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Biomedical subjects

Leena Bruckner-Tuderman

Publications and source records attributed to Leena Bruckner-Tuderman.

At least 19 recordsLinked to original sources
PubMed2007

Frequency, function and CLA expression of CD4+CD25+FOXP3+ regulatory T cells in bullous pemphigoid.

Bullous pemphigoid (BP) is an autoimmune blistering skin disease associated with autoantibodies to collagen XVII and tissue-separation along the dermo-epidermal junction. We addressed the question whether the loss of tolerance in BP patients is associated with a reduction and/or functional impairment of CD4+CD25+FOXP3+ regulatory T cells, which are essential for the active maintenance of self tolerance. The relative and absolute frequency of CD4+CD25+ and CD4+CD25(high) regulatory T cells in the peripheral blood of newly diagnosed, untreated patients was similar to that of healthy controls. Interestingly, more than 50% of circulating CD4+CD25(high) regulatory T cells from both patients as well as healthy controls expressed cutaneous lymphocyte-associated antigen. Considerable numbers of FOXP3+ cells were detected in lesional skin of patients. CD4+CD25+ regulatory T cells of patients were functionally intact as assessed by their ability to suppress allogeneic as well as antigen-specific T-cell proliferation. These data argue against a general defect of CD4+CD25+FOXP3+ regulatory T cells in patients with BP.

Aged↗
PubMed2006

Molecular basis of inherited skin-blistering disorders, and therapeutic implications.

Epidermolysis bullosa (EB) and associated skin-fragility syndromes are a group of inherited skin diseases characterised by trauma-induced blistering of the skin and mucous membranes. Mutations in at least 14 distinct genes encoding molecular components of the epidermis or the dermal-epidermal junction (DEJ) can cause blistering skin diseases that differ by clinical presentation and severity of the symptoms. Despite great advances in discerning the genetic basis of this group of diseases, the molecular pathways leading to symptoms are not yet fully understood. Unravelling these pathways by molecular analysis of the structure and in vitro assessment of functional properties of the human proteins involved, combined with genetic models in lower organisms, should pave the way for specific cures for inherited skin fragility.

Animals↗
PubMed2006

Kindlin-1 is a phosphoprotein involved in regulation of polarity, proliferation, and motility of epidermal keratinocytes.

A novel family of focal adhesion proteins, the kindlins, is involved in attachment of the actin cytoskeleton to the plasma membrane and in integrin-mediated cellular processes. Deficiency of kindlin-1, as a result of loss-of-function mutations in the KIND1 gene, causes Kindler syndrome, an autosomal recessive genodermatosis characterized by skin blistering, progressive skin atrophy, photosensitivity and, occasionally, carcinogenesis. Here we characterized authentic and recombinantly expressed kindlin-1 and show that it is localized in basal epidermal keratinocytes in a polar fashion, close to the cell surface facing the basement membrane, in the areas between the hemidesmosomes. We identified two forms of kindlin-1 in keratinocytes, with apparent molecular masses of 78 and 74 kDa, corresponding to phosphorylated and desphosphorylated forms of the protein. In kindlin-1-deficient skin, basal keratinocytes show multiple abnormalities: cell polarity is lost, proliferation is strongly reduced, and several cells undergo apoptosis. In vitro, deficiency of kindlin-1 in keratinocytes leads to strongly reduced cell proliferation, decreased adhesion, undirected motility, and intense protrusion activity of the plasma membrane. Taken together, these results show that kindlin-1 plays a role in keratinocyte adhesion, polarization, proliferation, and migration. It is involved in organization and anchorage of the actin cytoskeleton to integrin-associated signaling platforms.

Animals↗
PubMed2006

Molecular and diagnostic aspects of genetic skin fragility.

