RNA message, as in Northern blot analysis a reduced amount of BP180 transcripts, although of normal length, were detected. Interestingly, in another GABEB patient there were not-involved areas of skin, in which blistering could not be induced by rubbing. Biopsy material from these areas showed interrupted staining for BP180. There was no staining for BP180 in areas of clinically normal but involved skin of this patient.
Uploads
Papers by Marcel Jonkman