PubMed Health⌕ Search

Biomedical subjects

U R Müller

Publications and source records attributed to U R Müller.

At least 37 records · Page 2Linked to original sources
PubMed1995

Type I skin reactivity to native and recombinant phospholipase A2 from honeybee venom is similar.

Phospholipase A2 is the major allergen in honeybee venom. Recombinant phospholipase A2 was produced in prokaryotes and tested for its biologic activity by intracutaneous skin testing with serial 10-fold dilutions in comparison with native and deglycosylated phospholipase A2 in patients allergic to bee venom. Linear regressions of the log of the wheal area versus the log of the allergen concentration were calculated for all allergens in each patient. The relative allergenic potency of the various preparations was analyzed by comparing the linear regressions. Native phospholipase A2 was about 10 times more potent than whole bee venom. None of 58 patients allergic to bee venom was missed by testing with native phospholipase A2 alone. This allergen and deglycosylated native phospholipase A2 resulted in similar skin reactions, indicating that the sugar residues were of little relevance for IgE-binding in the patients tested. Native phospholipase A2 also had relative potency similar to that of recombinant refolded phospholipase A2, whereas recombinant nonrefolded phospholipase A2 had almost no biologic activity in skin testing. These results demonstrate in vivo activity of the recombinant bee venom allergen phospholipase A2. Although correct refolding is a prerequisite for type I skin reactivity, glycosylation seems to be less important.

Adult↗
PubMed1995

Ultra rush bee venom immunotherapy does not reduce cutaneous weal responses to bee venom and codeine phosphate.

BACKGROUND: The rapid administration of bee venom in cumulative doses exceeding the quantity contained in one bee sting is well tolerated by most of the patients during 3.5 h of ultra-rush bee venom immunotherapy (VIT). The mechanism of this tolerance is unknown. OBJECTIVE: The aim of the study was to verify the hypothesis that either slow mediator depletion of mast cells or blockade of their surface receptor mechanisms by increasing doses of allergen might be the major mechanisms of tolerance induced by ultra-rush VIT. METHODS: Nine bee venom allergic patients with a history of severe systemic reactions after a bee sting, positive skin tests and bee venom specific serum IgE antibodies were treated as follows: on the first day a cumulative dose of 111 micrograms was administered over 3.5 h under intensive care conditions. Further injections were given on day 7, day 21 and thereafter at 4 week intervals. Intradermal tests with codeine phosphate (non-specific mast cell degranulation) and bee venom were performed before the initiation of VIT and 30 min after the last injection on the same day as well as before the subsequent bee venom injections. RESULTS: No significant changes of skin reactivity to both codeine phosphate and bee venom were observed on day 1 (before initiation of VIT and after the last injection on the same day). CONCLUSIONS: Ultra-rush VIT does not induce mediator depletion or surface receptor blockade in skin mast cells.

Bee Venoms↗
PubMed1995

Selective restimulation of antigen or allergen preactivated T cells using OKT3 F(ab)2 results in the secretion of TH-1 or TH-2-like cytokine patterns.

BACKGROUND: The synthesis of IgE is regulated by cytokines secreted from T-helper cells. The studies on cytokine secretion by peripheral blood mononuclear cells (PBMC) upon stimulation with antigen or allergen are difficult due to low levels of cytokines, especially of interleukin-4 (IL-4). OBJECTIVE: In this study we tried to establish a culture system, which could enable the measurement of the cytokine profiles in specifically activated cultures. METHODS: Three methods to potentiate cytokine secretion were evaluated: PBMC from bee venom or house dust mite (Dermatophagoides pteronyssinus) allergic patients as well as normal subjects were stimulated either with the major bee venom allergen phospholipase A2 (PLA) or with the major D. pteronyssinus allergen (Der p 1) or with the control antigens tetanus toxoid (TT) and purified protein derivate (PPD). After 7 days of culture the cells were restimulated either with plastic bound OKT3 F(ab)2 monoclonal antibodies (MoAbs), with the appropriate antigen + antigen presenting cells or with IL-2. The secretion of cytokines (IL-4, IFN gamma) was measured after restimulation of the cultures (day 8). RESULTS: While OKT3 F(ab)2 was unable to activate resting T cells, it could restimulate preactivated cells. Restimulation with OKT3 F(ab)2 induced higher IL-4 and IFN gamma secretion than restimulation with IL-2 or antigen. TT and PLA stimulated a similar cytokine secretion profile in normal and PLA allergic subjects with substantial levels of both IL-4 and IFN gamma. In contrast, PPD induced virtually only IFN gamma secretion. Der p 1 stimulated mainly IL-4 secretion but also IFN gamma production in some mite allergic patients. CONCLUSION: We have established a cell culture system, which combines antigen specificity with a strong cytokine inducing signal provided by anti-CD3 MoAbs. TH-1 and TH-2 characteristic cytokine patterns can be observed in short-term PBMC cultures already after 8 days of culture.

