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Biomedical subjects

M Wagner

Publications and source records attributed to M Wagner.

At least 469 records · Page 26Linked to original sources
PubMed1994

Loss of chromosome arm 8p loci in prostate cancer: mapping by quantitative allelic imbalance.

A previous study of 18 primary or metastatic prostate cancers showed loss of genetic markers on chromosome 8; 10, or 16 in more than 50% of cases [Bergerheim USR et al. (1991) Genes Chromosom Cancer 3:215-220]. The small size and infiltrative nature of primary prostatic tumors have hindered efforts to assess allelic losses by traditional restriction fragment length polymorphism (RFLP)/Southern blotting methods. To improve the sensitivity and specificity of this analysis in early prostate cancer, we have amplified polymorphic microsatellite repeats by polymerase chain reaction (PCR), and have quantitated allelic imbalances with phosphor imaging technology. In this study, 63 primary prostate tumors and matched benign tissues obtained by radical prostatectomy were examined at 28 genetic loci on chromosome 8, all but five of which were located on the short arm. Twenty-nine (46%) of the 63 cases showed loss of at least one locus. Multiple adjacent loci, usually including the LPL and MSR genes in 8p22, were lost in 28 cases. In 10 of these, losses were observed at all informative loci on the p arm. In another 15 tumors, losses were restricted to subregions of the p arm by loci retained either distally toward the p terminus or proximally at the 8p12-8p21 border, or both. In three tumors, two discrete regions of loss were observed within 8p, separated by several retained loci. Allelic loss of 8p loci was associated with higher tumor grade. These data are complementary to previous reports of allelic deletions in colorectal, hepatocellular, and non-small cell lung cancers and suggest the existence of one or more pleotropic tumor suppressor genes on 8p.

Alleles↗
PubMed1994

Deletion screening of mitochondrial DNA via multiprimer DNA amplification.

We report an application of multiprimed polymerase chain reaction (PCR) which allows a rapid, nonradioactive detection of deletions in mitochondrial DNA using EDTA-blood and muscle samples. The use of two primer sets consisting of three forward and five reverse primers, respectively, allows a competitive PCR resulting in significant amplification products only in the presence of deletion-harbouring DNA species. Under the conditions described, deletions causing Kearns-Sayre syndrome (KSS) and progressive external ophthalmoplegia (PEO) have been successfully detected. The location of the primers on mitochondrial DNA used in this study should allow identification and localization of most of the large-scale deletions (i.e. more than 1 kb) of mitochondrial DNA reported so far.

Base Sequence↗
PubMed1994

Investigation of the role of signal transduction in attachment of ocular melanoma cells to matrix proteins: inhibition of attachment by calmodulin antagonists including tamoxifen.

We investigated the role of signal transduction systems in the attachment of human uveal melanoma cells to matrix proteins. Ocular melanoma cells established from primary tumours attached rapidly to all substrates examined. Preferred substrates of attachment were collagens type I, III and IV and fibronectin rather than laminin, gelatin, arginine-glycine-aspartine, vitronectin, poly-L-lysine or plastic. All cells showed rapid attachment to the preferred substrates (80% within 10 min). Manipulation of intracellular cyclic AMP or protein kinase C activity had relatively little effect on cell attachment. In contrast, attachment was significantly reduced by manipulating either intracellular calcium or calmodulin. After 15 min at 37 degrees C, the calcium ionophore ionomycin (5 microM) reduced attachment to 25%, and TMB8 (50 microM), which can reduce intracellular calcium, reduced attachment to 60%. The experimental calmodulin antagonist J8 (25 microM), a substituted naphthalene sulphonamide, reduced attachment to 40%. Similarly tamoxifen (25 microM), which has calmodulin antagonist activity in vitro, reduced attachment to 55%. Both J8 and tamoxifen inhibited cell attachment to a wide range of matrix proteins, suggesting that this effect on attachment is not dependent on the presence of specific adhesion receptors. Reduction of ocular melanoma tumour cell/matrix interactions through manipulation of intracellular calcium or calmodulin may therefore merit further investigation as a possible approach to reducing metastatic spread.

Calcium↗
PubMed1994

Hemispheric disconnection in Marchiafava-Bignami disease: clinical, neuropsychological and MRI findings.

