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Biomedical subjects

L Bruckner-Tuderman

Publications and source records attributed to L Bruckner-Tuderman.

At least 91 records · Page 5Linked to original sources
PubMed1994

Versican is expressed in the proliferating zone in the epidermis and in association with the elastic network of the dermis.

The expression of the large chondroitin sulfate proteoglycan versican was studied in human adult skin. For this purpose, bacterial fusion proteins containing unique portions of the versican core protein were prepared. Polyclonal antibodies against the fusion proteins specifically reacted with versican from a proteoglycan fraction of MG63 osteosarcoma cells. In immunohistochemical experiments, the affinity-purified antibodies localized versican in the stratum basale of the epidermis, as well as in the papillary and reticular layers of the dermis. An apparent codistribution of versican with the various fiber forms of the elastic network of the dermis suggested an association of versican with microfibrils. Both dermal fibroblasts and keratinocytes expressed versican in culture during active cell proliferation. In line with the observation that versican is absent in the suprabasal layers of the epidermis where keratinocytes terminally differentiate, culture conditions promoting keratinocyte differentiation induced a down-regulation of versican synthesis. In Northern blots versican mRNA could be detected in extracts from proliferating keratinocytes and dermal fibroblasts. Comparison of RNA preparations from semi-confluent and confluent fibroblast cultures demonstrated decreasing amounts of versican mRNA at higher cell densities. This inverse correlation of versican expression and cell density was confirmed by indirect immunofluorescence staining of cultured fibroblasts and keratinocytes. The localization of versican in the basal zone of the epidermis as well as the density dependence of versican in cell cultures suggest a general function of versican in cell proliferation processes that may not solely be confined to the skin.

Adult↗
PubMed1994

Intracellular accumulation of collagen VII in cultured keratinocytes from a patient with dominant dystrophic epidermolysis bullosa.

Expression of collagen VII, a candidate molecule for dystrophic epidermolysis bullosa, was analyzed in cultured keratinocytes from a patient with generalized dominant dystrophic epidermolysis bullosa (DEBD) of the Pasini subtype. Double immunofluorescence revealed an increased intracellular staining of collagen VII that co-localized with protein disulfide isomerase, a marker of the rough endoplasmic reticulum. Ultrastructural analysis of cultured DEBD cells showed dilated cisternae of the rough endoplasmic reticulum and numerous residual bodies, both of which contained abundant collagen VII as detected by immunoelectron microscopy. Immunoblotting of keratinocyte extracts indicated an increased ratio of cell-associated versus secreted soluble collagen VII in DEBD cells. Collagen VII mRNA was of normal size in the DEBD cells, but present in excessive amounts. The data suggest a mutation in the collagen VII gene that leads to intracellular accumulation and degradation of this collagen, and thus to a reduced number of anchoring fibrils at the dermo-epidermal junction, and subsequently to blistering of the skin in this family.

Adult↗
PubMed1994

Heterogeneity of severe dystrophic epidermolysis bullosa: overexpression of collagen VII by cutaneous cells from a patient with mutilating disease.

Severe mutilating recessive dystrophic epidermolysis bullosa presents with extensive blistering, scarring, and pseudosyndactylies. The skin of most affected individuals lacks normal anchoring fibrils and contains no, or drastically reduced amounts of, collagen VII, the major fibril component. Here we present evidence for molecular heterogeneity of the mutations underlying this phenotype. A patient with severe mutilating disease, but with apparently normal anchoring fibrils and abundant collagen VII, was defined. Indirect immunofluorescence examination of the patient's skin exhibited a strong staining for collagen VII at the dermo-epidermal junction and at the roof of a natural blister, and immunoblotting of skin extracts revealed collagen VII of normal size. The patient's keratinocytes expressed two- to threefold increased amounts of collagen VII at the mRNA and protein level compared to controls. Synthesis of matrix metalloproteases by the patient's keratinocytes was comparable to normal cells, indicating that the overexpression of collagen VII did not affect the synthesis of these enzymes. We hypothesize that in this patient a mutation affecting interactions of the anchoring fibrils with other components of the basement membrane zone underlies the disease.

Adult↗
PubMed1994

Collagen VII in severe recessive dystrophic epidermolysis bullosa: expression of mRNA but lack of intact protein product in skin and cutaneous cells in vitro.

