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Biomedical subjects

K Weber

Publications and source records attributed to K Weber.

At least 307 records · Page 17Linked to original sources
PubMed1991

Immunocytochemical criteria in the differential diagnosis of malignant melanoma versus carcinoma, lymphoma and sarcoma in fine needle aspirates.

Monoclonal antibodies to keratin, vimentin, leukocyte common antigen (LCA) and S-100 protein have been used in fine needle aspirates of 35 metastatic malignant melanomas, 136 carcinomas, 35 sarcomas and 82 non-Hodgkin's lymphomas in search for immunocytochemical criteria useful in differential diagnosis of melanoma versus carcinoma, non-Hodgkin's lymphoma and sarcoma. All melanomas expressed vimentin and did not express keratin. Six of 14 melanomas contained S-100 protein. All carcinomas were keratin positive. Some were also vimentin positive. All sarcomas expressed vimentin. Synovial sarcomas were also keratin positive. All NHLs were vimentin positive, keratin negative. All NHLs except one expressed also LCA. It is concluded that keratin, vimentin and LCA are useful markers in differential diagnosis of malignant melanoma versus carcinoma and non-Hodgkin's lymphoma in fine needle aspirates when used together with morphologic and clinical data. However, in differential diagnosis of malignant melanoma and sarcoma these markers are of little use.

Biopsy, Needle↗
PubMed1991

Dendritic accessory cells derived from rat bone marrow precursors under chemically defined conditions in vitro belong to the myeloid lineage.

Serum-free conditions have been developed to differentiate dendritic cells from a non-adherent fraction of rat bone marrow precursors by action of the multipotential and macrophage colony-stimulating factors further supplemented with linoleic acid, vitamin E, and vitamin D3. Accessory activity was demonstrated by the high potency of the dendritic cells to stimulate autologous T cell proliferation, whereas such cells were negative for Fc receptor-dependent phagocytosis, a characteristic macrophage feature. While the dendritic cells were weakly positive for alpha-naphtylbutyrate esterase, they strongly expressed RT.1 class II antigens. Apparently, these cells represent a more differentiated phenotype since they expressed the nuclear A/C lamins. By addition of serum to the cultures, the dendritic cells developed into macrophages, which were also lamin A/C-positive as well as strongly positive for alpha-naphtylbutyrate esterase. Thus, these dendritic cells belong to the myeloid lineage, and it appears as if serum factor(s) control differentiation at a mature level. Suitable conditions could also be established for large-scale cultures of dendritic cells, which would be useful for applications requiring higher numbers of cells.

Animals↗
PubMed1991

[Sleep-related respiratory disorders and coronary heart disease].

A review of the literature shows that more than 50% of examined patients suffering from coronary heart disease were also suffering from sleep-related apnea. We were able to diagnose a pathological sleep apnea in 9 out of 25 patients (36%) suffering from an angiographically confirmed coronary 2-vessel and 3-vessel disorder. Patients with this combination--this is the hypothesis derived from our study--are at risk due to nocturnal apnea-induced myocardial ischaemia and rhythmic disorders. In 15 patients with sleep apnea and coronary heart disease or small vessel disease, nocturnal polysomnography was conducted, in parallel a 6-channel ECG was recorded. The apnea index (second night) was on the average 33 phases/h, the maximal duration of an apnea phases being 120 seconds. The minimal blood gas saturation recorded during sleep was between 46 and 89% (median 76.0%). In 4 of the 15 patients it was possible to confirm myocardial ischaemia (correlated via REM and also via NREM) with a maximum duration of 60 seconds, mainly during the phases of maximal apnea activity and blood gas desaturation. On comparing the ventricular arrhythmias waking/sleep, the Lown class did not change in 12 patients; there was deterioration in 2 patients and in one patient a qualitative improvement during the sleep phase. Patients suffering from sleep-related respiratory disorders and coronary heart disease are at cardiac risk, the more so since long-lasting apneas can lead to conditions of hypoxia at the heart in pre-existing changes in the coronary arteries, restricted coronary reserves and reduced tolerance to hypoxia. Such hypoxia can in turn induce enhanced electrical instability and a disturbance of the contractile function.