Allergens↗
PubMed1994

[Insect venom allergy].

Systemic allergic reactions to hymenoptera stings are most often mediated by venom-specific IgE antibodies. The most frequent symptoms are urticaria, angioedema, asthma and anaphylactic shock. Rarely, cerebral or myocardial ischemia with permanent damage may result from severe anaphylactic shock. Each year several individuals die in Switzerland from allergic reactions following hymenoptera stings. The most valuable diagnostic tests are skin tests and estimation of venom-specific serum-IgE antibodies. The most important drugs for emergency treatment are adrenaline and antihistamines. During anaphylactic shock, volume substitution may be essential. All patients with a history of severe systemic reactions should receive venom immunotherapy which induces complete protection in a high percentage of the patients.

Acute-Phase Proteins↗
PubMed1994

[The value of immunotherapy in treatment of IgE-induced allergic diseases].

Until today, immunotherapy is the only causal treatment of IgE-mediated allergic disease. Its efficacy has been demonstrated in controlled studies for rhinitis and bronchial asthma due to various pollens, house-dust mite and animal dander as well as for systemic allergic reactions following hymenoptera stings. This treatment is especially successful in younger patients with seasonal allergy of rather short duration and with sensitization only to few allergens. In order to avoid allergic side effects of this treatment, measures of precautions must be strictly observed. A close cooperation between patient, family practitioner and allergist is important for the success of this treatment.

Allergens↗
PubMed1991

Extraction and differentiation of the Su autoantigen from calf thymus nuclei.

Antibodies to Su antigen have been reported previously as a distinct antigen-antibody system associated with connective tissue diseases; most specifically systemic lupus erythematosus and undifferentiated connective tissue disease. The Su antigen was first identified by double immunodiffusion using calf thymus nuclear extract (CTNE) as a source for Su antigen. In this report, enhanced extraction of Su antigen was achieved using deoxyribonuclease I (DNase) for preparation of CTNE. Only the Sm antigen was found in comparable quantities in the DNase CTNE. Western immunoblotting and immunoprecipitation employing DNase CTNE and extracts of [35S]methionine-labeled HeLa cells respectively were used for further characterization and differentiation of the Su antigen. Sera from patients positive for Su antibodies by double immunodiffusion were found to react most specifically with antigen components in a molecular weight range of approximately 50-55 kDa. These methods should assist in further understanding the biochemical properties of the Su antigen.

Ammonium Sulfate↗
PubMed1990

Effect of in vitro transcription on cruciform stability.

We have investigated the effect of in vitro transcription on cruciform stability. Replicative form DNA of phiX174 strain ins6240, containing a 48 bp synthetic palindrome in the J-F intercistronic region, was supercoiled in vitro to mean negative superhelical densities (sigma) ranging from 0 to 0.15. The presence of cruciforms was probed by limited digestion with the single-strand specific nuclease Bal31. The 48 bp palindrome was extruded at a mean sigma = -0.05, but only after heating the DNA. An in vitro transcription reaction with E. coli RNA polymerase and [alpha-32P]UTP gave identical transcripts with heated or unheated template DNA. The synthetic cruciform was stable upon binding of the RNA polymerase to the template, but it was destabilized upon movement of the transcription complex along the template. Transcription of unheated templates did not result in cruciform formation. We propose that cruciform structures in supercoiled template DNAs present no hindrance to RNA polymerase, and thus have no detectable effect on transcription elongation in vitro.

Bacteriophage phi X 174↗
PubMed1988

Effects of palindromes on in vivo DNA replication and mutagenesis in bacteriophage phi X174 RF DNA.

Bacteriophage phi X174 mutants with insertions of palindromic DNA sequences are rapidly outgrown by competing wild-type phage (Müller & Turnage, J. Mol. Biol. 189: 285). The basis for this defect was investigated and found to be due to an exclusion event early in the infectious cycle, in which phage genomes with palindromic inserts were preferentially excluded by wt phage. In addition, we have obtained further evidence for a palindrome induced genetic instability. Both defects are dependent on palindrome size and sequence, consistent with a model which involves formation of cruciforms, or cruciform-like structures. We propose that formation of unusual DNA secondary structures reduces the effectiveness of replicative form (RF) DNA to interact with limiting replication factors or membrane binding sites, possibly because of interaction with the host recombination system.