Genetic syndromes with skin fragility represent a heterogeneous group of very rare disorders caused by mutations in genes encoding proteins or protein subunits important for the mechanical resistance of keratinocytes and for cell-cell or cell-extracellular matrix adhesion. The common symptoms are skin blistering or peeling, with various degrees of severity and distribution, ranging from localized to generalized forms. Associated features include involvement of skin annexes, mucous membranes, teeth, muscles or the digestive tract. Morphological investigation of skin samples provides evidence for the tissue level of blister formation, while immunostainings may reveal defective proteins, providing clues concerning the genetic origin of the disease. Extensive mutation analysis and subsequent identification of new gene defects provide accurate diagnostics, and lead to better understanding of the functions of the respective proteins, with the potential for new therapeutic strategies.

Cell Adhesion↗
PubMed2006

C-terminal truncation impairs glycosylation of transmembrane collagen XVII and leads to intracellular accumulation.

Collagen XVII, a type II transmembrane protein in hemidesmosomes, is involved in the anchorage of stratified epithelia to the underlying mesenchyme. Its functions are regulated by ectodomain shedding, and its genetic defects lead to epidermal detachment in junctional epidermolysis bullosa (JEB), a heritable skin fragility syndrome, but the molecular disease mechanisms remain elusive. Here we used a spontaneously occurring homozygous COL17A1 deletion mutant in JEB to discern glycosylation of collagen XVII. The mutation truncated the distal ectodomain and positioned the only N-glycosylation site 34 amino acids from the newly formed C terminus, which impaired efficient N-glycosylation. Immunofluorescence staining of authentic JEB keratinocytes and of COS-7 cells transfected with the mutant indicated intracellular accumulation of collagen XVII precursor molecules. Cell surface biotinylation and quantification of ectodomain shedding demonstrated that only about 15% of the truncated collagen XVII reached the cell surface. The cell surface-associated molecules were N-glycosylated in a normal manner, in contrast to the molecules retained within the cells, indicating that N-glycosylation of the ectodomain is required for targeting of collagen XVII to the plasma membrane and that reduced accessibility of the N-glycosylation site negatively regulates this process. Functional consequences of the strong reduction of collagen XVII on the cell surface included scattered deposition of cell adhesion molecule laminin 5 into the extracellular environment and, as a consequence of faulty collagen XVII-laminin ligand interactions, aberrant motility of the mutant cells.

Amino Acid Sequence↗
PubMed2006

Molecular basis of Kindler syndrome in Italy: novel and recurrent Alu/Alu recombination, splice site, nonsense, and frameshift mutations in the KIND1 gene.

Kindler syndrome (KS) is a rare autosomal recessive disorder characterized by skin blistering in childhood followed by photosensitivity and progressive poikiloderma. Most cases of KS result from mutations in the KIND1 gene encoding kindlin-1, a component of focal adhesions in keratinocytes. Here, we report novel and recurrent KIND1 gene mutations in nine unrelated Italian KS individuals. A novel genomic deletion of approximately 3.9 kb was identified in four patients originating from the same Italian region. This mutation deletes exons 10 and 11 from the KIND1 mRNA leading to a truncated kindlin-1. The deletion breakpoint was embedded in AluSx repeats, specifically in identical 30-bp sequences, suggesting Alu-mediated homologous recombination as the pathogenic mechanism. KIND1 haplotype analysis demonstrated that patients with this large deletion were ancestrally related. Five additional mutations were disclosed, two of which were novel. To date, four recurrent mutations have been identified in Italian patients accounting for approximately approximately 75% of KS alleles in this population. The abundance of repetitive elements in intronic regions of KIND1, together with the identification of a large deletion, suggests that genomic rearrangements could be responsible for a significant proportion of KS cases. This finding has implications for optimal KIND1 mutational screening in KS individuals.

Adolescent↗
PubMed2006

Collagen XVII is expressed in human CNS neurons.

