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Biomedical subjects

B Müller

Publications and source records attributed to B Müller.

At least 235 records · Page 13Linked to original sources
PubMed1995

Monoclonal antibodies against Rous sarcoma virus integrase protein exert differential effects on integrase function in vitro.

We have prepared and characterized several monoclonal antibodies (MAbs) against the Rous sarcoma virus integrase protein (IN) with the aim of employing these specific reagents as tools for biochemical and biophysical studies. The interaction of IN with the purified MAbs and their Fab fragment derivatives was demonstrated by Western blot (immunoblot), enzyme-linked immunosorbent assay, and size exclusion chromatography. A series of truncated IN proteins was used to determine regions in the protein important for recognition by the antibodies. The MAbs described here recognize epitopes that lie within the catalytic core region of IN (amino acids 50 to 207) and are likely to be conformational. A detailed functional analysis was carried out by investigating the effects of Fab fragments as well as of intact MAbs on the activities of IN in vitro. These studies revealed differential effects which fall into three categories. (i) One of the antibodies completely neutralized the processing as well as the joining activity and also reduced the DNA binding capacity as determined by a nitrocellulose filter binding assay. On the other hand, this MAb did not abolish the cleavage-ligation reaction on a disintegration substrate and the nonspecific cleavage of DNA by IN. The cleavage pattern generated by the IN-MAb complex on various DNA substrates closely resembled that produced by mutant IN proteins which show a deficiency in multimerization. Preincubation of IN with substrate protected the enzyme from inhibition by this antibody. (ii) Two other antibodies showed a general inhibition of all IN activities tested. (iii) In contrast, a fourth MAb stimulated the in vitro joining activity of IN. Size exclusion chromatography demonstrated that IN-Fab complexes from representatives of the three categories of MAbs exhibit different stoichiometric compositions that suggest possible explanations for their contrasting effects and may provide clues to the relationship between the structure and function of IN.

Animals↗
PubMed1995

Estimate of severe autosomal recessive limb-girdle muscular dystrophy (LGMD2C, LGMD2D) among sporadic muscular dystrophy males: a study of 415 familes.

Ninety-five percent of cases of severe muscular dystrophy with early childhood onset result from mutations in the dystrophin region of the human X chromosome (DMD, McKusick 310200), whereas 5% are thought to result from mutations in autosomal genes. We examined a total of 415 families with at least one living patient whose clinical features suggested DMD. Based on formal genetics, haplotype analysis, and dystrophin determinations, we estimate that one in eight (11.8%) sporadic male patients carries autosomal rather than X chromosomal mutations.

Chromosome Aberrations↗
PubMed1995

Oguchi disease: suggestion of linkage to markers on chromosome 2q.

Oguchi disease is a rare autosomal recessive form of congenital stationary night blindness. The condition is associated with fundus discolouration and abnormally slow dark adaptation. Earlier studies suggested that the 48 kD protein S antigen may be involved in the recovery phase of light transduction. Previous cytogenetic and linkage studies have localised the S antigen gene (SAG) to chromosome 2q37.1. In the present study markers which map to distal chromosome 2q were typed in an inbred Oguchi pedigree. The segregation data obtained suggested that the affected subjects are homozygous by descent for a region between D2S172 and D2S345. An intragenic SAG polymorphism was homozygous in all affected people and a recombination event suggested that SAG maps proximal to D2S345. Collectively, these findings support the hypothesis that a defect in S antigen may be responsible for Oguchi disease.

Antigens↗
PubMed1995

Quantitative analysis of parenchymal and vascular alterations in NO2-induced lung injury in rats.