The collagen VII gene, COL7A1, is the candidate gene for both the recessive and dominant forms of dystrophic epidermolysis bullosa (EBD). Collagen VII is a structural protein of the anchoring fibrils, which are rudimentary or altered in several subtypes of EBD. In severe recessive mutilating EBD, anchoring fibrils and collagen VII are not detectable in skin of most patients. To elucidate the underlying pathogenetic mechanisms, we analyzed collagen VII expression in cutaneous cells of six patients with this severe EBD subtype. Neither keratinocytes nor fibroblasts synthesized detectable amounts of collagen VII protein; however, Northern blot analysis revealed small amounts of normal-size collagen VII mRNA in both EBD and control fibroblasts. When the mRNA was amplified using reverse transcription-polymerase chain reaction, correct amplimers were present in all specimens. The results demonstrate that transcription of the COL7A1 gene occurs in these patients with severe mutilating EBD and suggest that post-transcriptional or post-translational events lead to absence of collagen VII protein from skin.

Adult↗
PubMed1994

Childhood epidermolysis bullosa acquisita.

We report a 6-year-old boy suffering from acquired epidermolysis bullosa, who presented with extensive lesions of the mucous membranes and disseminated, herpetiform and 'cluster of jewels'-like vesicles and bullae arising on erythematous plaques. Direct immunofluorescence showed linear deposits of IgG and C3 at the epidermal basement membrane zone. Indirect immunofluorescence demonstrated circulating autoantibodies (titre 1:128-1:256) directed at the blister floor of human NaCl-split skin, and reacting specifically with collagen VII of the anchoring fibrils, as demonstrated by immunoblotting. The disease was controlled with a combination of dapsone and prednisone. This case demonstrates the importance of modern immunological techniques in classifying childhood autoimmune bullous diseases, as precise diagnosis is important in determining an appropriate therapeutic regimen.

Autoantibodies↗
PubMed1994

Frequency of sensitization to 13 common preservatives in Switzerland. Swiss Contact Dermatitis Research Group.

From February 1989 to January 1990, the Swiss Contact Dermatitis Research Group conducted a 1-year study to examine the frequency of sensitization to a series of 13 common preservatives. A group of 2295 consecutive outpatients with suspected allergic contact dermatitis (age range 7-90 years, with a mean age of 42; 911 males, 1384 females) was tested. The %s of positive reactions to the preservatives studied are as follows, in descending order: formaldehyde 5.7%, benzalkonium chloride 5.5%, Kathon CG 5.5%, thimerosal 4.2%, chlorhexidine digluconate 2.0%, DMDM hydantoin 1.7%, paraben mix 1.7%, chloroacetamide 1.5%, Bronopol 1.2%, imidazolidinyl urea 1.0%, quaternium 15 1.0%, triclosan 0.8%, 2,4-dichlorobenzyl alcohol 0.4%. These relatively high values suggest a heavy exposure of the Swiss population to topical preservatives. Compared to previous studies, the sensitization rate to Kathon CG has stabilized in Switzerland over the last 2 years. Sensitization to formaldehyde portrayed impressive geographical variation, with sensitization rates up to 9% in western and only 3% in eastern Switzerland. The low sensitization rate to parabens argues for their inclusion in a medicament or preservative series, rather than in the standard series.

Administration, Cutaneous↗
PubMed1994

Collagen VII and bullous disorders of the skin.

Recent advances in molecular and cell biology of the cutaneous basement membrane zone have drastically increased our knowledge of the pathomechanisms underlying skin blistering disorders. Defects in genes coding for the structural proteins of the basement membrane zone have been defined in epidermolysis bullosa (EB), and abnormal expression of the structural proteins in some of its subtypes. The underlying cause for several subtypes of dystrophic EB are mutations in the gene for collagen VII, COL7A1. In vitro studies utilizing cells derived from the skin of such patients have helped understanding the pathogenetic pathways that lead from the mutation to the symptom, skin blistering. The data accumulated from analyses of the genetic disorders will also yield indirect information on the normal physiology of the skin and be highly relevant for the clarification of the etiopathogenesis of acquired blistering diseases.

Collagen↗
PubMed1994

Epidermolysis bullosa: pathogenetic pathways from mutations to symptoms.

Recent developments of the molecular and cell biology of the cutaneous basement membrane zone have greatly advanced our understanding of the pathomechanisms underlying skin blistering disorders. The heritable blistering diseases, the epidermolysis bullosa group, have been investigated as model diseases. Defects in genes coding for the structural proteins of the basement membrane zone have been defined in some EB subtypes and abnormal expression of the structural proteins in others. In vitro studies utilizing cutaneous cells derived from epidermolysis bullosa skin have helped to understand the pathogenetic pathways that lead from the mutation to the symptom, skin blistering. The data accumulated from analyses of the genetic disorders will yield indirect information on the normal physiology of the skin and be highly relevant for discerning the etiopathogenesis of acquired blistering diseases and for dermal-epidermal interactions required for reparative processes, such as wound healing.