Arrhythmias, Cardiac↗
PubMed1990

Alkylation of cysteine 82 of p11 abolishes the complex formation with the tyrosine-protein kinase substrate p36 (annexin 2, calpactin 1, lipocortin 2).

p36 (annexin 2) is the major cytoplasmic target of the src tyrosine-kinase and forms in vitro and in vivo a stable tetrameric complex in which two p36 polypeptides interact with a dimer of a unique p11 polypeptide. p11 belongs into the superfamily of EF-hand proteins. Upon mild cysteine modification conditions, both cysteines (position 61 and 82) of the free p11 become substituted, and the ability to form the p36.p11 complex is lost. Under the same conditions, the 2 cysteines of p11 incorporated into the complex display differential reactivity. Here, cysteine 61 is fully substituted while cysteine-82 is protected. p11 derivatives substituted only on cysteine 61 retain binding activity for p36 unless cysteine 82 is substituted by a second cycle of modification of the isolated p11. Thus, the C-terminal extension protruding from the second EF-hand of the p11 molecule (residues 77-96) is important for the interaction with p36. As a consequence of our analysis, we report a new separation of p36 and p11 from the p36.p11 complex. This is based on a reversible cysteine modification and thus is an alternative to the denaturation and renaturation cycle used previously.

Alkylation↗
PubMed1990

Interaction of alpha-actinin and nebulin in vitro. Support for the existence of a fourth filament system in skeletal muscle.

Nebulin is a high molecular weight polypeptide (mass 0.6-0.8 million) which accounts for 3% of the myofibrillar mass in skeletal muscle. Due to its resistance to extraction under native conditions, relatively little is known about the biochemistry of the molecule. Here we report in vitro binding of alpha-actinin (a major Z-line protein) to nebulin. After solubilization with sodium dodecylsulfate myofibrillar polypeptides separated by gel electrophoresis were blotted on nitrocellulose and probed with 125I-labelled alpha-actinin. Nebulin is the only polypeptide decorated by alpha-actinin. This result gives biochemical support for the hypothesis, based on recent immunoelectron micrographs, that nebulin could form in skeletal muscle a fourth filament system, possibly extending to the Z-line.

Actinin↗
PubMed1990

Absorption and fluorescence spectroscopic studies of the Ca2(+)-dependent lipid binding protein p36: the annexin repeat as the Ca2+ binding site.

The existence of a single tryptophan residue in the protein p36, a member of a recently characterized family of Ca2+ binding proteins called annexins, is exploited to provide unique spectroscopic information on the annexin repeat motif and its role in Ca2+ binding. The differences in ultraviolet absorption and fluorescence excitation upon Ca2+ binding are interpreted solely in terms of this tryptophan, which, in view of the pronounced blue-shifts and the presence of vibronic structure, seems to reside in a highly nonpolar environment. The fluorescence emission from the protein is correspondingly blue-shifted, and it is found to transfer energy in resonance with Tb3+ absorption lines in the near-ultraviolet. This effect allows us to locate the Tb3+ and, by implication, the Ca2+ binding site to within ca. 8 A of the tryptophan residue.

Animals↗
PubMed1990

The two coiled coils in the isolated rod domain of the intermediate filament protein desmin are staggered. A hydrodynamic analysis of tetramers and dimers.

Desmin protofilaments and the proteolytically derived alpha-helical rod domain have been characterized by high-resolution gel permeation chromatography (GPC) using columns calibrated for the determination of viscosity radii. Additional characterization by chemical cross-linking and the determination of sedimentation values allowed the calculation of the molecular dimensions of the molecular species isolated. In dilute buffers GPC separated desmin rod preparations into two complexes: a dimer species (single coiled coil) with a length of 50 +/- 5 nm and a tetramer species (two coiled coils) with a length of 65 +/- 5 nm. Thus the two coiled coils in the tetramer are staggered by approximately 15 nm. The hydrodynamically derived lengths of the rod dimer and tetramer are supported by electron microscopy after metal shadowing. The hydrodynamic properties of desmin protofilaments follow that of the rod tetramer. The data on the hydrodynamic analysis of the rod tetramer of desmin in solution are in full agreement with the structural information recently deduced from paracrystals of the rod of glial fibrillary acid protein [Stewart, M., Quinlan, R.A. & Moir, R.D. (1989) J. Cell Biol. 109, 225-234]. Our results explain the inhomogeneity of molecules encountered in previous electron microscopical analyses.

