Biomedical subjects

C Klessen

Publications and source records attributed to C Klessen.

At least 19 recordsLinked to original sources

PubMed1996

A computer based, temperature controlled bipolar electrocoagulation system.

OBJECTIVE: To reduce complications during electrocoagulation particularly injury of other organs, a computer based, temperature controlled bipolar coagulation system has been developed. Under this system temperature in the tissue is kept between 60 degrees C and 95 degrees C during coagulation. A real time graphic display provides the surgeon with an actual temperature at the tip of the forceps during coagulation. STUDY DESIGN: Rabbit uterine horns were coagulated at 90 degrees C and were examined histologically and histochemically at 6 weeks after coagulation. RESULTS: Electrocoagulation at 90 degrees C induced a total loss of LDH activity in the tissue. Lumen of the uterus was completely occluded when examined 6 weeks later. CONCLUSION: This system with low temperature is sufficient for homogenous coagulation of the tissue.

Animals↗

PubMed1995

Immunocytochemical and immunoelectron microscopic demonstration of cathepsin B in human malignant melanoma.

Proteases are known to enhance the spread of tumour cells. Possible sources of these proteases are the tumour cells themselves or fibroblasts in the tumour tissue. Immunological staining with anticathepsin B antibody indicates that the subcellular distribution of cathepsin B in tumour cell lines differs from that in normal liver. The aims of this study were: (i) to show whether different types of melanoma differ in their production of cathepsin B; (ii) to identify the cathepsin B-producing cells; and (iii) to determine the subcellular distribution of cathepsin B in melanoma cells. All types of melanomas contained cell regions stained with anticathepsin B antibody. The intensity of the stain and the number of cells reacting with anticathepsin B antibody depended on the size of the tumour but not on the type of melanoma. Epithelioid cells stained more intensely with anticathepsin B antibody than spindle-shaped cells. Cells staining with anticathepsin B antibody were almost exclusively tumour cells. Anticathepsin B stain was located mainly in vesicular structures which did not contain a filamentous matrix. Additional anticathepsin B stain was detected at the extracellular spaces. Hypomelanotic melanoma cells, mainly of the epithelioid type, produced most of the cathepsin B. Cathepsin B may be involved in both the degradation of possibly abnormal melanosomes and the focal degradation of the extracellular matrix.

Antigens, Neoplasm↗

PubMed1995

Aminopeptidase M and dipeptidyl peptidase IV activity in epithelial skin tumors: a histochemical study.

The activities of microsomal alanylaminopeptidase (APM EC 3.4.11.2) and of dipeptidyl dipeptidase IV (DPP IV EC 3.4.14.5) were histochemically studied in frozen sections of normal skin, seborrheic keratosis, basal cell carcinoma, solar keratosis, Bowen's disease and squamous cell carcinoma using amino acid- or peptide-4-methoxy-2-naphthylamides as specific chromogenic substrates. Compared to biochemical and immunohistochemical methods, the histochemical technique used in this study allows distinct localization of protease activity within the tumor tissue and the tumor-associated stroma. Strong APM activity was detectable only in the stroma of basal cell carcinoma, a result which reflects the particular tumor-stroma interaction of this semimalignant tumor. APM activity was not detectable in either healthy epidermis or the tumor parenchyma. Altered activity of DPP IV was found in the tumor cells as well as in the surrounding connective tissue: precancerous dermatoses and basal cell carcinomas had higher levels of DPP IV-activity than normal skin or benign seborrheic keratosis. Poorly differentiated malignant squamous cell carcinomas, however, showed no histochemically detectable DPP IV-activity at all. This result is in line with reports of decreased activity of this enzyme in cases of malignancy.

Aminopeptidases↗

PubMed1993

Proteases of polymorphonuclear neutrophils: influence of different salts and buffers on pericellular lysis.

Polymorphonuclear neutrophils (PMN) contain neutral proteases which are cytochemically detectable on blood smears. The activity of these proteases can be visualized by pericellular lysis, i.e. through a disk like degradation of plasma and erythrocytes around centrally located neutrophils. This degradation is inhibited by protease inhibitors in a concentration-dependent manner. Striking differences in halo formation were found depending on the type of buffer and salt used and change of temperature. A halo formation was obtained in Tris-HCl buffer, pH 7.5 without salt, at room temperature (22 degrees C), and with short incubation periods (30 min). Borate buffer, pH 8.5, showed a comparable halo formation only at 37 degrees C, after addition of salts, and after longer incubation times (180 min). When different alkali chlorides were used, halo size increased with molecular mass. Optimum halo size was achieved at a concentration of 250 mM. Addition of MgCl2 and CaCl2 (250 mM) resulted in a small halo formation at 22 degrees C, but inhibited halo formation at 37 degrees C. The present study shows that the pericellular lysis caused around neutrophils by the release of proteases is affected by a complex interaction of salts, temperature, buffers and thermodynamic aspects, all of which should be evaluated carefully in these kinds of experiments.