Type XVII collagen (collagen XVII) is a component of hemidesmosomes, which connect epithelial cells to the underlying basement membrane. Previously, an association has been suggested between neurological disorders and the skin disease bullous pemphigoid, where autoimmunity is directed against collagen XVII. Furthermore, the lack of alpha6 integrin, a ligand of collagen XVII, has been implicated in defects of cortical organization in the mouse brain. In this study, we demonstrate for the first time the presence of collagen XVII in neurons of the human brain by in situ hybridisation, immunohistochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR). We propose that collagen XVII may be involved in the pathogenesis of various disorders affecting neuronal migration or synaptic plasticity.

Adult↗
PubMed2006

Novel and recurrent mutations in keratin KRT5 and KRT14 genes in epidermolysis bullosa simplex: implications for disease phenotype and keratin filament assembly.

Epidermolysis bullosa simplex (EBS) is a group of autosomal dominant genetic skin disorders caused by mutations of the keratin genes KRT5 and KRT14. It is characterised by lysis of basal keratinocytes leading to the development of intraepidermal blisters upon minor mechanical trauma. We investigated 27 EBS patients and families of mainly German origin by sequence analysis of the entire coding sequences of KRT5 and KRT14 and identified 12 novel and seven previously reported mutations within the KRT5 and KRT14 genes. The study discusses possible implications of the novel mutations on protein structure, keratin intermediate filament (KIF) formation and the corresponding phenotype, and summarises the spectrum of mutations reported so far in EBS. Detailed knowledge of the spectrum of EBS mutations and their genotype-phenotype correlation is essential for accurate genetic counselling and prenatal diagnosis.

DNA Mutational Analysis↗
PubMed2006

Expanding the COL7A1 mutation database: novel and recurrent mutations and unusual genotype-phenotype constellations in 41 patients with dystrophic epidermolysis bullosa.

Dystrophic epidermolysis bullosa (DEB), a heterogeneous hereditary skin disorder characterized by trauma-induced blistering and scarring, affects thousands of families worldwide. The clinical manifestations extend from minor nail dystrophy to severe life-threatening blistering, making early molecular diagnosis and prognostication of utmost importance for the affected families. DEB is caused by mutations in the COL7A1 gene encoding collagen VII in the skin. Molecular diagnostics and genotype-phenotype correlations in DEB remain complex owing to the gene structure, large variety of mutations, high rate of novel mutations, complex protein structure and assembly, and the heterogeneity of phenotypes. Here, we report an efficient strategy for COL7A1 mutation detection using direct automated DNA sequencing and implementation of software tools. With this approach, COL7A1 mutations of 41 DEB families were disclosed. Twenty-four mutations were novel and two recurrent. Elucidation of biological consequences of the mutations helped define disease mechanisms, but also revealed several unusual genotypic and/or phenotypic constellations, which impeded the diagnostics and prognostication. In addition, the studies disclosed a de novo mutation in recessive DEB and two new polymorphisms in the COL7A1 gene.

Collagen Type VII↗
PubMed2006

Lupus vulgaris--a case diagnosed more than 20 years after onset.

A 47-year-old Turkish woman developed an erythematous scaly plaque on her thigh over 20 years. Cutaneous sarcoidosis had been diagnosed eight years previously on a skin biopsy. We confirmed Mycobacterium tuberculosis infection by polymerase chain reaction and cultivation of the causative organism from lesional skin. After several months of antituberculous treatment with isoniazid, rifampicin, ethambutol and pyrazinamide, the lesion resolved.

Antitubercular Agents↗
PubMed2006

Tight junctions and compression therapy in chronic venous insufficiency.

