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Biomedical subjects

S Müller

Publications and source records attributed to S Müller.

At least 235 records · Page 13Linked to original sources
PubMed1998

Glove powder affects skin roughness, one parameter of skin irritation.

Most patients with immediate-type-hypersensitivity to natural rubber latex (NRL) give a history of hand eczema. Susceptibility to allergens is increased by irritant hand eczema due to damage to the skin barrier. Therefore, especially for employees in medical or paramedical professions, reduction in skin irritation is of importance. The present study reports the effect of glove powder on skin roughness, one feature of skin irritancy. Skin replicas, performed before and after wearing different types of gloves, were evaluated by laser profilometry. Significant alteration of skin roughness was evident and determined by different factors. Use of unpowdered Biogel gloves on prepowdered hands results in an increase in skin roughness, in contrast to the use of Biogel gloves on hands without any pre-application. On the other hand, powdered Manex neoderm gloves show no influence on the skin structure, whereas powdered Peha taft gloves also increase the roughness of the skin. Other potential candidates causing skin morphology alteration could be the glove pH, rubber chemicals with irritant potential and other characteristics of gloves, which are already under investigation.

Adult↗
PubMed1998

Purification of the inlB gene product of Listeria monocytogenes and demonstration of its biological activity.

Entry of Listeria monocytogenes into nonphagocytic cells requires the inlAB gene products. InlA and InlB are bacterial cell wall-associated polypeptides that can be released by sodium dodecyl sulfate treatment. By applying more gentle extraction methods, we have purified InlB in its native form. Treatment of bacteria with various nondenaturating agents including mutanolysin, thiol reagents, sodium chloride, and detergents like Triton X-100 or 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate did not release substantial amounts of InlB from the bacterial cell wall. Instead, InlB was nearly quantitatively extracted in a solubilized form by treatment of bacteria with 1 M Tris-Cl or other protonated amines at pH 7.5. However, the reduced solubility of the extracted InlB in low-salt buffers hampered further biochemical purification. A panel of monoclonal antibodies against listerial Tris-Cl extracts containing InlB was therefore produced to generate reagents for use in affinity chromatography. One of the monoclonal antibodies enabled purification of the InlB protein to homogeneity with relatively high yields. When added externally, purified InlB associated with the surface of noninvasive bacteria such as Listeria innocua or an L. monocytogenes inlB2 mutant, where it promoted entry of these strains into Vero cells >300- and 17-fold, respectively. This effect was even more dramatic for HeLa cells, where the observed invasion was increased about 9,000- and 4,000-fold, respectively. The availability of purified native, invasion-competent InlB will allow analysis of the molecular basis of InlB-mediated entry into tissue culture cell lines in greater detail.

Animals↗
PubMed1998

Dynamic three-dimensional echocardiography using parallel slicing: a promising diagnostic procedure in adults with congenital heart disease.

In 18 of 77 adult patients presenting with congenital heart disease, three-dimensional conceptualization was found to be insufficient using two-dimensional echocardiographic imaging. In these individuals, three-dimensional image reconstruction from digitized parallel images was carried out, providing additional information in 14 of them. Three-dimensional echocardiography and volumetry may, especially in patients with atrial septal defect, cor triatriatum and corrected transposition of the great arteries, be an asset amplifying the diagnostic information yielded by conventional two-dimensional echocardiographic approaches.

Adolescent↗
PubMed1998

Expression and synthesis of insulin-like growth factor-binding proteins in human glioma cell lines.