Buffers↗

PubMed1993

Immunelectron microscopic localization of cathepsin B in human exocrine glands.

The distribution of the lysosomal enzymes cathepsin B, lysozyme, chymotrypsin, and neutrophil elastase was examined in eccrine, apocrine, and sebaceous glands using a postembedding immunogold labeling procedure. Various amounts of cathepsin B were detected in all glands. Lysozyme, however, was detected in apocrine glands only. The other two lysosomal enzymes were not detectable immunologically. In apocrine and eccrine glands, anti-cathepsin B antibody labeled all secretory granules. In sebaceous glands, only the peripheral layer of cells showed immunological activity for cathepsin B. In apocrine glands, granules containing remnants of cristae were more intensively labeled than those lacking cristae which supports the assumption that both granules are derived from mitochondria by acquiring lysosomal enzymes. The enzymes convert mitochondria to granules with cristae and finally to granules without cristae. Thus the difference in morphology is part of a spectrum of the degradation of mitochondria to granules.

Apocrine Glands↗

PubMed1992

Demonstration of proteases in basal cell carcinomas. A histochemical study using amino acid-4-methoxy-2-naphthylamides as chromogenic substrates.

BACKGROUND: Proteases are reported to play an essential part in the proliferative, invasive, and metastasizing behavior of malignant tumors. The aim of the current study was to determine the activity and localization of proteases in basal cell carcinomas (BCC) histochemically. METHODS: Various proteases were identified histochemically in frozen sections of BCC. The following amino acid-4-methoxy-2-naphthylamides (MNA) were used as chromogenic substrates:alanine-MNA for the detection of aminopeptidase M (APM), glycyl-proline-MNA for dipeptidyl peptidase IV (DPP IV), lysyl-proline-MNA and lysyl-alanine-MNA for dipeptidyl peptidase II (DPP II), glycyl-arginine-MNA for dipeptidyl peptidase I (DPP I), and carbobenzoxy (CBZ)-arginyl-arginine-MNA for cathepsin B. RESULTS: APM activity was high in the peritumorous connective tissue, whereas the tumor epithelium and epidermis had negative results. DPP IV showed a highly positive reaction in both tumor epithelium and surrounding connective tissue. Cathepsin B and DPP I reacted strongly in the tumor epithelium but not in the peritumorous connective tissue. CONCLUSIONS: The marked activity of APM, DPP IV, DPP I, and cathepsin B may be related to the proliferation and invasive growth of BCC. The distribution of the activity of APM and DPP IV indicates dynamic interactions between the tumor epithelium and the adjacent connective tissue in the neoplastic process.

2-Naphthylamine↗

PubMed1992

Acetylene, a mammalian metabolite of 1,1,1-trichloroethane.

1,1,1-Trichloroethane (TCE) is a widely used industrial solvent of low acute toxicity. It is slowly oxidized to trichloroethanol and trichloroacetic acid by cytochrome P-450-dependent mono-oxygenases. Increased inhalative uptake by rats under hypoxia and spin-trapping experiments indicate that TCE is also reductively metabolized to a radical intermediate. Acetylene is formed as a metabolite, suggesting transfer of an additional electron to form the corresponding carbene. Hypoxia and induction of mixed-function mono-oxygenases accelerate the formation of acetylene. Experiments performed in vitro with rat liver microsomal fractions yield analogous results.

Acetylene↗

PubMed1991

Protein A-streptokinase fusion protein for immunodetection of specific IgG antibodies.

The streptococcal streptokinase gene truncated at its 5' end was fused to regions of the staphylococcal protein A gene encoding the Fc-binding domains A and B. The resultant fusion gene, when expressed in the Escherichia coli lacPO system or under the speA expression/secretion signals in S. sanguis, specified a bifunctional hybrid protein, SPA-SKC, capable of Fc binding and plasminogen activation. When used in immunoassays designed to titrate antisera raised against bovine chymosin, human serum albumin and fibrinogen, the assay using SPA-SKC compared well with that using a commercial SPA-enzyme conjugate. The simple preparative method together with its efficacy and ease of use, make SPA-SKC a potentially valuable detector reagent in quantitative immunology.

Antibodies, Anti-Idiotypic↗

PubMed1991

Destruction of tumour parenchyma in basal cell carcinoma by tumour-associated neutral proteases: a histochemical study.