Basement Membrane↗
PubMed1994

[Expression of the extracellular matrix proteoglycan, versican, in human skin].

Extracellular matrix chondroitin/dermatan sulfate proteoglycans are present in a wide variety of tissues including cartilage, placenta, aorta, tendon, brain and skin. They possibly participate in cellular processes such as cell adhesion, migration and proliferation. Recently, we have determined the entire primary structure of the large fibroblast proteoglycan, versican, on the basis of its cDNA sequence. Versican belongs to the family of large aggregating proteoglycans. Other members of the family, which have been characterized in terms of their primary structure, are aggrecan in cartilage and neurocan isolated from brain tissues. Due to the extensive sequence similarities between these three proteoglycans in the N- and C-terminal domains and due to the high degree of carbohydrate substitution, the generation of antibodies monospecific for versican has been difficult. To avoid cross-reactivity with aggrecan and neurocan, we therefore prepared unique portions of versican in a bacterial expression system and used them to immunize rabbits (Zimmermann et al., 1994). The affinity-purified anti-fusion protein antibodies specifically reacted with intact versican from an osteosarcoma cell line. First immunohistochemical experiments on cryo-sections of human skin revealed anti-versican staining in the stratum basale of the epidermis, as well as in the papillary and reticular layers of the dermis. By indirect immunofluorescence, Northern and Western blotting we could demonstrate that both, dermal fibroblasts and keratinocytes express versican in primary cultures. A striking inverse correlation between versican expression and cell density was observed. Analogous to the in vivo situation, keratinocytes induced to terminally differentiate ceased to express versican.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Keratinocytes from junctional epidermolysis bullosa do adhere and migrate on the basement membrane protein nicein through alpha 3 beta 1 integrin.

BACKGROUND: Junctional epidermolysis bullosa (JEB) encompasses several genodermatoses characterized by skin blistering, and possibly disturbed wound healing. Although the molecular defects underlying JEB are not known, we have demonstrated previously that nicein, an adhesive laminin-related basement membrane component, is immunologically altered in the very severe JEB of Herlitz type (H-JEB), and was expressed to a lesser extent in skin from patients with inversa JEB (I-JEB). In this study, we assessed adhesion and migration of H-JEB and I-JEB keratinocytes on exogenous nicein and laminin to get insights on the biologic function defective in JEB skin. EXPERIMENTAL DESIGN: Adhesion of cultured epidermal keratinocytes from H-JEB and I-JEB patients was assayed by quantitation of cell attachment 1 hour after seeding into microtiter wells coated with nicein or laminin. Cell migration and modulation by function-blocking antibodies to integrins was quantified by computer-assisted image analysis of the tracks left by the cells in a phagokinetic assay using gold particles coated with nicein or laminin. RESULTS: In spite of the fact that H-JEB keratinocytes do not produce normal immunoreactive nicein, they were able to adhere on exogenous nicein similarly to normal and I-JEB keratinocytes which produce nicein. Adhesion of both JEB and normal keratinocytes to laminin was weak compared with nicein. At low and high concentrations of nicein, a reduced migration response occurred with H-JEB keratinocytes whereas I-JEB cells behaved like their normal counterparts. Integrin alpha 3 beta 1 was dominantly involved in adhesion and migration of all these cells. Laminin did not support the migration of either JEB or normal keratinocytes. CONCLUSIONS: H-JEB and I-JEB keratinocytes which produce no or less nicein than normal keratinocytes are able to adhere and migrate on exogenous nicein. Integrin alpha 3 beta 1 which is specifically involved in migration and adhesion of keratinocytes on nicein does not appear altered in JEB. These data indicate that defective nicein rather than modifications of the nicein-recognizing receptor play a central role in the pathogenesis of H-JEB.

Adult↗
PubMed1994

Transforming growth factor-beta promotes deposition of collagen VII in a modified organotypic skin model.

