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Detlef Zillikens

Publications and source records attributed to Detlef Zillikens.

At least 19 recordsLinked to original sources
PubMed2007

Anti-p200 pemphigoid: a novel autoimmune subepidermal blistering disease.

Anti-p200 pemphigoid is a recently defined autoimmune subepidermal blistering disease characterized by circulating and tissue-bound autoantibodies to a 200-kDa protein (p200) of the dermal-epidermal junction (DEJ). This DEJ constituent is thought to be important for adhesion of basal keratinocytes to the underlying dermis. While the exact identity of p200 remains unknown, it has been demonstrated to be immunologically and biochemically distinct from all major autoantigens of the DEJ, including bullous pemphigoid antigens 180 and 230, laminin 1, 5 and 6, alpha6beta4 integrin, and type VII collagen. Clinically, most reported cases present with tense blisters as well as urticarial papules and plaques, closely resembling bullous pemphigoid. Histopathological examination of lesional skin biopsies shows subepidermal split formation and superficial inflammatory infiltrate typically dominated by neutrophils. Immunopathologically, linear deposits of immunoglobulin (Ig)G and C3 are detected along the DEJ by direct immunofluorescence microscopy of perilesional skin. Indirect immunofluorescence microscopy of patients' sera on NaCl-split human skin demonstrates circulating IgG autoantibodies labeling the dermal side of the split. By immunoblotting, these autoantibodies recognize a 200-kDa protein of human dermis. Biochemical characterization of the p200 molecule revealed a noncollagenous N-glycosylated acidic protein with an isoelectric point of approximately 5.5. We present an overview of the pathogenesis, clinical features, diagnosis and treatment of this new disease entity.

Autoantibodies↗
PubMed2006

Inhibition of Rho A activity causes pemphigus skin blistering.

The autoimmune blistering skin diseases pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are mainly caused by autoantibodies against desmosomal cadherins. In this study, we provide evidence that PV-immunoglobulin G (IgG) and PF-IgG induce skin blistering by interference with Rho A signaling. In vitro, pemphigus IgG caused typical hallmarks of pemphigus pathogenesis such as epidermal blistering in human skin, cell dissociation, and loss of desmoglein 1 (Dsg 1)-mediated binding probed by laser tweezers. These changes were accompanied by interference with Rho A activation and reduction of Rho A activity. Pemphigus IgG-triggered keratinocyte dissociation and Rho A inactivation were p38 mitogen-activated protein kinase dependent. Specific activation of Rho A by cytotoxic necrotizing factor-y abolished all pemphigus-triggered effects, including keratin retraction and release of Dsg 3 from the cytoskeleton. These data demonstrate that Rho A is involved in the regulation of desmosomal adhesion, at least in part by maintaining the cytoskeletal anchorage of desmosomal proteins. This may open the possibility of pemphigus treatment with the epidermal application of Rho A agonists.

Blister↗
PubMed2006

Initial Fc epsilon RI-mediated signal strength plays a key role in regulating basophil signaling and deactivation.

BACKGROUND: Mediator releases after high-affinity IgE receptor (Fc(epsilon)RI) cross-linking in basophils and mast cells crucially govern the symptoms of allergic disease and amplify underlying T(H)2-type responses. Interestingly, the dose-response curve for Fc(epsilon)RI activation is bell-shaped, with supraoptimal stimulation leading to reduced mediator release. OBJECTIVE: We sought to characterize the mechanisms responsible for this control of Fc(epsilon)RI-triggered basophil activation. METHODS: Human basophils were purified by means of Ficoll density centrifugation, elutriation, and negative selection with immunomagnetic beads. Various intracellular signal protein activities were assessed by means of Western blotting, and mediator releases were analyzed either spectrofluorometrically (histamine) or by means of ELISA (IL-4 and IL-13). RESULTS: Supraoptimal anti-IgE concentrations led to lower mediator release than optimal concentrations but simultaneously to considerably faster histamine release kinetics. In parallel, basophil signaling proteins (Syk, p38 mitogen-activated protein kinase, and extracellular signal-regulated kinases 1 and 2) were more rapidly phosphorylated at higher anti-IgE concentrations but more transiently activated in the supraoptimal range. This endogenous regulation most likely involved src homology 2 domain-containing inositol 5' phosphatase (SHIP), which was highly phosphorylated after supraoptimal anti-IgE triggering compared with lower stimulus concentrations. Conversely, N-formyl-methionyl-leucyl-phenylalanine-stimulated basophils failed to phosphorylate SHIP in the supraoptimal concentration range and did not display a bell-shaped dose-response curve. CONCLUSION: The kinetics of IgE-mediated signaling and mediator release in primary human Fc(epsilon)RI(+) cells varies substantially according to the magnitude of stimulation, and SHIP most likely plays an important role in terminating these events. CLINICAL IMPLICATIONS: The speed of allergic symptom generation depends on the degree of IgE receptor triggering, which is downregulated by SHIP, a potential target for allergy therapy.

