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Biomedical subjects

G Pohla-Gubo

Publications and source records attributed to G Pohla-Gubo.

At least 19 recordsLinked to original sources
PubMed2004

Telepathology using immunofluorescence/immunoperoxidase microscopy.

We evaluated low-cost, store-and-forward telepathology interpretation of digital images of skin sections stained immunohistochemically, using immunofluorescence (IF) and immunoperoxidase (IP). The sample comprised 17 patients with skin diseases characterized by cutaneous deposition of immunoglobulins, fibrinogen or complement components. Up to 11 digital IF or IP images (median 3) were transferred via email to centres in Graz, Austria, and Kurume, Japan. Both remote centres had expertise in reading immunohistochemical specimens. Although image files were relatively small (approximately 100 kByte), the IF images were of high quality and they were well suited to static telepathology. There was agreement on the diagnoses made by the local and both remote centres by physicians experienced in IF/IP microscopy in 14 of 17 cases (82%). These results suggest that telepathology evaluation of immunohistochemical specimens may be a useful procedure for the discussion of unusual skin disorders, training purposes and second-opinion consultations on difficult cases from centres of excellence in immunohistochemical diagnosis.

Austria↗
PubMed2003

Characteristic immunohistochemical and ultrastructural findings indicate that Kindler's syndrome is an apoptotic skin disorder.

BACKGROUND: Kindler's syndrome is a rare genodermatosis mainly characterized by the onset of skin blistering in early childhood, web formation of fingers and toes, photosensitivity, and progressive poikiloderma. There is still debate whether this disease represents a distinctive entity in the spectrum of congenital bullous poikilodermas or a variant of dystrophic epidermolysis bullosa. OBJECTIVE: To evaluate the recently proposed and debated characteristic immunohistochemical and ultrastructural features of Kindler's syndrome. PATIENT/METHODS: Immunofluorescence (IF) antigen mapping and transmission electron microscopy (TEM) were performed on a skin specimen from non-sun-exposed inner aspect of the upper arm of a 49-year-old patient with characteristic clinical features of Kindler's syndrome. RESULTS: IF studies revealed focally an extensively broadened, partly reticular staining pattern in the dermoepidermal basement membrane zone (BMZ) with antibodies against laminin-5 and type IV as well as type VII collagen. Anti-alpha6 and beta4 integrin staining revealed small gaps in the linear reactivity in the BMZ. Abundant keratin bodies, as detected by anti-immunoglobulin M (IgM) staining, were focally present in the dermis, indicating prominent epidermal apoptosis. This was verified by a histochemical apoptosis stain [terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) reaction]. Transmission electron microscopic examination showed manifold reduplications of the lamina densa (with attached anchoring fibrils) as well as a keratin body surrounded by a fibroblast in the upper dermis. CONCLUSION: We present characteristic immunohistochemical and ultrastructural features of Kindler's syndrome identical to those described by Shimizu et al. and provide evidence that Kindler's syndrome might primarily be an apoptotic disorder of basal keratinocytes.

Apoptosis↗
PubMed2003

A novel homozygous nonsense deletion/insertion mutation in the keratin 14 gene (Y248X; 744delC/insAG) causes recessive epidermolysis bullosa simplex type Köbner.

We report the sixth case of a human keratin 14 'knockout' mutation resulting in recessive epidermolysis bullosa simplex (EBS). A novel, homozygous nonsense mutation resulting from a deletion/insertion mutation (744delC/insAG) leads to a premature termination codon in the KRT14 gene (Y248X). The patient suffers from generalized cutaneous blistering since birth, mild nail dystrophy, involvement of mucous membranes and multiple epidermolysis bullosa naevi. The clinical variability noted in K14-deficient EBS patients suggests phenotypic modulation by additional genetic and/or epigenetic factors.

Child↗
PubMed2002

[Unilateral blepharochalasis with IgA-deposits].