We previously demonstrated the presence of insulin-like growth factors (IGFs) and IGF-receptors in human glioma cell lines derived from primary glioblastomas. The biological action of IGFs is modulated by specific IGF-binding proteins (IGFBP)-1 to -7. By means of polymerase chain reaction (PCR), we detected mRNA transcripts for IGFBP-1 in 42%, IGFBP-2 in 65%, IGFBP-3 in 97%, IGFBP-4 in 3%, IGFBP-5 in 74%, IGFBP-6 in 94% and IGFBP-7 in 87% of the glioma cell lines. The specificity of the PCR reaction was verified by direct sequencing of the PCR product. In addition, the content of the most prevalent IGFBP-3 was measured in conditioned medium from glioma cells by specific radioimmunoassay with levels ranging from < 1 to 620 ng/ml. Moreover, the presence of membrane-bound IGFBPs (44, 50 and 60 kDa) as well as IGF-II receptors was demonstrated by using 125I-labelled IGF-II as a ligand. In conclusion, IGFBPs may modulate the IGF-mediated effects in these cell lines.

Aged↗
PubMed1998

Activated CD4+ and CD8+ cytotoxic cells are present in increased numbers in the intestinal mucosa from patients with active inflammatory bowel disease.

The contribution of cell-mediated cytotoxicity to the pathogenesis of inflammatory bowel disease (IBD) is controversial, and results of in vitro assays vary according to experimental procedures. Therefore, we compared the frequency of cytotoxic effector cells in situ. On tissue sections of controls (n = 11), low frequencies of granzyme A and perforin mRNA-expressing cells are found in the lamina propria (1.77 +/- 0.15% and 1.46 +/- 0.12%, respectively) and in the epithelial cell layer (0.76 +/- 0.12% and 0.66 +/- 0.10%, respectively). In patients with IBD (n = 33), corresponding values were significantly (P < 0.02) higher, 6.1 +/- 0.40% and 5.92 +/- 0.57% for granzyme A and perforin expression in the lamina propria and 2.50 +/- 0.19% and 2.59 +/- 0.28%, respectively, in the epithelial compartment. Differences between ulcerative colitis and Crohn's disease are statistically not significant (P > 0.33). Activated cytotoxic cells are preferentially found at sites facing the intestinal lumen. Perforin mRNA-expressing cells are mainly CD8+ T cells. CD4+ T cells expressing perforin mRNA are mainly isolated from affected areas of patients with Crohn's disease. Immunostaining for perforin protein generally coincides with perforin mRNA in situ. These data demonstrate that cytotoxic cells are vigorously activated in situ in the intestinal mucosa of patients with active IBD.

Adult↗
PubMed1998

Mobilization and collection of allogeneic peripheral blood progenitor cells for transplantation.

A median dose of 11 (6-17) microg G-CSF per kg and day was given to 96 (49 female, 47 male) healthy family donors in order to mobilize and to collect peripheral blood progenitor cells (PBPC) for allogeneic transplantation. Donor age was 36 (17-76) years. The leukocytes of the donors increased to 46 (12-115) x 10(9)/l on days 4-6 of G-CSF treatment with a median of 71 (2-657) CD34+ cells per microl, respectively. Female and older donors seem to have a lower response to G-CSF. About 32% of the donors suffered from side-effects of G-CSF requiring analgetics. A total of 197 stem cell aphereses were performed using the COBE Spectra cell separator. Median apheresis time was 225 (118-300) min processing 11.8 (5.7-20) l blood, collecting 5.3 (1.7-14.9) x 10(10) nucleated cells and containing 0.7 (0.1-3.7)% CD34+ cells. Severe citrate toxicity occurred in 5% of the donors. Retransfusion of autologous platelets post apheresis was necessary in 16% of the donors because of a platelet count <80 x 10(9)/l. An insufficient number of stem cells was collected in four female donors due to a very poor response to G-CSF. In conclusion, the collection of allogeneic G-CSF-mobilized PBPC is safe and effective. One or two aphereses were sufficient in 91% of the donors to achieve >4 x 10(6) CD34+ cells per kg. In 4% of the donors an additional bone marrow harvest or the use of an alternative donor was necessary because of a poor mobilization.

Adolescent↗
PubMed1997

Inter-domain cross-linking and molecular modelling of the hairpin ribozyme.