Bacteriophage phi X 174↗
PubMed1987

Effects of palindrome size and sequence on genetic stability in the bacteriophage phi X174 genome.

By inserting palindromes of varying length and sequence into a non-essential region of the bacteriophage phi X174 genome we have investigated the effect of palindrome size and sequence on their genetic stability. Multimers of increasing size of the EcoRI linker CCGAATTCGG (E), the BamHI linker CCGGATCCGG (B) or mixtures of both (E, B) were inserted into the PvuII site of a previously constructed bacteriophage strain phi X174 J-F ins6. The largest inserts that could be maintained in the genome without significant loss of genetic stability were 2B, 4E, and 4(E, B), respectively. Polymers exceeding this size could be inserted but resulted in rapid and precise deletion from the phage genome, whereby nB was more unstable than nE, and nE was more unstable than n(E, B). Analysis of the resulting deletion mutants provided evidence for two different types of deletions. The more frequent deletion arose from either type palindrome and removed nucleotides in blocks of ten base-pairs (one linker unit), but only from the palindromic sequence, and always left at least an 18 base-pair long palindrome (one linker plus 8 neighboring base-pairs) behind. The less frequently occurring deletions arose only from nB type palindromes, removing the complete palindromic sequence plus adjacent nucleotides. At least the first type of deletion occurred in the absence of recA activity. Our results show a correlation between the sequence, as well as size, and the genetic stability of palindromes, i.e. sequences that could decrease the stability of a cruciform increased their genetic stability. This supports the theory that palindrome deletion occurs via extrusion of the palindrome into a cruciform or cruciform-like structure.

Bacteriophage phi X 174↗
PubMed1987

The effect of supercoil and temperature on the recognition of palindromic and non-palindromic regions in phi X174 replicative form DNA by S1 and Bal31.

The effect of supercoil and temperature on the topology of phi X174 replicative form (RF) DNA was studied using single-strand specific endonucleases S1 and Bal31 as probes for cruciform extrusion and other structural perturbations of the B-helix. Both enzymes were found to recognize specifically and reproducibly over 30 sites, most of which were cleaved by both enzymes independent of the superhelicity of the genome. A negative superhelical density exceeding 0.06 stabilized a transition in the DNA conformation that generated several new cleavage sites for Bal31. The underlying structures appeared to be only transiently stable and were lost from in vitro supercoiled DNA during brief incubation at 65 degrees C. They were generally absent from in vivo supercoiled RF DNA of equal superhelicity as a consequence of the extraction and storage procedure. Mapping of the cleavage sites suggested that they were preferentially located near the beginnings and ends of genes and that the structural basis for at least some of them was the extrusion of relatively small palindromes into the cruciform state. Insertion of a short synthetic palindromic sequence into the phi X174 genome generated a supercoil-dependent, temperature-sensitive secondary structure that was cleaved in the Bal31 but not the S1 reaction, further supporting the hypothesis that even small cruciforms with stem size of 7 or less base pairs may be transiently stable. Subjecting supercoiled RF DNA to the typical S1 reaction conditions induced a topological shift that diminished all but one of the supercoil-induced Bal31 recognition sites and promoted the formation of one major new site.

Bacteriophage phi X 174↗
PubMed1987

Immunotherapy with Hymenoptera venoms. Position paper of the Working Group on Immunotherapy of the European Academy of Allergy and Clinical Immunology.

Immunotherapy with Hymenoptera venoms is widely used throughout the world and is accepted as an effective treatment for most patients with Hymenoptera venom allergy. There are, however, still some unresolved problems with this form of treatment. At present there is no definite test which makes it possible to identify patients at risk - and thus candidates for immunotherapy - unequivocally. On the basis of prospective studies on the natural history of Hymenoptera allergy, venom immunotherapy is indicated in adults with severe systemic anaphylaxis. It is usually not necessary in patients with large local reactions only. Children with mild systemic reactions, e.g. urticaria, will need immunotherapy only in case of repeated reactions and/or a high risk of re-exposure. The selection of venoms for immunotherapy may lead to some confusion owing to common antigenic determinants shared by venoms of various Hymenoptera species. Many different regimens for immunotherapy have been proposed. At present, the three main are: rush, stepwise or clustered and classical. The maintenance dose of 100 micrograms usually protects from life-threatening reactions. However, in some patients 200 micrograms are necessary for complete protection. The usual interval between maintenance injections is 4 to 6 weeks. In many patients a strong increase of venom specific serum IgG-antibodies usually parallels clinical protection induced by venom immunotherapy, although many exceptions have been reported. Allergic side effects of venom immunotherapy are not rare, especially with honey bee venom and during the initial phase of dose increase. The question of the duration of venom immunotherapy is handled differently: although some authors recommend treatment for life, most suggest treating patients until skin tests and RAST become negative.