A 36-year-old male patient presented with unilateral periocular skin atrophy. The blepharochalasis developed without any obvious inflammation of the eyelids over the past 10 years. Interestingly, elongated blood vessels and microaneurysmatic vessel changes were found in the tarsal conjunctiva. A punch biopsy revealed a nearly complete loss of elastic fibres in the papillary and superficial reticular dermis. The contralateral side was histopathologically normal. On immunohistology IgA-deposits could be observed especially on perifollicular elastic fibres. Immunoelectronmicroscopy confirmed the diagnosis and suggested fibulin and fibronectin as potential binding sites for the autoantibodies. This further report of elastolysis in association with IgA-autoantibodies defines the autoantibody binding site in more detail and suggests that the immune mechanisms may also play a role in vessel changes of the conjunctiva.

Adult↗
PubMed2001

A compound heterozygous one amino-acid insertion/nonsense mutation in the plectin gene causes epidermolysis bullosa simplex with plectin deficiency.

Plectin is a cytoskeleton linker protein expressed in a variety of tissues including skin, muscle, and nerves. Mutations in its gene are associated with epidermolysis bullosa simplex with late-onset muscular dystrophy. Whereas in most of these patients the pathogenic events are mediated by nonsense-mediated mRNA decay, the consequences of an in-frame mutation are less clear. We analyzed a patient with compound heterozygosity for a 3-bp insertion at position 1287 leading to the insertion of leucine as well as the missense mutation Q1518X leading to a stop codon. The presence of plectin mRNA was demonstrated by a RNase protection assay. However, a marked reduction of plectin protein was found using immunofluorescence microscopy of the patient's skin and Western blot analysis of the patient's cultured keratinocytes. The loss of plectin protein was associated with morphological alterations in plectin-containing structures of the dermo-epidermal junction, in skeletal muscle, and in nerves as detected by electron microscopy. In an in vitro overlay assay using recombinant plectin peptides spanning exons 2 to 15 the insertion of leucine resulted in markedly increased self-aggregation of plectin peptides. These results describe for the first time the functional consequences of an in-frame insertion mutation in humans.

Base Sequence↗
PubMed1999

Ocular involvement in IgA-epidermolysis bullosa acquisita.

Epidermolysis bullosa acquisita (EBA) is an autoimmune bullous disease with frequent ocular involvement, but visual loss is rare. In contrast, EBA patients with predominant IgA autoantibodies more frequently develop severe ocular involvement, which tends to be refractory to therapy. We report two patients with 'IgA-EBA' with ocular involvement. Both initially presented with a generalized bullous disease, and direct immunofluorescence microscopy demonstrated IgA in the basement membrane zone of the skin, and in the conjunctiva and cornea of patient 1. On salt-split patient skin, IgA was found predominantly on the dermal side of the artificial split in both patients. Direct immunoelectron microscopy demonstrated IgA below the lamina densa in close association with the anchoring fibrils in both patients. In patient 1, who had a prolonged course of the disease, the skin disorder responded well to treatment with cyclosporin, but the ocular involvement ended in bilateral blindness despite repeated surgical treatment. In patient 2, the blister formation and scarring conjunctivitis was stopped by a combination of prednisolone and colchicine. These patients show that in subepithelial blistering diseases, early delineation of disease nosology is critical to detect subtypes with severe ocular involvement such as 'IgA-EBA'. In addition, colchicine may be a valuable alternative in the treatment of EBA with ocular involvement.

Adult↗
PubMed1997

Premature termination codons are present on both alleles of the bullous pemphigoid antigen 2/type XVII collagen gene in five Austrian families with generalized atrophic benign epidermolysis bullosa.

Patients with generalized atrophic benign epidermolysis bullosa (GABEB), an inherited subepidermal blistering disease, often have no immunologically detectable bullous pemphigoid antigen 2 (BPAG2) in their epidermal basement membrane. Recently, we analyzed the BPAG2 gene (GenBank no. M91669) in an Austrian family with GABEB and identified a homozygous deletion mutation, 4003delTC, that results in a downstream premature termination codon (PTC). This mutation has now been identified in additional descendants, suggesting transmission of this mutant allele through at least six generations. Screening of four other Austrian GABEB families revealed that affected members were homozygous for 4003delTC in two cases and heterozygous in two others. In the latter, mutational analysis identified two novel nonsense mutations, Q1403X and G803X, that were confirmed by restriction endonuclease digestions. Thus, PTCs on both alleles of BPAG2 are present in all of these GABEB families. Immunoprecipitation and northern blot studies of cultured keratinocytes from homozygous GABEB patients show that 4003delTC results in undetectable levels of BPAG2 protein and mRNA-findings consistent with the process of nonsense-mediated mRNA decay. Incubating keratinocytes with cycloheximide increased BPAG2 mRNA to a level detectable by northern analysis. When the latter was used in reverse transcription-PCR studies, the mutation was demonstrated, suggesting that cycloheximide may allow mutational analysis in cases where low transcript levels have previously thwarted RT-PCR studies. These findings account for the absence of BPAG2 in GABEB patients and attest to the importance of this protein in adhesion of epidermis to epidermal basement membrane.