Proteolytic activity was demonstrated histochemically in frozen sections of basal cell carcinomas (BCCs). After incubation of tissue sections in 0.1 M phosphate buffer with 0.25 M NaCl the tumour epithelium was almost completely destroyed. The basal and squamous cell layers of the epidermis disintegrated to varying degrees, particularly where they were directly in contact with tumour epithelium. Serine and metalloprotease inhibitors diminished this tissue destruction. Iodoacetate enhanced tumour destruction, urea and potassium thiocyanate even more so. The high proteolytic activity of BCC demonstrated in this study may be an important factor in the proliferative, invasive and destructive behaviour of this tumour.

Basal Cell Carcinoma↗

PubMed1991

[Temperature-controlled bipolar electrocoagulation].

The safety of electrocoagulation for laparoscopic surgery has been greatly improved by the introduction of the bipolar coagulation technique. But complications caused by coagulation, especially by the overheated forceps, are still reported. In order to reduce those complications, a temperature controlled bicoagulation system was developed. This system allows coagulation with lower temperatures which can be kept constant during the coagulation process. In experiments with human fallopian tubes obtained by adnexectomy and rabbit uterine horns it is shown that those lower temperatures are sufficient for homogeneous coagulation. On line monitoring of the temperature during coagulation provides the surgeon with the possibility of directly controlling the actual temperature at the tip of the forceps. The temperature curves can easily be stored in order to be able to demonstrate the particular coagulation process even after years.

Animals↗

PubMed1989

Lectin binding sites in normal and phenobarbitale/halothane treated rat liver. A histochemical study.

The content of carbohydrate residues of both normal and phenobarbitale-halothane-hypoxia exposed rat liver has been examined by means of lectin histochemistry. Eight biotinylated lectins specific to galactose, N-acetyl-galactosamine, N-acetyl-glucosamine, fucose and mannose were applied to paraffin sections of rat liver at light microscopic level. The most distinct binding was observed at the structures of the "perisinusoidal functional unit": Kupffer cells are bound by S-WGA, SBA and PNA. Bile canaliculi display binding sites for RCA I and WGA. Cytoplasm of hepatocytes appears lectin-negative, except for PSA. The enhanced reaction of S-WGA, PNA and SBA after the preincubation of the sections with neuraminidase indicates the occurrence of sialic acid in Kupffer cells. The phenobarbitale-halothane-hypoxia exposed rat liver shows centrolobular degeneration of hepatocytes with a diminished amount of hepatocyte and Kupffer cells as well. The lectin binding pattern of sinusoidal walls, membranes of hepatocytes and bile canaliculi remains the same compared to that of normal rat liver. This finding suggests that at least the carbohydrate content of membranes in the liver resists severe destruction under phenobarbitale-halothane-hypoxia. It is assumed that there exists a connection between intact carbohydrate residues and the regeneration of liver parenchyma.

Animals↗

PubMed1989

Complete secretion of activable bovine prochymosin by genetically engineered L forms of Proteus mirabilis.

To circumvent problems encountered in the synthesis of active chymosin in a number of bacteria and fungi, a recombinant DNA L-form expression system that directed the complete secretion of fully activable prochymosin into the extracellular culture medium was developed. The expression plasmid constructions involved the in-frame fusion of prochymosin cDNA minus codons 1 to 4 to streptococcal pyrogenic exotoxin type A gene (speA') sequences, including the speA promoter, ribosomal binding site, and signal sequence and five codons of mature SpeA. Secretion of fusion prochymosin enzymatically and immunologically indistinguishable from bovine prochymosin was achieved after transformation of two stable protoplast type L-form strains derived from Proteus mirabilis. The secreted proenzyme was converted by autocatalytic processing to chymosin showing milk-clotting activity. In controlled laboratory fermentation processes, a maximum specific rate of activable prochymosin synthesis of 0.57 x 10(-3)/h was determined from the time courses of biomass dry weight and product formation. Yields as high as 40 +/- 10 micrograms/ml were obtained in the cell-free culture fluid of strain L99 carrying a naturally altered expression plasmid of increased segregational stability. The expression-secretion system described may be generally useful for production of recombinant mammalian proteins synthesized intracellularly as aberrantly folded insoluble aggregates.

Animals↗

PubMed1988

Expression of the human interferon-alpha 1 gene under transcriptional and translational control of the speA gene.