Nitrogen dioxide (NO2), the oxidation product of nitric oxide (NO), is a reactive free radical forming gas, the inhalation of which has been reported to induce severe damage to distal airways. In order to quantify dose and time course of parenchymal and vascular damage, rats were exposed to 5, 10 and 20 ppm NO2 for 3 and 25 days, followed by quantitative histology and morphometry of the lung. Histological investigations of the short-term exposed animals showed structural alterations extending from slight interstitial oedema after exposure to 5 ppm, to epithelial necrosis and interstitial inflammatory infiltration after exposure to 10 ppm, and an additional intra-alveolar oedema after 20 ppm. The pulmonary arteries disclosed no qualitative changes, such as muscularization of intra-acinar vessels. Long-term exposure to 10 ppm and 20 ppm NO2 resulted in emphysema and slight centrilobular interstitial fibrosis. Morphometric analysis revealed the alveolar surface density to be significantly diminished after short-term exposure to 20 ppm NO2 and long-term exposure to 10 and 20 ppm NO2. The medial thickness of pulmonary arteries was significantly increased after short- and long-term exposure to 20 ppm NO2 and long-term exposure to 10 ppm NO2. In the 5 ppm short- and long-term exposure groups the pulmonary arterial medial thickness was significantly decreased compared to controls. Correlation analysis revealed a negative correlation between average medial thickness and alveolar surface density (coefficient of correlation: -0.56). We conclude that the extent of NO2-induced pulmonary parenchymal and vascular alterations are closely related and concentration- and time-dependent.

Animals↗
PubMed1995

Variations of steroid hormone metabolites in serum and urine in polycystic ovary syndrome after nafarelin stimulation: evidence for an altered corticoid excretion.

To evaluate the clinical relevance of testing pituitary-ovarian responses in patients suffering from polycystic ovary syndrome (PCOS) with the GnRH agonist nafarelin, a 1.2-mg dose of nafarelin was given intranasally to 19 women with PCOS and 15 healthy premenopausal women. The subsequent analysis of steroids in both serum and urine during the test was carried out at several time points for up to 24 h. Serum levels of 17 alpha-hydroxyprogesterone were elevated at all time points of the test in PCOS patients vs. controls [at baseline, 3.5 +/- 0.2 vs. 1.8 +/- 0.1 nmol/L (P < 0.001); at 24 h, 9.9 +/- 0.9 vs. 4.9 +/- 0.3 nmol/L (P < 0.001)]. Basal levels of androstenedione were higher in the patient group, but there was no significant change during the test in either group. Serum testosterone levels were also found to differ in PCOS patients compared with the control values at baseline (2.2 +/- 0.2 vs. 1.5 +/- 0.1 nmol/L; P < 0.05) and after nafarelin treatment (at 24 h, 3.2 +/- 0.4 vs. 1.8 +/- 0.2 nmol/L; P < 0.05). Serum estradiol levels rose significantly in both groups during the test; the posttest levels were significantly higher in PCOS than in controls. The PCOS patients displayed a significant increase in androgen and gestagen metabolites as well as in glucocorticoid metabolites excreted in the urine during the 24 h. In the control subjects, except for 17 alpha-hydroxypregnanolone, which rose significantly, none of the urinary steroids investigated showed relevant changes during the nafarelin test. The posttest excretion of allo-tetrahydrocortisol (1.4 +/- 0.2 vs. 0.3 +/- 0.1 mumol/g creatinine; P < 0.001) and the increase in 17 alpha-hydroxypregnanolone excretion (1.4 +/- 0.2 vs. 0.3 +/- 0.1 mumol/g creatinine; P < 0.001) were distinctly higher in PCOS patients than in the controls; the diagnostic sensitivity of the combination of both parameters was 89% at a 93% specificity. Thus, measurements of 17 alpha-hydroxyprogesterone levels in serum and of urinary allo-tetrahydrocortisol and 17 alpha-hydroxypregnanolone after nafarelin treatment make this stimulation test a valuable diagnostic tool for identifying PCOS patients. The significant changes in the excretion of urinary androgen and gestagen metabolites, unmasked by GnRH agonist stimulation, suggest a functional alteration of the pituitary-ovarian axis. The reason for the increased excretion of glucocorticoid metabolites after nafarelin stimulation remains to be clarified.

Administration, Intranasal↗
PubMed1995

Autosomal dominant exudative vitreoretinopathy: linkage analysis and its clinical application.

Close linkage without recombination (Zmax = 7.383 at Theta = 0.00) was found between the locus for autosomal dominant exudative vitreoretinopathy (Criswick-Schepens) and the locus D11S388 in 11q14.3-q21. We report on the application of data from linkage studies in the diagnostic management of this disease. To determine the disease risk for two newborn children of two affected mothers more precisely, indirect genotype analysis was performed by typing their DNA for three DNA polymorphisms, the loci of which have been shown to be closely linked to the locus of autosomal dominant exudative vitreoretinopathy.