Alleles↗
PubMed1996

A homozygous deletion mutation in the gene encoding the 180-kDa bullous pemphigoid antigen (BPAG2) in a family with generalized atrophic benign epidermolysis bullosa.

The 180-kDa bullous pemphigoid antigen (BPAG2) is a candidate gene/protein for mutations in some forms of junctional epidermolysis bullosa. In this study, we searched for mutations in BPAG2 in a large Austrian pedigree with generalized atrophic benign epidermolysis bullosa, a distinct nonlethal form of junctional epidermolysis bullosa, using polymerase chain reaction amplification of genomic DNA, heteroduplex analysis of the polymerase chain reaction products, and direct nucleotide sequencing. We identified a homozygous 2-bp deletion within the coding region of BPAG2 in the affected individuals. This mutation results in a frameshift and downstream stop codons on both alleles, predicting an absence of functional protein. These findings illustrate the molecular basis of the skin fragility in this family and attest to the importance of the 180-kDa bullous pemphigoid antigen in the attachment of the epidermis to the underlying dermoepidermal basement membrane.

Autoantigens↗
PubMed1995

Diminished expression of the extracellular domain of bullous pemphigoid antigen 2 (BPAG2) in the epidermal basement membrane of patients with generalized atrophic benign epidermolysis bullosa.

Generalized atrophic benign epidermolysis bullosa (GABEB) is a nonlethal form of junctional epidermolysis bullosa characterized by generalized skin and mucosal blisters that heal with atrophy; other features include alopecia, nail dystrophy, large melanocytic nevi, and autosomal recessive inheritance. The specific aim of this study was to identify an abnormality in epidermal basement membrane adhesion molecules in well characterized GABEB patients that would explain why these subjects' epidermis separates from their epidermal basement membrane. Cryostat sections of nonlesional skin from 8 GABEB patients in 5 different families as well as skin from normal volunteers (controls) were studied by indirect immunofluorescence microscopy using rabbit antiserum directed against a BPAG1 fusion protein or monoclonal antibodies directed against the extracellular domain of BPAG2 (HD18 and 233), epiligrin (P1E1), laminin 5 (GB3), types IV and VII collagen, or integrin subunits alpha 2, alpha 3, beta 1, alpha 6, or beta 4. In these studies, monoclonal antibodies HD18 and 233 showed no reactivity and diminished reactivity, respectively, to the epidermal BM of all GABEB patients. Interestingly, in one patient, the absent or diminished reactivities of monoclonal anti-BPAG2 antibodies were limited to well demarcated portions of an otherwise intact epidermal basement membrane. Moreover, BPAG1, epiligrin, laminin 5, types IV and VII collagen, and all integrin subunits under study were expressed in the same manner in both GABEB and normal human skin. These findings identify an abnormality in the extracellular domain of BPAG2 in the skin of GABEB patients. BPAG2 (type XVII collagen) is a transmembrane, hemidesmosome-associated molecule whose extracellular domain resides at the exact level where blisters develop in the skin of patients with GABEB.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1992

[Quality control in the allergy laboratory--4 years "Allergy Ring Trial" in Austria].