The hairpin ribozyme is a small catalytic RNA composed of two helical domains containing a small and a large internal loop and, thus, constitutes a valuable paradigm for the study of RNA structure and catalysis. We have carried out molecular modelling of the hairpin ribozyme to learn how the two domains (A and B) might fold and approach each other. To help distinguish alternative inter-domain orientations, we have chemically synthesized hairpin ribozymes containing 2'-2' disulphide linkages of known spacing (12 or 16 A) between defined ribose residues in the internal loop regions of each domain. The abilities of cross-linked ribozymes to carry out RNA cleavage under single turnover conditions were compared to the corresponding disulphide-reduced, untethered ribozymes. Ribozymes were classed in three categories according to whether their cleavage rates were marginally, moderately, or strongly affected by cross-linking. This rank order of activity guided the docking of the two domains in the molecular modelling process. The proposed three-dimensional model of the hairpin ribozyme incorporates three different crystallographically determined structural motifs: in domain A, the 5'-GAR-3'-motif of the hammerhead ribozyme, in domain B, the J4/5 motif of group I ribozymes, and connecting the two domains, a "ribose zipper", another group I ribozyme feature, formed between the hydroxyl groups of residues A10, G11 of domain A and C25, A24 of domain B. This latter feature might be key to the selection and precise orientation of the inter-domain docking necessary for the specific phosphodiester cleavage. The model provides an important basis for further studies of hairpin ribozyme structure and function.

Computer Simulation↗
PubMed1997

Identification and characterization of the functional amino acids at the active site of the large thioredoxin reductase from Plasmodium falciparum.

The thioredoxin system, composed of the pyridine nucleotide-disulfide oxidoreductase thioredoxin reductase, the small peptide thioredoxin, and NADPH as a reducing cofactor, is one of the major thiol-reducing systems of the cell. Recent studies revealed that Plasmodium falciparum and human thioredoxin reductase represent a novel class of enzymes, called large thioredoxin reductases. The large thioredoxin reductases are substantially different from the isofunctional prokaryotic Escherichia coli enzyme. The putative essential amino acids at the catalytic center of large thioredoxin reductase from P. falciparum were determined by using site-directed mutagenesis techniques. To analyze the putative active site cysteines (Cys88 and Cys93) three mutant proteins were constructed substituting alanine or serine residues for cysteine residues. Further, to evaluate the function of His509 as a putative proton donor/acceptor of large thioredoxin reductase this residue was replaced by either glutamine or alanine. All mutants were expressed in the E. coli system and characterized. Steady state kinetic analysis revealed that the replacement of Cys88 by either alanine or serine and Cys93 by alanine resulted in a total loss of enzymatic activity. These results clearly identify Cys88 and Cys93 as the active site thiols of large thioredoxin reductase. The replacement of His509 by glutamine yielded in a 95% loss of thioredoxin reductase activity; replacement by alanine provoked a loss of 97% of enzymatic activity. These results identify His509 as active site base, but imply that its function can be substituted, although inefficiently, by an alternative proton donor, similar to glutathione reductase. Spectral analysis of wild-type P. falciparum thioredoxin reductase revealed a 550-nm absorption band upon reduction which resembles the EH2 form of glutathione reductase and lipoamide dehydrogenase. This spectral feature, recently also reported for the human placenta protein (Arscott, L. D., Gromer, S., Schirmer, R. H., Becker K., and Williams, C. H., Jr. (1997) Proc. Natl. Acad. Sci. U. S. A. 94, 3621-3626), further illustrates the similarity between large thioredoxin reductases and glutathione reductases and stresses the profound differences to small E. coli thioredoxin reductase.

Animals↗
PubMed1997

Localization of O-glycosylation sites on glycopeptide fragments from lactation-associated MUC1. All putative sites within the tandem repeat are glycosylation targets in vivo.