We report a single-case study of a patient suffering from Marchiafava-Bignami disease (MBD), a rarely intra vitam diagnosed syndrome with focal demyelination in the corpus callosum, which is usually found in patients with chronic alcohol abuse. Our study explored a variety of motor, sensory and visual functions. Striking features were left apraxia caused by two lesions in the rostral area, and left hemidyslexia related to a large splenial lesion demonstrated by MRI. As symptoms of disconnection may remain undetected clinically, MRI is a most useful tool for the diagnosis of MBD.

Adult↗
PubMed1994

Event-related and motor responses to probes in a forewarned reaction time task in schizophrenic patients.

Surface-negative brain potentials indicate increased excitability of the underlying cortical neural networks. Consequently, deviant patterns of event-related potentials in schizophrenic patients reveal an atypical regulation of cortical excitability. Twelve patients with a chronic schizophrenic disorder and 12 matched control subjects were investigated using a probe paradigm: A contingent negative variation (CNV) was evoked in a forewarned reaction time paradigm. Clicks were presented before, during and after elicitation of the CNV. Click-evoked responses allow one to 'probe' the current brain state, particularly neuronal excitability, which is also reflected by the slow potentials. During the measurements, subjects pressed one button in response to the offset of the visual warning stimulus and a different button in response to the acoustic probes, the latter button press being a behavioral indication of the brain's excitability. In the forewarned reaction time task, patients developed a CNV with a frontal maximum, while the CNV in control subjects was predominantly centro-parietal. This atypical topographical pattern of the CNV may indicate a different spatio-temporal regulation of cortical preparatory processes in schizophrenics. Motor responses were accelerated during negative potential shifts in both patients and controls, with responses being slower overall in patients. In patients, probe-evoked potentials revealed a smaller N100, but a larger P300, than in controls. The covariation of these brain waves with slow potentials, however, turned out to be similar for both groups.

Adult↗
PubMed1994

Rescue of renal transplants with distal ureteral complications by pyelo-pyelostomy.

Secondary ureteroneocystostomy is the procedure most commonly used for repair of ureteral stenosis or necrosis in renal transplant patients. However, when the transplant ureter available for reconstruction is too short, ureteropyelostomy is the standard procedure. Another option is pyelo-pyelostomy, which has been used infrequently to date. Between 1979 and 1988 we managed 6 patients (4 men and 2 women 42 to 62 years old) with renal allografts who required repair of the transplant ureter for long segmented ureteral necrosis (3), long segmented ureteral stenosis (2) and an iatrogenic lesion of the ureter (1) with pyelo-pyelostomy. Of the 6 patients 2 had undergone previous rescue operations. Pyelo-pyelostomy with the renal pelvis of the graft was performed as first described by Gil-Vernet and Caralps in 1968. Median followup was 6 years (range 3 to 12 years). Two patients died of chronic rejection and pulmonary embolism, respectively. Four patients are still alive with functioning transplants and serum creatinine levels of less than 210 mumol/l. According to our experiences with 6 patients, we believe that pyelo-pyelostomy is a promising rescue maneuver that is worth consideration in cases of total or subtotal ureteral necrosis and long segmented ureteral stenosis after renal transplantation.

Adult↗
PubMed1994

JM9 strains, a new type of group B streptococci from Japan.

JM9 strains isolated from human carriers and patients in several districts of Japan, represent a new serotype of group B streptococci (GBS, Streptococcus agalactiae). They were first detected in 1983 but are meanwhile prevailing among all other GBS serotypes in Japan. Outside of Japan, strains of this type have not been reported until now. In the present work, N-acetylneuraminic acid was detected in all strains investigated, by chemical analysis as well as by interaction with a sialic acid-binding lectin. This component is characteristic of all analyzed GBS type polysaccharides. In a chicken embryo model, all strains exhibited a very strong virulence. Examination of the antibiotic sensitivity revealed that all strains were susceptible to penicillin, cephalothin, clindamycin, mezlocillin, azlocillin, erythromycin, methicillin, chloramphenicol, tetracyclin, oxacillin and sulfamethoxazole/trimethoprim, whereas all strains were resistant against gentamicin, kanamycin and neomycin. Electron microscopic studies revealed for these strains relatively small capsules but unusually thick cell walls. By immunogold labelling, the type polysaccharide, the group polysaccharide, the lipoteichoic acid and in some strains, the protein R were localized.