Tight junctions (TJs) provide a barrier function, inhibiting solute and water flow through the paracellular space. There had been no analysis until now as to how tight junction molecules could be involved in the pathology of patients with chronic venous insufficiency. The aim of the study was to analyse the expression pattern of TJ-molecules occludin (OCLN), claudin-1 (CLDN-1), claudin-3 (CLDN-3) and claudin-5 (CLDN-5) on mRNA and protein level in patients with edema, venous leg ulcers and healthy controls. Biopsy specimens were taken in healthy individuals and in patients before, and four weeks after compression therapy. mRNA-expression was determined by using reverse-transcriptase and polymerase chain reaction (RT-PCR) and the protein-expression was determined by Western blotting from tissue specimens. Quantification performed determining the expression for TJ-molecules displayed diminished expression for CLDN-1 (p<0.01) and CLDN-5 (p<0.01) in patients with chronic venous insufficiency in comparison with healthy controls on mRNA as well as protein level. No statistical differences were detected for OCLN and CLDN-3 between the edema group and healthy controls. There was a significantly elevated expression (p<0.01) on mRNA and protein level between the leg ulcer group and healthy controls for OCLN and CLDN-3. Densitometric evaluation revealed a more significantly elevated expression (p<0.01) for CLDN-1 and CLDN-5 on mRNA and protein level after four weeks of compression therapy in comparison with prior to treatment for the edema as well as the leg ulcer group. Compression therapy tightens the paracellular barrier via elevated expression of specific TJs and prevents thereby the progression of chronic venous insufficiency due to inhibited permeability of fluid into the perivascular tissue.

Bandages↗
PubMed2005

Imatinib mesylate as a novel treatment option for hypereosinophilic syndrome: two case reports and a comprehensive review of the literature.

Hypereosinophilic syndromes (HES) are a heterogenous group of rare disorders characterized by sustained and otherwise unexplained overproduction of eosinophils with organ involvement and consecutive dysfunction. Recent reports document the efficacy of imatinib mesylate in a large proportion of HES patients (65%). Rearrangements involving the platelet-derived growth factor receptor genes (PDGFRA and PDGFRB), both tyrosine kinase receptors, have been demonstrated to be pathogenetically linked to the dysregulated clonal overproduction of eosinophils. This refined hypothesis has been confirmed by the discovery of the novel FIP1L1-PDGFRA fusion gene, which is a gain-of-function gene on chromosome 4q12. Its product is an imatinib-sensitive tyrosine kinase, which can be found in a subset of patients with HES, particularly in those responding to treatment with imatinib mesylate. Here, we sum up recent knowledge of clinical features, pathophysiology and novel treatment aspects of HES by performing a comprehensive search of the available literature and report on 94 patients. We particularly address the issue of organ involvement and specific characteristics of the variable clinical pictures. In addition, two cases will be presented, which illustrate typical clinical scenarios and treatment outcome.

Adult↗
PubMed2005

Shedding of collagen XVII ectodomain depends on plasma membrane microenvironment.

Collagen XVII, a hemidesmosomal component, mediates the adhesion of epidermal keratinocytes to the underlying basement membrane. It exists as a full-length transmembrane protein and a soluble ectodomain that is proteolytically released from the cell surface by sheddases of a disintegrin and metalloproteinase (ADAM) family; TACE, the tumor necrosis factor-alpha-converting enzyme, is the major physiological proteinase. Because both collagen XVII and the ADAMs are transmembrane proteins, their plasma membrane microenvironment can influence shedding. Lipid rafts, assemblies of sphingolipids and cholesterol within the plasma membrane, are responsible for the separation of membrane proteins and are thought to regulate shedding of cell surface proteins. In this study we analyzed the influence of the cholesterol-depleting agent methyl-beta-cyclodextrin (MbetaCD), which disintegrates lipid rafts, on the shedding of collagen XVII in HaCaT keratinocytes and in transfected COS-7 cells. Increasing concentrations of MbetaCD led to a dose-dependent decrease of membrane cholesterol levels and to stimulation of collagen XVII shedding. The stimulation was completely inhibited by sheddase inhibitors, and experiments with COS-7 cells co-transfected with TACE and collagen XVII demonstrated that TACE mediated the low cholesterol-dependent shedding. Co-patching analysis by double immunofluorescence staining revealed co-localization of collagen XVII with the raft resident phosphatidylinositol-linked placental alkaline phosphatase and segregation from the non-raft protein human transferrin receptor, indicating that a majority of collagen XVII molecules was incorporated into lipid rafts. These data deliver the first evidence for the role of plasma membrane lipid organization in the regulation of collagen XVII shedding and, therefore, in the regulation of keratinocyte migration and differentiation.