BACKGROUND: Anchoring fibrils, which consist mainly of collagen VII, form a three-dimensional network below stratified squamous epithelia thus attaching the epithelial basement membrane to the underlying connective tissue. In skin, collagen VII is mainly produced by keratinocytes, and in vitro findings of keratinocyte-fibroblast cocultures suggest that mesenchymal-epithelial signaling plays a major role in the regulation of its expression. The matrix cytokine transforming growth factor-beta (TGF-beta) is a candidate mediator of such interactions, and exogenous TGF-beta has been shown to stimulate collagen VII expression in vitro. In the present study, the role of TGF-beta and other growth factors as mediators of mesenchymal-epithelial interactions in the regulation of collagen VII synthesis and deposition was investigated. EXPERIMENTAL DESIGN: The synthesis of collagen VII in monolayer cultures of normal human skin cells and in an organotypic skin equivalent system was investigated by indirect immunofluorescence staining. The role of soluble factors in collagen VII regulation was examined in a two-compartment coculture system, and the effect of medium conditioned by cocultures was tested on monocultures. Neutralizing antibodies to TGF-beta 2 were used to establish the relevance of this growth factor in the regulation of collagen VII synthesis. Modified skin equivalent cultures were constructed by cultivating an epithelium on top of devitalized human dermis deprived of the basement membrane. RESULTS: TGF-beta 2 was found to be a major autocrine or paracrine stimulator of collagen VII synthesis in cocultures. Incubation of the cultures with neutralizing antibodies to this growth factor abolished expression of collagen VII almost completely. Among other putative regulators of extracellular matrix metabolism, such as insulin-like growth factor, basic fibroblast growth factor, platelet-derived growth factor, epidermal growth factor, and calcium, insulin-like growth factor and calcium stimulated collagen VII synthesis by keratinocytes, but the extent was clearly less than by TGF-beta. Fibroblasts did not respond to either growth factor. However, experiments with fibroblasts grown on a keratinocyte-derived extracellular matrix suggested that collagen VII synthesis by fibroblasts is enhanced by matrix-mediated stimuli. A modified organotypic skin equivalent proved to be a suitable model to study the expression and deposition of basement membrane components. Collagen VII was expressed by basal keratinocytes after 3 weeks in culture under serum-free conditions. Its deposition at the dermoepidermal junction, however, depended on the addition of TGF-beta. CONCLUSIONS: TGF-beta is an important mediator of mesenchymal-epithelial interactions regulating collagen VII expression, and the deposition of collagen VII at the dermoepidermal interface is promoted by this growth factor. TGF-beta is a candidate factor for stabilization of the dermoepidermal junction in vivo during wound healing and in conditions where the junction zone is pathologically altered, e.g., dystrophic epidermolysis bullosa.

Collagen↗
PubMed1993

Epidermolysis bullosa junctionalis progressiva in three siblings.

Three siblings of Swiss origin with epidermolysis bullosa junctionalis progressiva are described. The following clinical features were present from school age: dystrophy of the nails, non-scarring blistering of the skin, mild skin atrophy, hypodontia and dental caries. Light microscopy showed subepidermal blistering. Direct immunofluorescence was negative. On indirect immunofluorescence staining of a fresh spontaneous blister, bullous pemphigoid antigen and laminin were localized to the blister roof, and collagen IV and collagen VII to the blister base, indicating junctional splitting. Electron microscopy revealed a normal dermo-epidermal junction zone, including normal hemidesmosomes. There were no deposits of electron-dense amorphous material.

Adolescent↗
PubMed1993

[Lupus erythematosus discoid-like dermatosis in a carrier of septic granulomatosis].

We present a clinically atypical case of discoid lupus erythematosus in the mother of a boy with chronic granulomatous disease. In this disorder, the phagocytes are unable to produce superoxide anion to degrade incorporated microorganisms. In addition to a discoid lupus erythematosus-like skin disease, recurrent stomatitis aphthosa, hidradenitis suppurativa and Raynaud phenomenon are markedly associated with heterozygote carriers of chronic granulomatous disease. Based on this conspicuous association, diverse models concerning the pathogenesis of lupus erythematosus are discussed.

Adult↗
PubMed1993

[The dermo-epidermal juncture. Recent aspects of pathophysiology].

The dermo-epidermal junction anchors the epidermis with the dermis but separates simultaneously these two layers. The complex structure of this zone is important for the coherence of the skin and for resistance against shearing forces. Abnormalities of the zone that result form e.g. binding of immune deposits or genetic defects, can lead to dermo-epidermal splitting and blister formation. Recent morphologic, biochemical and molecular biological investigations have generated new information that allows faster and easier diagnosis of bullous diseases. Investigations of patients with hereditary blistering disorders have revealed some of the mechanisms that underlie the different forms of epidermolysis bullosa.