Basophil Degranulation Test↗
PubMed2006

Induction of complement-fixing autoantibodies against type VII collagen results in subepidermal blistering in mice.

Experimental models reproducing an autoimmune response resulting in skin blistering in immunocompetent animals are lacking. Epidermolysis bullosa acquisita (EBA) is a bullous skin disease caused by autoantibodies to type VII collagen. In this study, we describe an active disease model of EBA by immunizing mice of different strains with murine type VII collagen. All mice developed circulating IgG autoantibodies that recognized type VII collagen and bound to the lamina densa of the dermal-epidermal junction. Importantly, subepidermal blisters developed in 82% of SJL-1, 56% of BALB/c mice, and 45% of Fc gammaRIIb-deficient mice, but not in SKH-1 mice. In susceptible animals, deposits of IgG1, IgG2, and complement C3 were detected at the dermal-epidermal junction. In contrast, in the nondiseased mice, tissue-bound autoantibodies were predominantly of the IgG1 subclass and complement activation was weak or absent. This active disease model reproduces in mice the clinical, histopathological, and immunopathological findings in EBA patients. This robust experimental system should greatly facilitate further studies on the pathogenesis of EBA and the development of novel immunomodulatory therapies for this and other autoimmune diseases.

Animals↗
PubMed2006

Localisation of bullous pemphigoid antigen 180 (BP180) in cultured human keratinocytes: functionally relevant modification by calcium.

The expression of BP180 has previously been demonstrated to be influenced by both calcium (Ca(2+)) concentration and binding of anti-BP180-antibodies in cultured keratinocytes of the skin squamous cell carcinoma line DJM-1. Here, BP180 expression was studied in cultured normal human epidermal keratinocytes by confocal laser scanning microscopy. We exploited an experimental system, in which BP180 was previously shown to mediate, upon incubation with anti-BP180 antibodies, a specific signal-transducing event that leads to the release of inflammatory mediators, such as IL-8 from cultured normal human epidermal keratinocytes (NHEK). We found that without addition of BP180-specific IgG, BP180 is predominantly expressed on the cell surface irrespective of the Ca(2+) concentration. In contrast, cell surface BP180 was greatly reduced in NHEK kept in high Ca(2+) medium after incubation with BP180-specific IgG for 12 h compared to low Ca(2+) medium. This effect was seen with antibodies to both N- and C-terminal fragments of the BP180 ectodomain, respectively. In addition, a slightly higher BP180 expression was found in NHEK cultured in low compared to high Ca(2+) medium by Western blotting. Interestingly, in contrast to NHEK kept under low Ca(2+ )conditions, in NHEK grown in high Ca(2+) medium, no elevated levels of IL-8 were released after treatment of cells with anti-BP180 IgG compared to normal IgG. Our data indicate that the Ca(2+)-modulated expression of BP180 is functionally relevant. This finding sheds further light on the complex pathomechanism in blister formation of BP180-related autoimmune blistering skin diseases.

Animals↗
PubMed2006

Immunoadsorption against two distinct epitopes on human type XVII collagen abolishes dermal-epidermal separation induced in vitro by autoantibodies from pemphigoid gestationis patients.