Adult↗
PubMed1994

Theoretical and experimental epitope mapping of thymosin beta 4.

Two rabbit polyclonal antisera, one directed against thymosin beta 4 and the other one against the peptide fragment thymosin beta 4 (1-14) were characterised by epitope mapping. Hexapeptides representing the whole sequence of the native peptide and overlapping by one amino acid were synthesised on polystyrene pins. The antigenic determinants were identified in microtitre plates with an ELISA procedure. The polyclonal serum against thymosin beta 4 detected three epitopes (N-terminal, mid-region and C-terminal) whereas the polyclonal serum against the fragment contained only antibodies specific for the N-terminal epitope. These experimental results are consistent with theoretical predictions.

Amino Acid Sequence↗
PubMed1994

Selective recognition of cyclic RGD peptides of NMR defined conformation by alpha IIb beta 3, alpha V beta 3, and alpha 5 beta 1 integrins.

The binding of purified fibrinogen receptor alpha IIb beta 3, vitronectin receptor alpha V beta 3, and fibronectin receptor alpha 5 beta 1 to their corresponding ligands in solid-phase binding assays was used to examine the inhibitory activity of various linear and cyclic penta- and hexapeptides of different conformation containing RGD or RAD sequences. Cyclic peptides with different defined backbone conformations were designed by introducing a single D-amino acid or a proline at different positions in the ring. The data were calibrated for alpha IIb beta 3 integrin incorporated into a planar lipid bilayer by a physical method (total internal reflection fluorescence microscopy) which yielded KD = 1.7 microM for a linear RGD peptide and KD = 0.03 microM for fibrinogen. With this integrin, three cyclic hexapeptides ([GRGDFL], [ARGDFV], [GRGDFV]) were 2-4-fold more inhibitory than the linear GRGDS peptide in solid-phase assays and showed similar inhibition as the fibrinogen ligand. Six peptides had the same or a 2-fold lower activity as the linear reference peptide, and three peptides were up to 7-fold less active. Replacement of Arg or Asp by their stereoisomers or Gly by Ala resulted in a 100-1000-fold reduction in activity. With the two other integrins, a single cyclic pentapeptide [RGDFV] was 10-fold more active (alpha V beta 3) or equal in activity (alpha 5 beta 1) to linear GRGDS, while all of the other cyclic peptides were moderately or distinctly less active. Changes in the RGD sequence caused a less dramatic reduction in binding strength for alpha V beta 3 and alpha 5 beta 1 than for alpha II beta 3. Inhibitory activity was compared with the distance between the C beta atoms of Arg and Asp residues as determined by NMR and indicated that the optimum distance is in the range of 0.75-0.85 nm for alpha IIb beta 3 and at or below 0.67 nm for alpha V beta 3 and alpha 5 beta 1. This indicates that alpha IIb beta 3 less sensitive to variations in the RGD backbone structure and can accommodate a larger distance than the integrins alpha V beta 3 and alpha 5 beta 1.

Amino Acid Sequence↗
PubMed1994

Assay of zatebradine in plasma by fully automated sample clean-up, capillary gas chromatography and ammonia chemical ionisation mass spectrometry.

A method has been developed for the measurement of zatebradine (UL-FS 49), a heart-rate lowering drug, suitable for the treatment of stable angina pectoris. The method comprises a fully automated liquid-solid extraction using a Zymark Benchmate, a capillary gas chromatography and ammonia chemical ionisation (CI) mass spectrometry using hexadeuterated zatebradine for the internal standard. The assay has a mean between-batch imprecision of 4.9% and a mean inaccuracy of 1.5%. The calibration curve covers the range of 1-30 ng/ml. About 60 samples can be handled per day. The assay has been successfully applied to human pharmacokinetic studies.

Ammonia↗
PubMed1994

[Evaluation of advertisements of weight-reducing cures in the press].

