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K Scharffetter

Publications and source records attributed to K Scharffetter.

18 recordsLinked to original sources
PubMed1992

Occlusive microangiopathy by immunoglobulin (IgM-kappa) precipitation: pathogenetic relevance in paraneoplastic cryoglobulinemic neuropathy.

Histological, immunohistochemical and ultrastructural sural nerve and skin biopsy findings in a case of cryoglobulinemia secondary to an IgM-kappa-producing non-Hodgkin lymphoma are described. The main finding was an occlusive microangiopathy present in both the sural nerve and the skin. Widespread cryoglobulin deposits of the proliferated vasa nervorum were associated with pronounced changes probably evoked by ischemia. Moderate perivascular inflammation, but no florid vasculitis was additionally present. Our observations indicate that occlusive microangiopathy by precipitated cryoglobulins may be a relevant pathogenetic factor in cryoglobulinemic peripheral neuropathy.

B-Lymphocytes↗
PubMed1991

UVA irradiation induces collagenase in human dermal fibroblasts in vitro and in vivo.

We report the effect of UVA irradiation on collagen metabolism of fibroblasts, including both synthesis of the collagen degrading enzyme collagenase and de novo synthesis of type I collagen as the major structural component of the dermis. For this purpose confluent fibroblast monolayers were irradiated under standardized conditions (5, 15, 35, 60 J/cm2 using UVASUN 3000, Mutzhas, Munich, FRG, and UV source Sellas sunlight type 2.001, Sellas, Gevelsberg, FRG). Subsequently, total RNA was isolated and subjected to dot blot and northern blot analysis using oligolabelled cDNA clones for human type I collagen, collagenase and beta-actin. Collagen type I and beta-actin mRNA levels remained unaltered following irradiation, suggesting that the synthetic pathway of collagen metabolism at the pretranslational level is not affected by short-term UVA irradiation. However, collagenase mRNA was found to be dose-dependently induced in fibroblasts after irradiation, thus probably contributing to the actinic damage to the dermis. These in vitro data were confirmed in vivo using in situ hybridization on frozen sections of biopsy material obtained from UVA irradiated patients.

Cell Division↗
PubMed1991

[Polymerase chain reaction with subsequent direct gene sequence analysis. A possibility for molecular analysis in dermatology].

Molecular analyses are of increasing importance in clinical research as well as for diagnostic evaluations. Gene alterations are involved in the pathogenesis of numerous diseases and are described in an increasing number of conditions. A method is presented which allows for rapid and detailed nucleotide sequence analysis of a segment of a gene. Buccal epithelial cells of a patient are obtained by means of a mouth wash with 10 ml water. DNA molecules are amplified in vitro using the polymerase chain reaction, followed by direct sequence analysis of the product. Using this technique, a sequence analysis of a gene segment is possible within 1-2 days. As an example, the analysis of a segment of the gene for uroporphyrinogen decarboxylase of patients with cutanea tarda is described.

Base Sequence↗
PubMed1991

[Polymerase chain reaction--principle and application in medicine].

The polymerase chain reaction, a new molecular technique, is of increasing importance in many areas of medical research and diagnosis. Within hours DNA sequences can be amplified millionfold with very high specificity, making detection and further analysis possible. Using multiple rounds of exponential amplification, even one copy of a gene of interest can be detected by agarose gel electrophoresis and other methods. Furthermore, minute amounts of DNA even from tissue damaged by embedding in paraffin can be analysed. Polymerase chain reaction methodology has already gained significance in many areas of medical research, e.g. diagnosis of inherited diseases, detection of viral DNA in clinical samples, cancer research and diagnosis, and characterization of gene expression.

DNA, Viral↗
PubMed1991

[Erythropoietic protoporphyria: synopsis of 20 patients].