Animals↗
PubMed1994

TGN38 recycles basolaterally in polarized Madin-Darby canine kidney cells.

Sorting of newly synthesized plasma membrane proteins to the apical or basolateral surface domains of polarized cells is currently thought to take place within the trans-Golgi network (TGN). To explore the relationship between protein localization to the TGN and sorting to the plasma membrane in polarized epithelial cells, we have expressed constructs encoding the TGN marker, TGN38, in Madin-Darby canine kidney (MDCK) cells. We report that TGN38 is predominantly localized to the TGN of these cells and recycles via the basolateral membrane. Analyses of the distribution of Tac-TGN38 chimeric proteins in MDCK cells suggest that the cytoplasmic domain of TGN38 has information leading to both TGN localization and cycling through the basolateral surface. Mutations of the cytoplasmic domain that disrupt TGN localization also lead to nonpolarized delivery of the chimeric proteins to both surface domains. These results demonstrate an apparent equivalence of basolateral and TGN localization determinants and support an evolutionary relationship between TGN and plasma membrane sorting processes.

Amino Acid Sequence↗
PubMed1994

In situ probing of gram-positive bacteria with high DNA G + C content using 23S rRNA-targeted oligonucleotides.

23S-rRNA-targeted oligonucleotide probes were designed for the phylogenetic group 'Gram-positive bacteria with high G + C content of DNA' (GPBHGC). A sequence idiosyncrasy in two adjacent base pairs in the stem of helix 69 in domain IV of the 23S rRNA is present in all hitherto analysed strains of GPBHGC. An oligonucleotide probe targeted to this region hybridized only with strains of GPBHGC and was successfully used for in situ monitoring of these cells in activated sludge. Another unique feature of the 23S rRNA molecules of GPBHGC is a large insertion in domain III. Fluorescent oligonucleotides targeted to the highly variable regions of the rRNA within the insertions of Corynebacterium glutamicum DSM 20300, Aureobacterium testaceum DSM 20166 and Brevibacterium sp. DSM 20165 hybridized specifically to their target strains, whereas probing with oligonucleotides complementary to the rRNA-coding strand of the 23S rDNA and to the spacer between 16S and 23S rRNA of C. glutamicum did not result in detectable fluorescence. This confirmed that the large 23S insertions are indeed present in 23S rRNAs of GPBHGC and provide potential target sites for highly specific nucleic acid probes.

Base Sequence↗
PubMed1994

T-cell lines derived from lesional skin of lichen planus patients contain a distinctive population of T-cell receptor gamma delta-bearing cells.

Lichen planus is characterized by a dense infiltrate of T lymphocytes at the dermoepidermal junction. To determine the phenotypic and functional characteristics of the infiltrating lymphocytes, T-cell lines from normal and lesional skin from the same patients with lichen planus were established by culture with interleukin 2 followed by stimulation every 14 d with phytohemagglutinin and irradiated allogeneic feeder cells. Resultant T-cell lines were immunophenotyped by staining with monoclonal antibodies and their reactivity tested by determining their cytolytic activity to selected targets. T-cell lines from 13 lesional and nine normal biopsy specimens were studied. T-cell lines from normal skin were 61% CD4+ and 32% CD8+, whereas lines from lesional skin had significantly fewer CD4+ cells (13%) and more CD8+ cells (62%). T-cell lines from lesional skin contained a distinctive population of gamma delta T cells that was rarely present in lines derived from normal skin. We were able to culture gamma delta T cells out of the lesional skin of 12 of 13 patients. In these 12 patients, lesional T-cell lines were 17% gamma delta+ (range 2% to 47%). Only one T-cell line from normal skin contained significant numbers of gamma delta T cells. The gamma delta population from lesional skin was commonly V delta 1J delta 1+. These results suggest that CD8+ and TCR gamma delta+ T lymphocytes may be involved in the development or the maintenance of lichen planus.

Cell Line↗
PubMed1994

In situ analysis of microbial consortia in activated sludge using fluorescently labelled, rRNA-targeted oligonucleotide probes and confocal scanning laser microscopy.