The aim of this study was to obtain an overview of the weight control programmes advertised in the Swiss press and to explore them in terms of their scientific basis and success rates. 875 advertisements and questionnaires from 507 patients and 52 advertisers were analyzed. Scientifically questionable methods usually remained ineffective. In contrast, methods based on a reasonably balanced long term diet proved more successful. With the latter, an average weight loss of 15 kg was achieved in 32 weeks at a cost of Sfr. 456.-. In doing so, 56.1% of the patients reported they had reached their target weight and it was entirely or partly maintained for a prolonged period by 39.6%. In terms of cost/benefit ratios, scientifically based programmes which follow the principles of physiology and behavioural psychology are the most favourable, and physicians should know and recommend them. Unfortunately, in Switzerland health insurance still pays for programmes which were shown to be unsuccessful in the long term, such as appetite lowering medication, or costly, such as short-term courses of treatment in hotels.

Advertising↗
PubMed1994

Best's vitelliform dystrophy (VMD2) maps between D11S903 and PYGM: no evidence for locus heterogeneity.

Vitelliform macular dystrophy, also known as Best's disease (BD), is an autosomal dominant disorder typically characterized by an accumulation of yellowish material in the macular area. The disease is slowly progressive and eventually results in atrophy of the retinal pigment epithelium and photoreceptor cells, thus severely impairing central vision. The biochemical defect underlying this condition is unknown. More recently, the BD locus (VMD2) was mapped to chromosome 11 by genetic linkage to microsatellite markers at D11S871 and INT2. In the present study, we report a detailed genetic analysis in three multigeneration Best's disease families using eight microsatellite markers spanning approximately 26 cM around the putative BD locus. We demonstrate linkage between Best's disease and the markers used. Furthermore, haplotype analysis in our unrelated Best's disease families identified three distinct haplotypes associated with the disease, strongly suggesting independent origins of the BD mutation. Finally, we characterized two recombinant BD chromosomes that significantly refine the location of the disease gene to a 3.7-cM interval between markers at D11S903 and PYGM. PCR-hybrid mapping sublocalized this interval to the pericentromeric region of chromosome 11.

Chromosome Mapping↗
PubMed1994

Mapping of the autosomal dominant exudative vitreoretinopathy locus (EVR1) by multipoint linkage analysis in four families.

Autosomal dominant exudative vitreoretinopathy is a disorder affecting primarily the development of the human retinal vascular system. The disease locus has recently been assigned to 11q13-q23 by linkage studies in two families. Two-point analysis on a total of four families has now revealed close linkage (Zmax = 8.34 at theta = 0.00) between the disease locus and D11S873. Multipoint linkage analysis mapped the disease locus between D11S527/D11S533 and D11S35 with a maximum lod score of over 11 directly at D11S873. No evidence appeared for genetic/linkage heterogeneity among the four families examined.

Chromosome Mapping↗
PubMed1994

Processing of Holliday junctions by the Escherichia coli RuvA, RuvB, RuvC and RecG proteins.

Recent work has led to significant advances in our understanding of the late steps of genetic recombination and the post-replicational repair of DNA. The RuvA and RuvB proteins have been shown to interact with recombination intermediates and catalyse the branch migration of Holliday junctions. Although both proteins are required for branch migration, each plays a defined role with RuvA acting as a specificity factor that directs RuvB (an ATPase) to the junction. The RuvB ATPase provides the motor for branch migration. The next step is catalysed by RuvC protein which recognises Holliday junctions and promotes their resolution by endonucleolytic cleavage. New data indicates an alternative pathway for Holliday junction processing. This pathway involves RecG, a branch migration protein which is functionally analogous to RuvAB, and a protein (activated by a rus mutation) which works with RecG to process intermediates independently of RuvA, RuvB and RuvC.

Adenosine Triphosphatases↗
PubMed1994

Purification and analytical characterization of an anti-CD4 monoclonal antibody for human therapy.

A purification process for the monoclonal anti-CD4 antibody MAX.16H5 was developed on an analytical scale using (NH4)2SO4 precipitation, anion-exchange chromatography on MonoQ or Q-Sepharose, hydrophobic interaction chromatography on phenyl-Sepharose and gel filtration chromatography on Superdex 200. The purification schedule was scaled up and gram amounts of MAX.16H5 were produced on corresponding BioPilot columns. Studies of the identity, purity and possible contamination by a broad range of methods showed that the product was highly purified and free from contaminants such as mouse DNA, viruses, pyrogens and irritants. Overall, the analytical data confirm that the monoclonal antibody MAX.16H5 prepared by this protocol is suitable for human therapy.

Animals↗