Basement Membrane↗
PubMed1992

Transforming growth factor-beta stimulates collagen VII expression by cutaneous cells in vitro.

Collagen VII, the major component of cutaneous anchoring fibrils is expressed at a low level by normal human keratinocytes and fibroblasts in vitro. In cocultures of these two cell types, signals from fibroblasts enhance expression of collagen VII by keratinocytes and vice versa. In this study, the effects of a possible mediator of such a stimulation, transforming growth factor-beta (TGF-beta), were investigated. Its effect on the expression and deposition of the highly insoluble collagen VII was assessed in a semiquantitative manner by a newly developed enzyme-linked immunoassay which is based on immunoblotting. In keratinocyte monocultures, 0.5-20 ng/ml of TGF-beta 2 induced a dose-dependent stimulation of collagen VII expression as measured per microgram of DNA. The maximal enhancement was about sevenfold compared to controls. The effect of TGF-beta 2 was observed already after 12 h, with a steady increase at least up to 3 d. As previous studies have implicated, untreated cocultures of keratinocytes and fibroblasts exhibited a higher basic level of collagen VII expression, which could be further stimulated about twofold by TGF-beta 2. Fibroblasts alone synthesized very minor quantities of collagen VII and could be only weakly stimulated by TGF-beta 2. This growth factor seems a specific enhancer of collagen VII since the expression of laminin, collagen IV, as well as total protein was increased to a much lesser extent. Our data suggest that TGF-beta may be an important mediator of epithelial-mesenchymal interactions and may regulate the synthesis of the anchoring fibrils at the skin basement membrane zone.

Cell Communication↗
PubMed1992

Evidence for a structural abnormality of collagen VII in a patient with dystrophic epidermolysis bullosa inversa.

Recent studies indicate that in skin of patients with dystrophic epidermolysis bullosa (EB) inversa, anchoring fibrils have an abnormal ultrastructure, but the major protein of these fibrils, collagen VII, is expressed and detectable with antibodies at the dermo-epidermal junction. For molecular characterization of this rare EB phenotype, skin biopsies from a patient with dystrophic EB inversa were investigated with indirect immunofluorescence, immunoelectron microscopy, and immunoblotting. Ultrastructural analysis of clinically uninvolved skin showed sublamina densa splitting. In unblistered areas, focal groups of anchoring fibrils that appeared loosely polymerized and without a distinct crossbanding pattern were observed. Indirect immunofluorescence staining with antibodies to collagen VII exhibited a linear fluorescence at the dermo-epidermal junction and at the roof of a spontaneous blister. Immunoelectron microscopy demonstrated staining of the poorly assembled anchoring fibrils, but no significant reaction in areas where no fibrillar structures could be discerned. In contrast to normal control skin, immunoblotting showed immunoreactive collagen VII in both epidermal and dermal extracts. Moreover, the dermis-associated collagen VII appeared as a distinct doubleband that contained the tissue form of collagen VII (250-300 kD) and an additional band with a slightly smaller molecular weight. In epidermal extracts one band, of the size of the tissue form, was detected. The studies on the present patient suggest that a structural abnormality of collagen VII that prevents its aggregation to stable dimers or polymerization to distinct anchoring fibrils may contribute to the etiopathogenesis of dystrophic EB inversa.

Adult↗
PubMed1992

Keratinocytes and fibroblasts from a patient with mutilating dystrophic epidermolysis bullosa synthesize drastically reduced amounts of collagen VII: lack of effect of transforming growth factor-beta.

Keratinocytes and fibroblasts derived from skin of a patient with recessive dystrophic mutilating epidermolysis bullosa (EB) did not synthesize collagen VII as assessed by indirect immunofluorescence staining or immunoblotting, but expressed another basement membrane protein, laminin, in a normal manner. In contrast to control cells, no stimulation of collagen VII production was achieved in co-cultures of EB keratinocytes and fibroblasts. Further, treatment of normal keratinocytes or co-cultures with TGF-beta 2 significantly increased their expression of collagen VII, whereas the cytokine failed to induce its synthesis in the EB cells. Mixed co-cultures were constructed with normal fibroblasts and EB keratinocytes and vice versa. Both combinations showed strong expression of collagen VII in the normal cells but no synthesis in the EB counterparts. These results suggest that in this patient with severe mutilating dystrophic EB, inability of cutaneous cells to synthesize sufficient amounts of collagen VII underlies the lack of anchoring fibrils and skin fragility.

Cells, Cultured↗