The human gene for mature interferon-alpha 1 (IFN-alpha 1) was inserted in a new transcription-translation fusion vector system based on the expression and secretion signals of the gene for type A streptococcal pyrogenic exotoxin, speA. As deduced from the known nucleotide sequences of the component elements, the encoded IFN-alpha 1 was a fusion protein carrying an N-terminal extension of 17 amino acids. When inserted in appropriate vectors capable of replication in Escherichia coli, Bacillus subtilis and Streptococcus sanguis, this expression configuration directed the synthesis of antiviral activity in all 3 organisms, as judged by the cythopathic effect inhibition assay of Vesicular Stomatitis Virus. In E. coli JM101, IFN activity was found mainly in the cytoplasmic protein fraction whereas in the gram-positive hosts, it was completely secreted into the culture medium.

Amino Acid Sequence↗

PubMed1988

Survey of the pSM10 family of streptococcal cloning vectors.

A survey of the pSM10 family of streptococcal plasmid cloning vectors is provided. These vectors are derived from a macrolide-lincosamide-streptogramin B resistance plasmid and include general cloning vectors, shuttle vectors, helper plasmids, expression vectors, gene fusion vectors and plasmids. Besides describing their general properties, this paper also provides their genetic and physical maps.

Cloning, Molecular↗

PubMed1988

Tripartite streptokinase gene fusion vectors for gram-positive and gram-negative procaryotes.

A specific 1,596 bp HincII fragment ('skc) from the chromosome of Streptococcus equisimilis contains an active streptokinase (SK) gene (skc) lacking, in addition to the expression signals, codons 1 through 39 of wild-type skc but retaining the remainder of the skc coding sequence together with the transcription terminator. Using this fragment as an indicator gene, we constructed two types of vectors which in appropriate hosts resulted in the synthesis of SK fusion proteins after insertional activation of 'skc. The first type are open reading frame (ORF) vectors in which 'skc was inserted into pUC18 out of frame with respect to lacZ', thus conferring an SK-negative phenotype. Any DNA fragments representing ORFs inserted between the lacZ' expression signals and 'skc such that the skc reading frame was restored resulted in the production of tripartite proteins which exhibited SK activity. The second type of vector, which functioned in both gram-positive and gram-negative bacteria, used the streptococcal speA expression and secretion signals in front of the ORF to activate 'skc insertionally. Using a large fragment from the chymosin gene as the target sequence, the usefulness of these vectors for studying foreign gene expression in streptococci as well as Escherichia coli was demonstrated.

Amino Acid Sequence↗

PubMed1987

Tetrachloromethane metabolism in vivo under normoxia and hypoxia. Biochemical and histopathological effects relative to alkane exhalation.

About 250 mumol/kg tetrachloromethane is metabolized by male Sprague-Dawley rats receiving a dose of 500 mumol/kg by inhalation. Determination of enzyme activities and histopathological assessment show that various parameters reflecting cellular injury do not differ for the same amount of tetrachloromethane metabolized under different oxygen partial pressures. On the other hand, the amounts of ethane and pentane exhaled under hypoxic conditions are greater than under normoxia. The previously reported differences in the toxicity of tetrachloromethane upon exposure under normoxia or hypoxia seem to be mainly due to the different amounts of tetrachloromethane metabolized under both conditions.

Alkanes↗

PubMed1987

Lectinhistochemical demonstration of galactose- and N-acetyl-D-galactosamine residues in cells of the rat anterior pituitary.

Lectins are a useful tool for identification of differently glycosylated hypophyseal hormones, prohormones and glycoconjugates without hormone function. Beta-D-galactose and beta-N-acetyl-D-galactosamine (GalNAc) containing glycoconjugates were identified by light microscopy with biotinylated lectins in immunocytochemically localized cells of the anterior pituitary of the rat. Galactose, histochemically detectable by the peanut lectin (PNA), was found at penultimate position of the carbohydrate chain after removal of sialic acid. Galactose containing cells correspond to gonadotrophs and thyrotrophs located mainly in medioanterior regions of the pituitary. The lectins from the soybean (SBA) and horse gram (DBA) both specific for GalNAc residues, are bound to round and also polygonal cells corresponding again to gonadotrophs and thyrotrophs.

Acetylgalactosamine↗

PubMed1987

Galactose and fucose binding sites in anterior pituitary cells of the rat. Detection by means of biotinylated lectins.

The oligosaccharide chains of adenohypophyseal glycoprotein hormones fulfill important functions concerning their stability and biological activity. Galactose, histochemically detectable by the Peanut lectin (PNA), could be found after removal of sialic acid in cells situated mainly in the medioanterior region of the pituitary. The Ulex europaeus I lectin (UEA I) reacted with fucose containing sites predominantly in the Golgi-apparatus of nearly all cells. After incubation with neuraminidase, however, fucose may be detected also in the cytoplasm of cells of the medioanterior region. The biological significances of the results is discussed.

Animals↗