Pemphigoid gestationis (PG) is a subepidermal autoimmune blistering disease characterized by self-reactive T and B cells specific for the transmembrane hemidesmosomal protein type XVII collagen/BP180. Major T and B cell epitopes are located within the immunodominant 16th non-collagenous domain A (NC16A) of type XVII collagen. The aim of the present study was to map the pathogenically relevant epitopes targeted by blister-inducing patients' autoantibodies. For this purpose, we used an in vitro model of autoantibody-induced leukocyte-dependent dermal-epidermal separation. Pre-adsorption against a recombinant form of the NC16A region abolished the blister-inducing potential of autoantibodies from all PG patients. Using overlapping synthetic peptides, we demonstrated that PG autoantibodies bind to two defined epitopes within the NC16A region (aa 500-514 and aa 511-523). Importantly, pre-adsorption using an affinity matrix containing these epitopes completely abolished dermal-epidermal separation induced by PG autoantibodies. This study identifies the epitopes relevant for blister induction in PG and should facilitate the development of an antigen-specific immunoadsorption therapy for this disease.

Antibody Specificity↗
PubMed2006

Scarring autoimmune bullous disease in a Ugandan patient with autoantibodies to BP180, BP230, and laminin 5.

We report on a 24-year-old, male Ugandan patient with a 2-week history of itchy papules, vesicles, erosions, and crusts distributed on the entire body, accompanied by minor erosions on the palate, tongue, and lower lip. Conjunctivae and genital mucosa were not involved. Circulating IgG and IgA autoantibodies were found against recombinant full-length BP180, BP180 4575, and the C-terminus of BP230. In addition, IgG reactivity was observed against the 16th noncollagenous region of the BP180 ectodomain, the cell-derived soluble ectodomain of BP180 (linear IgA disease antigen 1), and the alpha3 and gamma2 chains of laminin 5. No reactivity was detected with type VII collagen, alpha6beta4 integrin, and the p200 protein. Oral prednisolone and dapsone led to clearance of lesions that mostly healed with scarring and milia formation. Here, we describe a scarring mucocutaneous variant of an autoimmune blistering skin disorder that extends the current clinical and immunopathologic spectrum of this group of diseases.

Adult↗
PubMed2006

Generation and characterization of monoclonal antibodies against the intracellular domain of hemidesmosomal type XVII collagen.

Type XVII collagen, also referred to as bullous pemphigoid antigen 2 (BPAG2) or bullous pemphigoid antigen 180 (BP180), is a transmembrane protein of the hemidesmosomal complexes of keratinocytes. Type XVII collagen has an unusual type II orientation with its N-terminus intracellularly located and with a large extracellular domain that spans lamina lucida of the dermal-epidermal junction. Type XVII collagen is an autoantigen in patients with pemphigoid diseases and its gene is mutated in patients with junctional epidermolysis bullosa. In the present work, we generated new monoclonal antibodies (MAbs) against the intracellular domain of type XVII collagen. We further characterized reactivity and fine specificity of an MAb (clone V58) from this panel of antibodies. The epitope recognized by the mAb V58 was mapped to a stretch of type XVII collagen corresponding to residues 234-398 of its sequence. Possible applications of this new MAb include antigen mapping in patients with hereditary epidermolysis bullosa and immunoaffinity purification of cell-derived type XVII collagen.

Animals↗
PubMed2006

Autoimmune subepidermal blistering diseases in Uganda: correlation of autoantibody class with age of patients.

BACKGROUND: No data are available on the incidence and immunoreactivity of autoimmune subepidermal blistering skin diseases in East Africa. METHODS: All patients with frank blisters/erosions on the skin and/or mucous membranes that attended the Department of Dermatology at Mbarara University, Uganda, from May 2000 to June 2002, were investigated. The diagnosis was based on the clinical presentation and on the presence of circulating autoantibodies detected by indirect immunofluorescence microscopy on 1 m NaCl-split human skin and by Western blotting of recombinant and cell-derived forms of BP180, BP230, and type VII collagen. RESULTS: Twenty-two patients with autoimmune subepidermal blistering skin disorders were identified, including nine with bullous pemphigoid (41%), four with linear immunoglobulin A (IgA) disease (18%), three with mucous membrane pemphigoid (14%), two with linear IgG/IgA bullous dermatosis (9%), and one each with cicatricial pemphigoid and epidermolysis bullosa acquisita (5%). In addition, two patients with immunoreactivity to both the epidermal and dermal side of salt-split skin by indirect immunofluorescence microscopy, who were unreactive to type VII collagen, were provisionally diagnosed as "mixed pemphigoid" (9%). In patients with subepidermal blistering diseases, IgG reactivity correlated significantly with old age, whereas younger patients preferentially developed IgA autoantibodies (P = 0.024). CONCLUSIONS: The age of patients with autoimmune subepidermal blistering diseases appears to influence the immunoglobulin class of autoantibodies. The high frequency of IgA autoantibodies in Ugandan patients may be explained by the age distribution of the Ugandan population.