Since there is no consensus sequence directing the initial GalNAc incorporation into mucin peptides, O-glycosylation sites are not reliably predictable. We have developed a mass spectrometric sequencing strategy that allows the identification of in vivo O-glycosylation sites on mucin-derived glycopeptides. Lactation-associated MUC1 was isolated from human milk and partially deglycosylated by trifluoromethanesulfonic acid to the level of core GalNAc residues. The product was fragmented by the Arg-C-specific endopeptidase clostripain to yield tandem repeat icosapeptides starting with the PAP motif. PAP20 glycopeptides were subjected to sequencing by post-source decay matrix-assisted laser desorption ionization mass spectrometry or by solid phase Edman degradation to localize the glycosylation sites. The masses of C- or N-terminal fragments registered for the mono- to pentasubstituted PAP20 indicated that GalNAc was linked to the peptide at Ser5,Thr6 (GSTA) and Thr14 (VTSA) but contrary to previous in vitro glycosylation studies also at Thr19 and Ser15 located within the PDTR or VTSA motifs, respectively. Quantitative data from solid phase Edman sequencing revealed no preferential glycosylation of the threonines. These discrepancies between in vivo and in vitro glycosylation patterns may be explained by assuming that O-glycosylation of adjacent peptide positions is a dynamically regulated process that depends on changes of the substrate qualities induced by glycosylation at vicinal sites.

Acetylgalactosamine↗
PubMed1997

Quantitative tissue Doppler in comparison with two-dimensional and Doppler echocardiographic indices in normal subjects.

To assess normal values of left ventricular wall motion velocities with respect to sequential phases of the cardiac cycle 95 normal subjects (mean age 34+/-12 years) were examined using a recently developed method based on the computer software TDI-LaborR enabling a variable time and space related measurement of regional wall velocity from tissue Doppler (TD) images. The study population consisted of three age groups: 16-29 years (group 1, n=34 (36%)), 30-49 years (group 2, n=45 (49%)), >50 years (group 3, n=16 (17%)). Using TD M-mode of the left ventricular posterior wall the mean subendocardial and subepicardial velocity as well as the transmural myocardial velocity were determined for the rapid, slow and atrial filling phases as well as the systole. In addition, standard mitral inflow parameters-E and A velocities were determined by pulsed Doppler echocardiography. The left parasternal window was used. The subendocardial rapid filling and atrial filling myocardial velocity were found to be age significantly dependent. The myocardial rapid filling were correlated to early diastolic mitral inflow velocity and to the ratio of early and atrial flow velocities (r ranges from 0.23 to 0.36, P<0.01). We conclude that aging is accompanied by an increase of atrial filling compensating a reduced myocardial relaxation already before 50 years. Whereas mitral inflow velocities reflect diastolic function indirectly, it is directly indicated by TD myocardial velocity parameters.

Adolescent↗
PubMed1997

Molecular characterization and expression of Onchocerca volvulus glutathione reductase.

Glutathione metabolism represents a potential target for anti-parasite drug design. The central role of glutathione reductase (GR) in maintenance of the thiol redox state and in anti-oxidative defence has to be evaluated in more detail in order to establish the essential function of this enzyme for the survival of the filarial parasite Onchocerca volvulus. The O. volvulus GR (OvGR) gene was cloned and sequenced. The gene is composed of 13 exons and 12 introns and spans 4065 bp. The first intron is located within the 5'-untranslated region of the gene, 16 nucleotides upstream of the translation initiation codon. Southern-blot analysis and structural characterization of the genomic sequence indicate that OvGR is encoded by a single-copy gene. Isolation of various cDNA clones revealed a polymorphism of polyadenylation initiation with no consensus polyadenylation sites in any of the cDNAs analysed. The entire cDNA is 1977 bp long and carries the nematode-specific spliced leader sequence SL1 at its 5' end, 236 nucleotides upstream of the first in-frame methionine. The cDNA codes for a polypeptide of 462 amino acids with 53.5% sequence identity with human GR (HsGR). A total of 18 out of 19 residues contributing to glutathione binding are identical in OvGR and HsGR. However, one of the arginine residues (Arg-224 in HsGR) involved in discrimination between NADPH and NADH in all known GRs is substituted by tryptophan (Trp-207 in OvGR). The coding region of OvGR was expressed in Escherichia coli as a histidine-fusion protein, and it was established that the parasite protein still favours the binding of NADPH (Km 10.9 microM) over NADH (Km 108 microM). The histidine-fusion protein has a subunit size of 54 kDa and is active as a homodimer of 110 kDa.