Animals↗
PubMed1990

Protein chemical analysis of purified murine lamin B identifies two distinct polypeptides B1 and B2.

Lamin B purified from murine EAT cells was characterized by partial protein sequences. Contrary to the current view that mammals express only a single lamin B polypeptide corresponding to a characterized murine cDNA clone, our analysis documents two distinct B lamins. One protein follows the established cDNA sequence while the other identifies a novel murine lamin B. Comparison with the two chicken lamin B sequences established by cDNA cloning identifies the first murine lamin B sequence as a B1 type and the second as a B2 type. We conclude that mammals express two distinct lamin B forms as established by others for chicken.

Amino Acid Sequence↗
PubMed1990

Structural analysis of p36, a Ca2+/lipid-binding protein of the annexin family, by proteolysis and chemical fragmentation.

Limited proteolysis of the core domain of the 36-kDa protein p36 by trypsin gives a first insight into the structural organization of the four annexin repeats. Trypsin opens only a single peptide bond, situated between residues 204 and 205. The two fragments (of 20 kDa and 15 kDa), each containing two annexin repeats, remain as a tight complex (nicked core), which binds phospholipids in a Ca2(+)-dependent manner. After denaturation by 9 M urea, the nicked core is again formed upon renaturation provided both fragments are present. If the fragments are separated by chromatography in urea prior to renaturation, they show different behaviour. The 15-kDa C-terminal repeats aggregate, while the 20-kDa N-terminal repeats stay in solution. In comparison to p36, fragments with two (20-kDa fragment) or one (N-terminal CNBr fragment) annexin repeats show a conformational alteration in CD spectroscopy and hydrodynamics and display an increased susceptibility to proteases. In line with these differences, their Ca2(+)-dependent affinity to phospholipids is more than 10-20-fold decreased. Thus the four annexin repeats form together an integrated domain with multiple contacts between the repeats. Although stable derivatives with less than four repeats can be obtained, their Ca2+/phospholipid binding affinities are noticeably reduced.

Amino Acids↗
PubMed1990

[Sleep apnea].

Explore the source record for details and available documents.

Humans↗
PubMed1990

Structure of an invertebrate gene encoding cytoplasmic intermediate filament (IF) proteins: implications for the origin and the diversification of IF proteins.

The structure of the single gene encoding the cytoplasmic intermediate filament (IF) proteins in non-neuronal cells of the gastropod Helix aspersa is described. Genomic and cDNA sequences show that the gene is composed of 10 introns and 11 exons, spanning greater than 60 kb of DNA. Alternative RNA processing accounts for two mRNA families which encode two IF proteins differing only in their C-terminal sequence. The intron/exon organization of the Helix rod domain is identical to that of the vertebrate type III IF genes in spite of low overall protein sequence homology and the presence of an additional 42 residues in coil 1b of the invertebrate sequence. Intron position homology extends to the entire coding sequence comprising both the rod and tail domains when the invertebrate IF gene is compared with the nuclear lamin LIII gene of Xenopus laevis presented in the accompanying report of Döring and Stick. In contrast the intron patterns of the tail domains of the invertebrate IF and the lamin genes differ from those of the vertebrate type III genes. The combined data are in line with an evolutionary descent of cytoplasmic IF proteins from a nuclear lamin-like progenitor and suggest a mechanism for this derivation. The unique position of intron 7 in the Helix IF gene indicates that the archetype IF gene arose by the elimination of the nuclear localization sequence due to the recruitment of a novel splice site. The presumptive structural organization of the archetype IF gene allows predictions with respect to the later diversification of metazoan IF genes. Whereas models proposing a direct derivation of neurofilament genes seem unlikely, the earlier speculation of an mRNA transposition mechanism is compatible with current results.

Amino Acid Sequence↗
PubMed1990

Protein-protein recognition via short amphiphilic helices; a mutational analysis of the binding site of annexin II for p11.