Animals↗
PubMed2005

Collagen XVII and BPAG1 expression in the retina: evidence for an anchoring complex in the central nervous system.

The ectoderm gives rise not only to the skin but also to the entire CNS. This common embryonic lineage suggests that some molecular isoforms might serve analogous functions in both tissues. Indeed, not only are laminins important components of dermal adhesion mechanisms, but they also regulate some aspects of synaptic development in both the CNS and the PNS. In the skin, laminins are part of a hemidesmosome complex essential for basal keratinocyte adhesion that includes collagen XVII (BP180) and BPAG1 (dystonin/BP230). Here, we show that CNS neurons also express collagen XVII and BPAG1 and that these molecules are expressed in the adult and developing retina. In the retina, isoforms of collagen XVII and BPAG1 are colocalized with laminins at photoreceptor synapses and around photoreceptor outer segments; both molecules are expressed by rods, whereas cones express collagen XVII but not BPAG1. Moreover, biochemical data demonstrate that collagen XVII complexes with retinal laminins. We propose that collagen XVII and BPAG1 isoforms may help to anchor elements of the rod photoreceptor cytomatrix to the extracellular matrix.

Animals↗
PubMed2005

Substrate-specific modulation of a multisubstrate proteinase. C-terminal processing of fibrillar procollagens is the only BMP-1-dependent activity to be enhanced by PCPE-1.

Members of the bone morphogenetic protein-1/tolloid (BMP-1/Tld) family of metalloproteinases, also known as procollagen C-proteinases (PCPs), control multiple biological events (including matrix assembly, cross-linking, cell adhesion/migration and pattern formation) through enzymatic processing of several extracellular substrates. PCP activities on fibrillar procollagens can be stimulated by another family of extracellular proteins, PCP enhancers (PCPE-1, PCPE-2), which lack intrinsic enzymatic activity. While PCPs have multiple substrates, the extent to which PCPEs is involved in the processing of proteins other than fibrillar procollagens is unknown. In the experiments reported here, PCPE-1 was found to have no effect on the in vitro BMP-1 processing of procollagen VII, the procollagen V N-propeptide, the laminin 5 gamma2 chain, osteoglycin, prolysyl oxidase, or chordin. In contrast, PCPE-1 enhanced C-terminal processing of human fibrillar procollagen III but only when this substrate was in its native, disulfide-bonded conformation. Surprisingly, processing of procollagen III continued to be enhanced when essentially all the triple-helical region was removed. These and previous results (Ricard-Blum, S., Bernocco, S., Font, B., Moali, C., Eichenberger, D., Farjanel, J., Burchardt, E. R., van der Rest, M., Kessler, E., and Hulmes, D. J. S. (2002) J. Biol. Chem. 277, 33864-33869; Bernocco, S., Steiglitz, B. M., Svergun, D. I., Petoukhov, M. V., Ruggiero, F., Ricard-Blum, S., Ebel, C., Geourjon, C., Deleage, G., Font, B., Eichenberger, D., Greenspan, D. S., and Hulmes, D. J. S. (2003) J. Biol. Chem. 278, 7199-7205) indicate that the mechanism of PCPE-1 action involves recognition sites in both the C-propeptide domain and in the C-telopeptide region of the procollagen molecule. PCPEs therefore define a new class of extracellular adaptor proteins that stimulate proteinase activity in a substrate-specific manner, thereby providing a new target for the selective regulation of PCP activity on fibrillar procollagen substrates.

Amino Acid Sequence↗