In 1988 the Austrian Society for Allergology and Immunology initiated an external quality control program for the in vitro allergy diagnosis. In 12 mailings, 62 sera from allergic patients were sent to selected laboratories in order to determine total and antigen specific IgE according to the laboratory-specific methods. The values for total IgE varied considerably (73.9% were within +/- 1 SD, 94.3% within +/- 2 SD and 99.1% within +/- 2 SD, but only 0.9% beyond that). Sources of error were mainly attributable to inappropriate equipment and low quality reagents, but also bad test performance, with respect to personnel and the routines. In contrast, antigen specific IgE against pollen, mites, moulds, insect venoms, animal danders, drugs, parasites, environmental and food antigens revealed quite homogeneous results. Out of 1492 data, only 46 (3.1%) had to be declared as "wrong", and the variability of the RAST-classes was low. Whereas the quality of the reagents from all the different suppliers was not absolutely reliable at the beginning of the study, it improved considerably with time, as consequence of our complaints. The comparability of the methods for detecting total-IgE were non-satisfactory, whereas those for antigen specific IgE were generally good. The variety of methods employed (radio- and enzyme immunologic, fluoro- and nephelometric methods, etc.) should entail appropriate consequences, especially critical comparisons within one and the same laboratory; in addition, international standardization of the "normal" values should replace company-standards, and quality control programs for each test system before it is marketed should be mandatory. Our results confirm, that external control should be obligatory for any laboratory.(ABSTRACT TRUNCATED AT 250 WORDS)

Allergens↗
PubMed1988

Diagnosis of Chlamydia trachomatis infection--culture versus serology.

The diagnostic value of different laboratory methods in detecting Chlamydia trachomatis infections in high risk groups was analysed. The efficiency of a direct specimen test was compared with serology (IgG and IgM ELISA) and culture in L929 cells, stained either with fluorescein conjugated monoclonal antibodies or with iodine. Patients (no. = 1041) with localized genital infections attending a STD clinic, sexual contacts and patients with ascending infections from urological and gynecological clinics were examined. Chlamydia trachomatis was detected in 225 patients: 210 (93.3%) were reactive in the direct test (smears stained with monoclonal antibodies), whereas culture missed only 5 (sensitivity 97.8%) when stained by the same method. Cultures stained with iodine produced the lowest recovery rate (73.8%), but this rate increased to 80.9% when a second passage was performed. In addition the prevalence of Neisseria gonorrhoeae, Mycoplasma hominis, Ureaplasma urealyticum, Candida albicans and Trichomonas vaginalis was investigated. In patients with non-gonococcal urethritis (no. = 331) and cervicitis (no. = 353), Chlamydia trachomatis was isolated in 32.3% and 12.8% respectively. However, this pathogen could be isolated in only 3 (15.8%) out of 19 patients with epididymitis and 15 (14%) out of 107 patients with adnexitis, although 66.7% and 93.3% respectively had specific IgG antibodies. Specific IgM could by detected with a sandwich ELISA in patients with adnexitis (46.7%), epididymitis (33.3%), cervicitis (22.2%), non-gonococcal urethritis (14%) and in the sexual partners of patients with genital infections (35.7%). The direct specimen test with monoclonal antibodies is the method of choice for the diagnosis of a C. trachomatis infection in patients with urethritis and cervicitis.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMedDate not supplied

Ocular involvement in anti-epiligrin cicatricial pemphigoid.

PURPOSE: To report an anti-epiligrin cicatricial pemphigoid (AECP) patient with severe ocular involvement and to provide a practical approach to distinguishing AECP patients from those with other subepidermal blistering diseases. METHODS: Techniques included direct and indirect immunofluorescence microscopy, Western blot and immunoprecipitation studies, as well as interdisciplinary examinations of mucous membranes and skin. RESULTS: This study describes a patient with clinical features of cicatricial pemphigoid, circulating anti-basement membrane zone IgG antibodies, and subepidermal blisters. Histopathology and immunofluorescence analysis suggested the diagnosis of a cicatricial pemphigoid-like type of epidermolysis bullosa acquisita. However, Western blot and immunoprecipitation studies demonstrated that the patient's serum contained autoantibodies against laminin 5 alpha3 subunit, leading to the diagnosis of an AECP. CONCLUSION: Since patients with AECP have an increased relative risk for malignant tumors, it is important to distinguish this entity within the spectrum of cicatricial pemphigoid patients by additional studies such as Western blot or immunoprecipitation.

Autoantibodies↗