Activated sludge flocs are complex consortia of various micro-organisms. The community structures of samples taken from municipal sewage treatment plants were characterized using fluorescently labelled, 16S and 23S rRNA-targeted oligonucleotide probes in combination with confocal scanning laser microscopy (CSLM). In comparison with conventional epifluorescence microscopy, CSLM considerably improved the capability to visualize directly the spatial distribution of defined bacterial populations inside the sludge flocs. Analyses could be performed at high resolution undisturbed by problems such as autofluorescence or limited spatial resolution in thick samples. In addition, CSLM was used to analyse some structural properties of paraformaldehyde-fixed activated sludge flocs, such as floc size and homogeneity. Typical floc sizes were found to be in the range between 5 and 50 microns. Whereas most of the flocs were completely colonized by bacteria, there were also examples of flocs containing gas bubbles or particles in the interior.

Bacteria↗
PubMed1994

The sialic acid-containing lipopolysaccharides of Salmonella djakarta and Salmonella isaszeg (serogroup O: 48): chemical characterization and reactivity with a sialic acid-binding lectin from Cepaea hortensis.

Lipopolysaccharides (LPS) of Salmonella djakarta and Salmonella isaszeg, as well as of a spontaneous R-mutant of S. djakarta were investigated as to their content in neuraminic acid (Neu) and its individual linkage. The two Salmonella serovars both belong to the O:48 serogroup of Salmonella, but to two different subgroups. LPS of both S-forms contained high amounts of Neu, although in different quantities, whereas the R-form was completely devoid of it. Methylation analysis indicated that Neu is exclusively terminally linked in S. djakarta whereas both terminal and 4-linked Neu were recognized in S. isaszeg. Although terminally linked, a sialidase from Arthrobacter ureafaciens was unable to split Neu even after prolonged incubation from both S-type LPSs. When LPS was first treated by mild alkali, however, the total amount of Neu from S. djakarta LPS and about 50% from that of LPS of S.isaszeg could be removed. In contrast, alkali-treated LPS, but also the non-treated one, proved to be effective inhibitors for a sialic acid-binding lectin from Cepaea hortensis. The resistance of terminal Neu towards sialidase may be due to the presence of an O-acetyl group which would be removed during the methylation analysis but would, especially when linked to C-4, not interfere with the reactivity of the lectin.

Animals↗
PubMed1994

Tamoxifen inhibition of ocular melanoma cell attachment to matrix proteins.

Tamoxifen plays a major role in the management of breast cancer in women and is currently used to a lesser extent in other neoplasias. Many of the pharmacological properties of tamoxifen are consistent with anti-estrogen activity, but it also has significant, although lesser, benefit in patients whose tumours are estrogen-receptor negative. We recently reported that murine B16 melanoma cell attachment to extracellular matrix proteins can be inhibited by calmodulin antagonists. In seeking a calmodulin antagonist that could be used clinically, we investigated tamoxifen, which is known to have calmodulin antagonist activity in vitro, and confirmed that it will inhibit murine melanoma cell attachment in vitro. In the current study, we examined the effect of tamoxifen on the attachment of human ocular melanoma cell lines to a range of extracellular matrix substrates to evaluate the potential relevance of calmodulin antagonists, including tamoxifen, to reducing metastatic spread of these tumours. We report that six ocular melanoma cell lines established from choroidal melanoma tumours showed rapid attachment to a range of substrates and that this attachment can be significantly reduced by an experimental calmodulin antagonist (J8) and by tamoxifen. In summary, we conclude that the ability of calmodulin antagonists, including tamoxifen, to inhibit ocular melanoma cell attachment to matrix proteins in vitro merits further investigation as it may offer another approach to reducing metastatic spread of these tumours.

Animals↗
PubMed1994

Development of an rRNA-targeted oligonucleotide probe specific for the genus Acinetobacter and its application for in situ monitoring in activated sludge.