Amino Acid Sequence↗
PubMed1997

Randomized comparison of angioplasty of complex coronary lesions at a single center. Excimer Laser, Rotational Atherectomy, and Balloon Angioplasty Comparison (ERBAC) Study.

BACKGROUND: The purpose of this study was to test whether coronary revascularization with ablation of either excimer laser or rotational atherectomy can improve the initial angiographic and clinical outcomes compared with dilatation (balloon angioplasty) alone. METHODS AND RESULTS: At a single center, a total of 685 patients with symptomatic coronary disease warranting elective percutaneous revascularization for a complex lesion were randomly assigned to balloon angioplasty (n = 222), excimer laser angioplasty (n = 232), or rotational atherectomy (n = 231). The primary end point was procedural success (diameter stenosis < 50%, absence of death, Q-wave myocardial infarction, or coronary artery bypass surgery). The patients who underwent rotational atherectomy had a higher rate of procedural success than those who underwent excimer laser angioplasty or conventional balloon angioplasty (89% versus 77% and 80%, P = .0019), but no difference was observed in major in-hospital complications (3.2% versus 4.3% versus 3.1%, P = .71). At the 6-month follow-up, revascularization of the original target lesion was performed more frequently in the rotational atherectomy group (42.4%) and the excimer laser group (46.0%) than in the angioplasty group (31.9%, P = .013). CONCLUSIONS: Procedural success of rotational atherectomy is superior to laser angioplasty and balloon angioplasty; however, it does not result in better late outcomes. The role of plaque debulking before balloon dilatation in percutaneous coronary revascularization remains to be fully defined.

Adult↗
PubMed1997

Degradation of T cell receptor (TCR)-CD3-zeta complexes after antigenic stimulation.

T cell activation by specific antigen results in a rapid and long-lasting downregulation of triggered T cell receptors (TCRs). In this work, we investigated the fate of downregulated TCR- CD3-zeta complexes. T cells stimulated by peptide-pulsed antigen-presenting cells (APCs) undergo an antigen dose-dependent decrease of the total cellular content of TCR-beta, CD3-epsilon, and zeta chains, as detected by FACS(R) analysis on fixed and permeabilized T-APC conjugates and by Western blot analysis on cell lysates. The time course of CD3-zeta chain consumption overlaps with that of TCR downregulation, indicating that internalized TCR-CD3 complexes are promptly degraded. Inhibitors of lysosomal function (bafilomycin A1, folimycin) markedly reduced zeta chain degradation, leading to the accumulation of zeta chain in large Lamp1(+) vesicles. These results indicate that in T cell-APC conjugates, triggered TCRs are rapidly removed from the cell surface and are degraded in the lysosomal compartment.

Amino Acid Sequence↗
PubMed1997

Purification, structural analysis, and function of natural ATAC, a cytokine secreted by CD8(+) T cells.

Recently, we identified a novel putative human cytokine expressed by activated CD8(+) T cells, which was designated ATAC (activation-induced, T cell-derived, and chemokine-related; the same molecule has been identified independently as lymphotactin and single cysteine motif-1). In this report, we provide evidence that ATAC is a secreted 93-amino acid protein that is generated from its precursor by proteolytic cleavage between Gly21 and Val22. An estimated 60% of ATAC (Val22-Gly114) is secreted as an unmodified protein with a molecular mass of 10,271.72 Da (apparent molecular mass of 12 kDa in SDS-polyacrylamide gel electrophoresis) and in which Cys32 and Cys69 are linked by a disulfide bridge. Unmodified ATAC is a cationic protein with a pI of 11.35 and is capable of binding to heparin. Some 40% of ATAC is O-glycosylated within 25 min of synthesis, giving rise to the appearance of a homogeneous 15-kDa (minor fraction) and a heterogeneous, terminally sialylated 17-19-kDa (major fraction) protein species in SDS-polyacrylamide gel electrophoresis. The secretion of all ATAC protein variants is completed within 30-40 min of synthesis. In terms of function, various ATAC protein forms were consistently ineffective in chemotaxis assays. In contrast, both purified natural ATAC and a chemically synthesized aglycosyl analog induced locomotion (chemokinesis) in purified CD4(+) and CD8(+) T cell populations at 400 ng/ml.