Annexin II (p36) interacts with its ligand p11 via the short stretch of 12 amino acids (Ac-S-T-V-H-E-I-L-C-K-L-S-L) situated at the N-terminus. We have now synthesized some 37 tetradecapeptides, which differ from the original p11 binding sequence (Ac1-14) by single amino acid substitutions. The relative affinity of each peptide for p11 was determined by fluorescence spectroscopy using a competitive binding assay. The binding behaviour of the different peptides confirms the model of an amphiphilic alpha-helix induced upon binding to p11. The apparent affinities delta delta Gbind of the mutant peptides revealed that the N-acetyl group of serine 1 and the hydrophobic side chains at positions 3, 6, 7 and 10 contribute most to the binding. The observed destabilization of the complex upon removal of signal methyl groups from the hydrophobic side of the helix is comparable with the destabilization of proteins in which methyl groups have been removed from the inner core. We conclude that upon binding to p11 the hydrophobic side of the amphiphatic alpha-helix becomes fully buried.

Amino Acid Sequence↗
PubMed1990

Electrical impedance, rCBF, survival and histology in Mongolian gerbils with forebrain ischaemia.

Survival, quantitative morphology of the hippocampus, cerebral tissue impedance and regional cerebral blood flow (rCBF) were studied in the Mongolian gerbil after 15 minutes of bilateral common carotid occlusion. A subgroup of animals was placed in cages with free access to running-wheels for two weeks preceding ischaemia to measure voluntary locomotor activity. Survival was enhanced in the running-wheel subgroup, with 90% of the animals still alive after 14 days as compared to 48% of the non-running group. Neuronal loss was found in all animals in the hippocampus (CA1, CA2, CA3 and CA4), and was most pronounced in the CA1 sector. In the running-wheel group, however, neuronal loss was significantly lower in sectors CA2, CA3 and CA4. The increases of cerebral impedance, which indicate ischaemic cell swelling, reached 190% in both groups during ischaemia. During postischaemic recirculation, however, impedance normalized more rapidly in the running-wheel group, indicating earlier resolution of ischaemic cell swelling. In wheel-running gerbils, postischaemic hyperperfusion evolved earlier and was more pronounced as compared to non-runners. No differences in systemic blood pressure were observed during cerebral ischaemia or thereafter.

Animals↗
PubMed1990

[Do immunologic markers facilitate differentiation between histologic types of stomach cancer?].

Seventeen gastric carcinomas (intestinal n = 12; diffuse n = 1; mixed type n = 4) and one Barrett's carcinoma were prospectively studied by immunohistochemistry for the expression of different keratin polypeptides and of the brush border markers villin, sucrase isomaltase and aminopeptidase N. All carcinomas expressed the keratin polypeptides 8, 18, and 19 and were stained by the broad specific keratin antibody KL1, irrespective of histologic type. Keratin 7, however, was expressed in only one carcinoma in most tumor cells and in two further carcinomas in some tumor cells. Thus, specific differentiation of the various histologic types of gastric carcinoma does not seem to be aided by the use of keratin antibodies. Villin was positive in 80% of the tumors and sucrase isomaltase and aminopeptidase N were positive in 67% respectively with no obvious histologic difference. The frequent positivity of the brush border markers, usually typical for intestinal epithelium, reflects the high degree of intestinal differentiation of gastric carcinomas, but again does not seem to be associated with a particular histologic type.

Adenocarcinoma↗
PubMed1990

Induction of nuclear lamins A/C in macrophages in in vitro cultures of rat bone marrow precursor cells and human blood monocytes, and in macrophages elicited in vivo by thioglycollate stimulation.

Hemopoietic cells from blood and bone marrow of mammals usually do not express lamins A/C but only lamin B, and this feature distinguishes these cells from the vast majority of somatic cells of the adult animal, which reveal lamins A/C as well as lamin B. Here we have cultivated rat bone marrow precursor cells and human monocytes isolated from peripheral blood in tissue culture supplemented with certain growth factors. These conditions allow bone marrow precursor cells and monocytes to differentiate almost quantitatively into accessory cells and/or mature macrophages. The different cell types in the cultures can be identified both morphologically and by other assays. Antibodies specific for mouse A/C lamins, human A/C lamins, or B lamins have been used to define the lamin complement as a function of time in culture and of cell type. A dramatic increase in lamin A/C-positive cells was observed in the first 3 days of culture with both accessory cells and macrophages expressing lamins A/C as soon as such cell types could be identified. Parallel in vivo experiments showed that treatment with thioglycollate caused the percentage of lamin A/C-positive peritoneal macrophages to increase from 5 to 80% between Days 0 and 6.