Enhanced biological phosphate removal in an anaerobic-aerobic activated sludge system has generally been ascribed to members of the genus Acinetobacter. A genus-specific 16S rRNA-targeted oligonucleotide probe was developed to investigate the role of Acinetobacter spp. in situ. Nonisotopic dot blot hybridization to 66 reference strains, including the seven described Acinetobacter spp., demonstrated the expected probe specificity. Fluorescent derivatives were used for in situ monitoring of Acinetobacter spp. in the anaerobic and aerobic compartments of a sewage treatment plant with enhanced biological phosphate removal. Microbial community structures were further analyzed with oligonucleotide probes specific for the alpha, beta, or gamma subclasses of the class Proteobacteria, for the Cytophaga-Flavobacterium cluster, for gram-positive bacteria with a high G + C DNA content, and for all bacteria. Total cell counts were determined by 4',6-diamidino-2-phenylindole staining. In both the anaerobic and the aerobic basins, the activated sludge samples were dominated by members of the class Proteobacteria belonging to the beta subclass and by gram-positive bacteria with a high G + C DNA content. Acinetobacter spp. constituted less than 10% of all bacteria. For both basins, the microbial community structures determined with molecular techniques were compared with the compositions of the heterotrophic saprophytic microbiota determined with agar plating techniques. Isolates on nutrient-rich medium were classified by whole-cell hybridization with rRNA-targeted probes and fatty acid analysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Acinetobacter↗
PubMed1994

Comparative studies on vascular endothelium in vitro. I. Cytokine effects on the expression of adhesion molecules by human umbilical vein, saphenous vein and femoral artery endothelial cells.

Endothelial cells (ECs) are very responsive to proinflammatory cytokines. ECs are stimulated by these substances to increase expression of cell surface adhesion molecules, leading to dramatically altered interactions with leukocytes. In these interactions, E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) are suggested to play the most important role. Recent evidence has suggested diversity in the responses of ECs from different regions of the vascular system. Human umbilical vein ECs (HUVECs) are the most often used EC culture model, although there are few studies comparing their response with other human EC types from the adult organism. In this study the expression of E-selectin, ICAM-1 and VCAM-1 on cultured human adult ECs from the saphenous vein (HSVECs) and from the femoral artery (HAFECs), as well as HUVECs was studied. Using a cell enzyme immunoassay as well as immunoelectron microscopical methods, we found that both HSVECs and HAFECs respond in a similar way to HUVECs to exogenous stimulation by IL-1 beta, TNF alpha or LPS. IL-1 beta and TNF alpha increased the expression of E-selectin on the cytoplasmic membranes of HUVECs, HSVECs and HAFECs and elicited even similar absolute quantities of this molecule, comparing the different cell types. ICAM-1 and VCAM-1 appeared to be regulated dose dependently by IL-1 beta, independent of the EC type. HUVECs as well as HSVECs and HAFECs gave a reproducible constitutive ICAM-1 expression, whereas E-selectin and VCAM-1 were absent on nonstimulated ECs.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

Brefeldin A causes structural and functional alterations of the trans-Golgi network of MDCK cells.

The trans-Golgi network (TGN) of MDCK cells is exquisitely sensitive to the fungal metabolite brefeldin A (BFA), in contrast to the refractory Golgi stack of these cells. At a concentration of 1 microgram/ml, BFA promoted extensive tubulation of the TGN while the medical Golgi marker alpha-mannosidase II was not affected. Tubules emerging minutes after addition of the drug contained both the apical marker influenza hemagglutinin (HA), previously accumulated at 20 degrees C, and the fusion protein interleukin receptor/TGN38 (TGG), a TGN marker that recycles basolaterally, indicating that, in contrast to TGN vesicles, TGN-derived tubules cannot sort apical and basolateral proteins. After 60 minutes treatment with BFA, HA and TGG tubules formed extensive networks widely spread throughout the cell, different from the focused centrosomal localization previously described in non-polarized cells. The TGG network partially codistributed with an early endosomal tubular network loaded with transferrin, suggesting that the TGG and endosomal networks had fused or that TGG had entered the endosomal network via surface recycling and endocytosis. The extensive structural alterations of the TGN were accompanied by functional disruptions, such as the extensive mis-sorting of influenza HA, and by the release of the TGN marker gamma-adaptin. Our results suggest the involvement of BFA-sensitive adaptor proteins in TGN-->surface transport.

Adaptor Protein Complex gamma Subunits↗