The authors present 20 patients (9 men, 11 women) with erythropoietic protoporphyria (EPP). The diagnosis was made on the basis of photosensitivity and porphyrin analysis. The disease first became apparent in the first years of life. The following acute symptoms were induced after exposure to sunlight: pruritus with or without skin changes, burning, pain and erythema, sometimes with petechiae, vesiculation and, in two cases, systemic symptoms. Chronic skin changes included hyalinosis cutis-like skin lesions, scarring, and also petechiae. Phototesting provoked only subjective symptoms, and none of the skin lesions characteristic of EPP could be induced. Postnatal diagnosis was attempted in three newborns, each of whom had one parent with proven EPP, by measuring the porphyrins in erythrocytes of cord blood. In all three normal porphyrin values were determined, and during an observation period of 3 years none has developed EPP. Therapy with carotenoids has yielded good to very good results in two-thirds of the patients. So far, a diagnosis of EPP has been established in 30 patients in Düsseldorf: one has died of liver cirrhosis and another has liver damage.

Adolescent↗
PubMed1990

Tumor necrosis factor alpha induces invasiveness of human skin fibroblasts in vitro.

The chemotactic response of fibroblasts plays an important role during wound healing and fibrosis. Several substances have been found to mediate fibroblast migration in vitro. In the tissue, however, fibroblasts have also the potential to pass through connective tissue barriers following a chemotactic stimulus. Since tumor necrosis factor alpha (TNF alpha) is a cytokine released by mononuclear cells during wound healing, we have been interested in studying its effect on the regulation of fibroblast chemotaxis and invasive migration. TNF alpha did not attract skin fibroblasts nor did it alter their chemotactic response towards other chemoattractants like fibroblast conditioned medium or fibronectin. However, whereas normal skin fibroblasts did not invade a collagen I gel, preincubation of the cells with TNF alpha markedly induced their invasive migration into the gel. This seems to be associated with a specific degradation of type I collagen, because TNF alpha did not promote the invasion of skin fibroblasts through a reconstituted basement membrane (Matrigel).

Adult↗
PubMed1990

Localization of cytoplasmic collagen mRNA in human aortic coarctation: mRNA enhancement in high blood pressure-induced intimal and medial thickening.

Enhanced synthesis and deposition of extracellular matrix components, including collagen, contribute significantly to arteriosclerotic changes in the arterial vessel wall. We localized cells actively synthesizing collagen by hybridizing 35S-labeled RNA probes complementary to type I and III collagen mRNA with cytoplasmic mRNA in frozen sections of surgically removed aortic coarctations. These were chosen as a model for comparing mRNA levels in areas of high blood pressure-induced wall thickening and in unaffected post-stenotic areas. In situ hybridization revealed increased expression of type I and III collagen mRNA in intimal cells and in cells adjacent to the medial-adventitial border in the pre-stenotic part of the coarctation. In contrast, cells of the post-stenotic area showed only a very low signal. No immunohistologically detectable macrophages were seen in the pre-stenotic subendothelial areas where mRNA levels were enhanced. Higher collagen mRNA levels therefore occur in particular regions of high blood pressure-induced arterial wall thickening in the absence of macrophages. The results suggest that in situ hybridization is suitable for detection of locally occurring transcriptional activation of cells for collagens in the vessel wall.

Adolescent↗
PubMed1990

[Scleroderma].

Sclerodermas may occur in two basic forms: localized sleroderma (LSc) and systemic scleroderma (SSc). Pseudoscleroderma as well as overlap syndromes have to be differentiated from these two variants. From the clinical point of view, localized scleroderma can be subdivided into type I = plaque-like LSc (= morphea), type II = linear LSc, and type III = deep LSc. According to the degree of the cutaneous involvement, systemic scleroderma can likewise be classified into type I = sclerodactylia, type II = acrosclerosis, and type III = scleroderma with primary involvement of the trunk (diffuse scleroderma). In LSc, we never find systemic involvement; SSc, in contrast, is almost always associated with Raynaud's phenomenon, changes of the esophagus, as well as an increased titer of antinuclear antibodies (Hep-2 cell test). Only 23% of our patients with LSc showed elevated ANA titers. We present and discuss data of 56 patients with LSc and 52 patients with SSc. Evidence in the literature as well as our own findings suggest that the pathogenesis of LSc is different from that of SSc. The influence of various mediators and cytokines on the collagen metabolism might be regarded as a theoretical approach in order to develop new therapeutic regimens. This is even more important since there is still no efficient mode of treatment for neither localized nor systemic scleroderma.