Adolescent↗
PubMed2006

The 21st century renaissance of the basophil? Current insights into its role in allergic responses and innate immunity.

Basophils and mast cells express all the three subchains of the high-affinity immunoglobulin E (IgE) receptor Fc epsilon RI and contain preformed histamine in the cytoplasmic granules. However, it is increasingly clear that these cells play distinct roles in allergic inflammatory disease. Despite their presence throughout much of the animal kingdom, the physiological function of basophils remains obscure. As rodent mast cells are more numerous than basophils, and generate an assortment of inflammatory cytokines, basophils have often been regarded as minor players in allergic inflammation. In humans, however, basophils are the prime early producers of interleukin (IL)-4 and IL-13, T helper (Th)2-type cytokines crucial for initiating and maintaining allergic responses. Basophils also express CD40 ligand which, in combination with IL-4 and IL-13, facilitates IgE class switching in B cells. They are the main cellular source for early IL-4 production, which is vital for the development of Th2 responses. The localization of basophils in various tissues affected by allergic inflammation has now been clearly demonstrated by using specific staining techniques and the new research is shedding light on their selective recruitment to the tissues. Finally, recent studies have shown that basophil activation is not restricted to antigen-specific IgE crosslinking, but can be caused in non-sensitized individuals by a growing list of parasitic antigens, lectins and viral superantigens, binding to non-specific IgE antibodies. This, together with novel IgE-independent routes of activation, imparts important new insights into the potential role of basophils in both adaptive and innate immunity.

Animals↗
PubMed2006

[Anti-laminin 5 mucous membrane pemphigoid].

A 70-year-old patient presented erosions on the oral mucosa and skin, predominantly in perianal and inguinal areas. Histopathologically, subepidermal blistering was observed. By direct immunofluorescence microscopy, linear deposits of IgG and C(3) were seen at the dermal-epidermal junction. By indirect immunofluorescence microscopy, binding of IgG autoantibodies to the dermal side of NaCl-split skin was detected. These autoantibodies were shown to target both the non-processed and processed alpha(3)- as well as the beta(3)-subunit of laminin 5 by immunoblot analysis using extracellular matrix of cultured human keratinocytes. Based on these results, the diagnosis of an anti-laminin 5 mucous membrane pemphigoid was established. Oral treatment with dapsone, combined with intravenous dexamethasone pulse therapy, led to rapid improvement.

Aged↗
PubMed2006

IgA pemphigus--occurrence of anti-desmocollin 1 and anti-desmoglein 1 antibody reactivity in an individual patient.

BACKGROUND: IgA pemphigus is a rare pustular autoimmune disease with exclusive IgA anti-keratinocyte cell surface antibody reactivity. Two subtypes have been discerned: in the subcorneal pustular dermatosis type, desmocollin 1 has been identified as a targeted autoantigen, while in few cases of the intraepidermal neutrophilic type, IgA anti-desmoglein 1 or IgA anti-desmoglein 3 reactivity has been demonstrated. PATIENTS AND METHODS: A 48-year-old white male presented with generalized large confluent pustules. Skin pathology was assessed by histology and direct immunofluorescence analysis. IgG/IgA autoantibodies against desmoglein 1/3 and desmocollin 1 were measured by ELISA and indirect immunofluorescence using desmocollin 1 cDNA-transfected COS7 cells, respectively. RESULTS: Histopathology revealed subcorneal pustules and direct immunofluorescence microscopy exclusively showed in vivo bound IgA with an intercellular pattern in the epidermis. Desmocollin 1 was identified as a target of IgA autoantibodies by indirect immunofluorescence microscopy utilizing desmocollin 1 cDNA-transfected COS7 cells. In addition, IgA anti-desmoglein 1 reactivity was demonstrated by ELISA. Neither IgA anti-desmoglein 3 nor IgG anti-desmoglein 1/3 autoantibodies were present. CONCLUSIONS: Both desmocollin 1 and desmoglein 1 were autoantigens in this patient with IgA pemphigus and a distinct clinical presentation. To our knowledge, this is the first IgA pemphigus case with dual autoantibody reactivity.