CD4-Positive T-Lymphocytes↗
PubMed1997

Comparative chromosome painting in mammals: human and the Indian muntjac (Muntiacus muntjak vaginalis).

We have used human chromosome-specific painting probes for in situ hybridization on Indian muntjac (Muntiacus muntjak vaginalis, 2n = 6, 7) metaphase chromosomes to identify the homologous chromosome regions of the entire human chromosome set. Chromosome rearrangements that have been involved in the karyotype evolution of these two species belonging to different mammalian orders were reconstructed based on hybridization patterns. Although, compared to human chromosomes, the karyotype of the Indian muntjac seems to be highly rearranged, we could identify a limited number of highly conserved homologous chromosome regions for each of the human chromosome-specific probes. We identified 48 homologous autosomal chromosome segments, which is in the range of the numbers found in other artiodactyls and carnivores recently analyzed by chromosome painting. The results demonstrate that the reshuffling of the muntjac karyotype is mostly due to fusions of huge blocks of entire chromosomes. This is in accordance with previous chromosome painting analyses between various Muntjac species and contrasts the findings for some other mammals (e.g., gibbons, mice) that show exceptional chromosome reshuffling due to multiple reciprocal translocation events.

Animals↗
PubMed1997

Selectivity of beta-adrenergic receptor kinase 2 for G protein betagamma subunits.

Phosphorylation of G protein-coupled receptors by beta-adrenergic receptor kinases (betaARK) requires the presence of G protein betagamma subunits. We have investigated the ability of the two betaARK isoforms to distinguish between defined recombinant betagamma subunits. betaARK2 had an about 25% lower specific activity than betaARK1 towards rhodopsin and the beta2-adrenergic receptor but the two kinases shared the selectivity for betagamma subunits: betagamma complexes consisting of beta1 or beta2 in combination with gamma2, gamma5, and gamma7 were more efficacious than those with gamma3 or beta1 gamma1. Thus, while betaARKs differentiate between defined betagamma subunits, betagamma complexes do not discriminate between betaARK isoforms.

Animals↗
PubMed1997

Cortical reorganization in patients with facial palsy.

Possible changes in the organization of the cortex in patients with facial palsy, serving as a model of peripheral motor deefferentation, were investigated by using transcranial magnetic stimulation (TMS) and positron emission tomography (PET). With TMS, the size of the area producing muscle-evoked potentials (MEPs) of the abductor pollicis brevis muscle, the sum of MEP amplitudes within this area, and the volume over the mapping area were compared between both hemispheres in 8 patients. With PET, increases in regional cerebral blood flow, measured with the standard H2(15)O2 bolus injection technique, were compared between 6 patients and 6 healthy volunteers during sequential finger opposition. Patients moved the hand ipsilateral to the facial palsy, the control subjects the right hand. Of 9 patients in total, 5 participated in both experiments. With both methods, an enlargement of the hand field contralateral to the facial palsy was found, extending in a lateral direction, into the site of the presumed face area. The PET data showed that the enlargement of the hand field in the somatosensory cortex (SMC) is part of a widespread cortical reorganization, including the ipsilateral SMC and bilateral secondary motor and sensory areas. We report for the first time, using two different noninvasive methods, that peripheral, mere motor deefferentation is a sufficient stimulus for reorganizational changes in the healthy adult human cortex.

Adaptation, Physiological↗