Antibodies, Antinuclear↗
PubMed1989

An automatic analysis method for in situ hybridization using high-resolution image analysis.

Specific mRNA for alpha 1 (I) collagen has been detected on a cellular level by in situ hybridization using radioactively labelled alpha 1 (I) antisense RNA probes. We here present an automatic, quantitative, evaluation technique for the determination of grain densities using the high-resolution image analysis system IPS KONTRON. The reliability and objectivity of this method were evaluated by comparing the values of grains per cell obtained by conventional and automatic techniques following in situ hybridization with alpha 1 (I) collagen DNA probes in various specimens. The correlation coefficients between conventional and automatic analysis of grain densities were 0.97 for fibroblasts embedded into a three-dimensional collagen gel, 0.94 for dermal fibroblasts in skin obtained from a patient with progressive systemic scleroderma, and 0.90 for fibroblasts in a 2-week-old scar. All correlation coefficients were significant on the P less than 0.0001 level. This new analysis technique therefore allows a more rapid and reliable quantitative evaluation of in situ hybridization and may also be helpful in differentiating between various cell populations characterized by different biosynthetic capacities.

Autoradiography↗
PubMed1989

In situ hybridization--a useful tool for studies on collagen gene expression in cell culture as well as in normal and altered tissue.

We report the application of antisense RNA probes for in situ hybridization to identify collagen type I and type III mRNA synthesizing fibroblasts under in vitro and in vivo conditions in normal and wounded human skin. Non-specific hybridization was excluded by specific distribution patterns of alpha 1(I)- and alpha 1(III) probes in mouse fetuses. In addition, the specificity of hybridization was checked by sense probes, radioactively labelled transcripts of Gemini vectors and a keratin probe. In normal skin weakly activated fibroblasts were sparsely scattered within the dermis, while in wound healing processes mRNA both for alpha 1(I) and for alpha 1(III) was dramatically increased, thus suggesting that collagen synthesis is at least partly regulated at a pretranslational level. In addition, the intensity of the labelling, as defined by image analysis and the distribution pattern of collagen mRNA synthesizing cells, provide strong evidence that wound healing by primary intention starts within the deep dermis.

Adult↗
PubMed1989

Synergistic effect of tumor necrosis factor-alpha and interferon-gamma on collagen synthesis of human skin fibroblasts in vitro.

The effect of tumor necrosis factor-alpha (TNF alpha) and interferon-gamma (IFN gamma) on collagen metabolism by human diploid fibroblasts in confluent monolayer culture was examined. Recombinant TNF alpha reduced collagen mRNA levels 2-fold and stimulated collagenase mRNA levels 5-fold, while recombinant IFN gamma affected only collagen mRNA levels. The combination of TNF alpha (10 ng/ml) and IFN gamma (100 ng/ml) resulted in a much stronger (about 30-fold) reduction of collagen mRNA levels indicating that the two cytokines act synergistically. In contrast no such synergism was observed with respect to collagenase mRNA levels. The effect of TNF alpha and IFN gamma on collagen metabolism reported here indicates a complex interaction of different cytokines in the control of tissue remodeling that occurs during inflammation, repair, or atrophy.

Adult↗
PubMed1989

Proliferation kinetics-study of the growth of keratocysts. Morpho-functional explanation for recurrences.

The mode of growth of the odontogenic keratocyst is of decisive importance in the strategy and radicality of its surgical removal. The high rate of recurrence reported in the literature is striking. Staining with haematoxylin-eosin for histological examination provides only limited information on growth. Serial sections of a keratocyst obtained by fractionated curettage were prepared and the proliferation patterns of epithelium and connective tissue were studied by means of autoradiography and DNA cytophotometry and compared with those of an odontogenic reference cyst of the radicular type. The epithelium of the keratocyst examined clearly showed a higher rate of proliferation than that of the radicular cyst. The connective tissue was also growing very actively with a high rate of proliferation and this growth behaviour is at least partly responsible for the invasive and destructive growth of the keratocyst. In contrast to the radicular cyst, proliferation of the epithelium of the keratocyst is multicentric being simultaneous within the growth centres. As a result the epithelium "buds" with formation of satellite cysts in the alveolar bone and peripheral epithelial islands located in the connective tissue. A case is reported of a keratocyst treated by radical curettage with subsequent freshening of adjacent bone with a bur. Our morphological findings are an important contribution to the present discussion of the various methods of treating keratocysts.