Anti-Bacterial Agents↗
PubMed2005

Pemphigus foliaceus IgG causes dissociation of desmoglein 1-containing junctions without blocking desmoglein 1 transinteraction.

Autoantibodies against the epidermal desmosomal cadherins desmoglein 1 (Dsg1) and Dsg3 have been shown to cause severe to lethal skin blistering clinically defined as pemphigus foliaceus (PF) and pemphigus vulgaris (PV). It is unknown whether antibody-induced dissociation of keratinocytes is caused by direct inhibition of Dsg1 transinteraction or by secondary cellular responses. Here we show in an in vitro system that IgGs purified from PF patient sera caused cellular dissociation of cultured human keratinocytes as well as significant release of Dsg1-coated microbeads attached to Dsg-containing sites on the keratinocyte cellular surface. However, cell dissociation and bead release induced by PF-IgGs was not caused by direct steric hindrance of Dsg1 transinteraction, as demonstrated by single molecule atomic force measurements and by laser trapping of surface-bound Dsg1-coated microbeads. Rather, our experiments strongly indicate that PF-IgG-mediated dissociation events must involve autoantibody-triggered cellular signaling pathways, resulting in destabilization of Dsg1-based adhesive sites and desmosomes.

Autoantibodies↗
PubMed2005

Differential role for mitogen-activated protein kinases in IgE-dependent signaling in human peripheral blood basophils: in contrast to p38 MAPK, c-Jun N-terminal kinase is poorly expressed and does not appear to control mediator release.

BACKGROUND: Exposure of human basophils to allergens results in a rapid secretion of histamine, LTC(4), IL-4 and IL-13, which dominate both the symptomology of allergic diseases and support the underlying Th2/IgE predominance associated with these reactions. The IgE-dependent release of these mediators in basophils crucially involves PI 3-kinase and the subsequent activation of p38 MAPK and ERK1&2. Here, we investigated the role of the third major member of the mitogen activated kinase family, namely the c-Jun amino terminal kinase (JNK), which is rapidly activated following IgE receptor cross-linking in murine mast cells. METHODS: Human basophils were highly purified by magnetic cell sorting. The activities of various intracellular signaling components, in basophils that had been stimulated under various conditions, were assessed by Western blotting. Mediator secretions were also determined using either spectrofluorometric analysis (histamine) or ELISA (LTC(4), IL-4 and IL-13). RESULTS: Our results show that while JNK is moderately expressed in human basophils, it is not consistently phosphorylated upon anti-IgE stimulation. Phosphorylation of the transcription factor c-Jun, a downstream target of JNK, was also undetected in contrast to p38 MAPK and ERK1&2, which were clearly activated following anti-IgE stimulation of the cells. Additionally, inhibitors of the JNK pathway failed to prevent basophil mediator release and had no effect on the phosphorylation of p38 MAPK or ERK1&2 at concentrations which were specific for JNK blockade. CONCLUSIONS: These data suggest major differences in utilizing various members of the mitogen-activated kinase family in the signal transduction cascade of IgE-receptor-bearing cells.

Basophils↗
PubMed2005

Nerve growth factor influences IgE-mediated human basophil activation: functional properties and intracellular mechanisms compared with IL-3.

NGF and IL-3 play a unique role in supporting human basophil differentiation and mediator secretion. Their importance in allergic disease is underlined further by studies showing elevated levels of these factors in asthmatics. Here, we compared the abilities of IL-3 and NGF to stimulate basophil histamine, IL-4 or IL-13 release, either directly or in conjunction with IgE-dependent stimulation and assessed the intracellular signals responsible. Our results show that the ability of IL-3 and NGF to enhance IgE-dependent histamine release are similar. Both factors also potentiated IgE-dependent IL-13 secretion to a greater degree than the release of histamine or IL-4. At high concentrations (100 ng/ml), IL-3 and NGF alone were capable of releasing cytokines but little histamine. These abilities of IL-3 and NGF to modulate basophil activation were sensitive to blockade by specific inhibitors of PI 3-kinase, p38 MAPK and PLC, but not PKC, suggesting that their effects are mediated considerably by pathways comparable to IgE-dependent signalling.

Basophils↗