Animals↗
PubMed1990

Differential distribution of subsets of myofibrillar proteins in cardiac nonstriated and striated myofibrils.

Cultured cardiac myocytes were stained with antibodies to sarcomeric alpha-actinin, troponin-I, alpha-actin, myosin heavy chain (MHC), titin, myomesin, C-protein, and vinculin. Attention was focused on the distribution of these proteins with respect to nonstriated myofibrils (NSMFs) and striated myofibrils (SMFs). In NSMFs, alpha-actinin is found as longitudinally aligned, irregular approximately 0.3-microns aggregates. Such aggregates are associated with alpha-actin, troponin-I, and titin. These I-Z-I-like complexes are also found as ectopic patches outside the domain of myofibrils in close apposition to the ventral surface of the cell. MHC is found outside of SMFs in the form of discrete fibrils. The temporal-spatial distribution and accumulation of the MHC-fibrils with respect to the I-Z-I-like complexes varies greatly along the length of the NSMFs. There are numerous instances of I-Z-I-like complexes without associated MHC-fibrils, and also cases of MHC-fibrils located many microns from I-Z-I-like complexes. The transition between the terminal approximately 1.7-microns sarcomere of any given SMF and its distal NSMF-tip is abrupt and is marked by a characteristic narrow alpha-actinin Z-band and vinculin positive adhesion plaque. A titin antibody T20, which localizes to an epitope at the Z-band in SMFs, precisely costains the 0.3-microns alpha-actinin aggregates in ectopic patches and NSMFs. Another titin antibody T1, which in SMFs localizes to an epitope at the A-I junction, typically does not stain ectopic patches and NSMFs. Where detectable, the T1-positive material is adjacent to rather than part of the 0.3-microns alpha-actinin aggregates. Myomesin and C-protein are found only in their characteristic sarcomeric locations (even in just perceptible SMFs). These A-band-associated proteins appear to be absent in ectopic patches and NSMFs.

Actinin↗
PubMed1990

The coiled coil of in vitro assembled keratin filaments is a heterodimer of type I and II keratins: use of site-specific mutagenesis and recombinant protein expression.

Recombinant DNA technology has been used to analyze the first step in keratin intermediate filament (IF) assembly; i.e., the formation of the double stranded coiled coil. Keratins 8 and 18, lacking cysteine, were subjected to site specific in vitro mutagenesis to change one amino acid in the same relative position of the alpha-helical rod domain of both keratins to a cysteine. The mutations lie at position -36 of the rod in a "d" position of the heptad repeat pattern, and thus air oxidation can introduce a zero-length cystine cross-link. Mutant keratins 8 and 18 purified separately from Escherichia coli readily formed cystine homodimers in 2 M guanidine-HCl, and could be separated from the monomers by gel filtration. Heterodimers with a cystine cross-link were obtained when filaments formed by the two reduced monomers were allowed to oxidize. Subsequent ion exchange chromatography in 8.5 M urea showed that only a single dimer species had formed. Diagonal electrophoresis and reverse phase HPLC identified the dimer as the cystine containing heterodimer. This heterodimer readily assembled again into IF indistinguishable from those obtained from the nonmutant counterparts or from authentic keratins. In contrast, the mixture of cystine-stabilized homodimers formed only large aberrant aggregates. However, when a reducing agent was added, filaments formed again and yielded the heterodimer after oxidation. Thus, the obligatory heteropolymer step in keratin IF assembly seems to occur preferentially at the dimer level and not during tetramer formation. Our results also suggest that keratin I and II homodimers, once formed, are at least in 2 M guanidine-HCl a metastable species as their mixtures convert spontaneously into heterodimers unless the homodimers are stabilized by the cystine cross-link. This previously unexpected property of homodimers explains major discrepancies in the literature on the keratin dimer.

Actin Cytoskeleton↗