Autoradiography↗
PubMed1989

Localization of collagen alpha 1(I) gene expression during wound healing by in situ hybridization.

The cellular localization of collagen alpha 1(I) chain gene expression during wound healing in rats was investigated using in situ hybridization. Activation of collagen gene expression was first found within fibroblastic cells around the deep layers of the granulation tissue as early as 16 h post wounding. Heavily labeled cells were also detected near the intact wound edge and around hair follicles. At day 6 intense alpha 1(I) collagen gene expression was found within most cells of the granulation tissue and after day 8 most of the activity was localized to cells directly underneath the epidermis. 26 d after the induction of the wounds hardly any alpha 1(I) collagen gene expression could be demonstrated, which indicates a close, time-dependent control of collagen synthesis during repair processes.

Animals↗
PubMed1988

[In situ hybridization--an expansion of dermatologic histopathology].

By means of in-situ-hybridization, we are able to detect specifically the mRNAs of defined proteins at a cellular level in frozen sections. This technique not only makes it possible to characterize the biosynthesis of defined proteins at a pretranslational level but can also identify activated cells in their histological surroundings. We employed this technique in order to detect collagen mRNAs in fibrotic diseases of the skin. Using cDNA clones for collagen type I, we proved that in early stages of scleroderma the fibrosis starts in the deep dermis and the subcutaneous fatty tissue. In addition, the regulation of the collagen synthesis seems to be disturbed at a pretranslational level and accompanied by dramatically increased levels of collagen mRNA in fibroblasts. This technique offers objective parameters for collagen mRNA and may be especially useful to estimate the activity of fibrotic diseases in individual patients as well as during therapeutical trials.

Base Sequence↗
PubMed1988

Regulation of collagen synthesis in fibroblasts within a three-dimensional collagen gel.

Fibroblasts cultivated within a three-dimensional collagen gel display an elongated, spindle-like morphology, reduce their proliferation rate, contact the gel to a very dense tissue, and modify their metabolic activity as compared to monolayer cultures. Collagen synthesis measured as protein-bound hydroxyproline is reduced to 5% of the values found in monolayer culture. The reduction involving type I and type III collagen is due to decreased de novo synthesis and not to enhanced degradation. Dot blot hybridization, Northern blot analysis, and in situ hybridization using collagen I- and III-specific cDNA probes demonstrate that reduced biosynthesis rates are reflected by a marked reduction of pro alpha 1 (I), pro alpha 2 (I), and pro alpha 1 (III) collagen mRNA indicating pretranslational regulation. A similar reduction was observed for actin mRNA whereas levels of tubulin mRNA were similar for fibroblasts in monolayer culture or cultivated within the three-dimensional collagen gels. The data suggest a specific reprogramming of various cellular activities in response to contact with the reconstituted extracellular matrix.

Blotting, Northern↗
PubMed1988

Localization of collagen mRNA in normal and scleroderma skin by in-situ hybridization.

Scleroderma is a fibrotic disease occurring in a localized or systemic form. Disturbed regulation of connective tissue metabolism plays an important role in its pathogenesis. However, until now, most of the data available were obtained from studies of fibroblasts in culture and there is considerable doubt that fibroblasts in a monolayer reflect the in-vivo situation. Using in-situ hybridization with specific antisense RNAs on frozen sections of skin, cells were detected displaying enhanced messenger RNA levels for type I and type III collagen in patients with localized and systemic scleroderma. Activated fibroblastic cells were often located near blood vessels in the deep dermis of patients with early stages of the disease and were mostly surrounded by mononuclear cells. These findings are in agreement with the concept that the interaction of fibroblasts with 'immunocompetent cells' is crucial in the initial activation of connective tissue metabolism in fibrosis.

Adult↗