Adult↗
PubMed1986

Insertions of palindromic DNA sequences into the J-F intercistronic region of bacteriophage phi X174 interfere with normal phage growth.

The effect of hairpin (cruciform) size on the regulation of gene expression was investigated by cloning a series of palindromic sequences into the non-essential J-F intercistronic region of the bacteriophage phi X174 ins6 genome. Genetic stability of the insert sequence and its effect on the growth efficiency of the phage was used as an initial measure of the biological consequence of hairpin insertions. Multimers of increasing size of the BamHI linker sequence C-C-G-G-A-T-C-C-G-G were inserted into the PvuII site of the parental strain ins6. The largest hairpin that could be constructed and maintained in the phi X174 genome had a stem length of 22 base-pairs and a loop size of four nucleotides (linker tetramer). However, this structure proved to be disadvantageous to the phage and was rapidly deleted from its genome. Trimer inserts were more stable, but were eventually deleted also. Monomer and dimer inserts, though genetically stable, decreased the growth efficiency of the phage as judged by competitive growth experiments and measurements of burst size. The physical formation of these hairpins was shown by restriction digests of single-stranded DNA with BamHI and HpaII. We argue that these secondary structures form in vivo, at least in the single-stranded genome and the polycistronic mRNAs, and were responsible for the observed growth defects.

Bacteriophage phi X 174↗
PubMed1986

Combined active and passive immunotherapy in honeybee-sting allergy.

Five honeybee sting-allergic patients in whom bee venom immunotherapy (VIT) had previously been stopped because of repeated allergic side effects (SE) were resubmitted to bee VIT after pretreatment with beekeeper gammaglobulin. The tolerated bee venom dose was increased five to 800 times after this passive immunotherapy, and the maintenance dose of 100 micrograms of honeybee venom (BV) was reached in all patients after 6 to 15 days of a rush hyposensitization. The IgG response to BV was not suppressed by the beekeeper-gammaglobulin infusion. After 2 to 9 weeks of continued bee VIT, SE recurred in all patients. Therapy had to be stopped in one patient. The other four patients are currently on a maintenance dose of 60 to 80 micrograms. The possibility of a qualitative inferiority of the patients own IgG response was studied by crossed radioimmunoelectrophoresis and IgG subclass analysis. Lack of an IgG response to particular BV components to which IgE was present was demonstrated in three patients. The IgG subclass antibody pattern to BV at the time of recurrence of SE was similar to that found in beekeepers with a prominent IgG4 response. In contrast to beekeepers, however, there was no IgG2 response in patients.

Adult↗
PubMed1985

The biological significance of G-T/G-U mispairing in nucleic acid secondary structure.

We have computed the expected distribution of the potential for hairpin-like secondary structures with small loops (3-20 bases) and uninterrupted stems and compared that to the distribution observed in the complete genomes of seven DNA viruses from animals, plants and bacteria, as well as a bacterial plasmid. The formation of G-T mismatches in the stems of these structures was allowed. Furthermore we have analyzed the distribution of the potential for such structures along the genetic maps of these genomes, specifically around the start sites of known genes. Our data reveal that the potential for mismatch containing structures with stem length exceeding eight base pairs is over-represented and non-randomly distributed, but to a much lesser degree than that for perfect structures of equal size. Moreover, the potential for both types of structures is preferentially located near functional start codons. From this we deduce that in general G-T/G-U containing nucleic acid secondary structures are biologically relevant, though possibly less significant than perfect ones.

Bacteriophage phi X 174↗
PubMed1985

Eclipse kinetics as a probe of quaternary structure in bacteriophage phi X174.

The extracellular form of bacteriophage phi X174 consists of single-stranded DNA within an icosahedral capsid, which has short spikes at each of its vertices. Each spike is composed of gene G and H proteins, while the capsid itself consists of gene F protein. Since several molecules of gene H protein are injected into the cell along with the DNA, specific protein--protein and DNA--protein interactions must be broken when the genome exits and leaves an intact capsid structure at the receptor site. To demonstrate this we examined the eclipse (DNA ejection) reaction with two types of phi X174 mutants. The first contains missense mutations in a capsid or spike protein gene, and the second involves insertions or deletions in non-coding regions of the DNA. Using an improved procedure, the eclipse rate in vivo of the eclipse mutants Fcs70 has been redetermined over a larger temperature range than in previous studies. The three- to fivefold decrease in rate between 37 degrees C and 25 degrees C is due to an increase in both the enthalpy and entropy of activation when compared to the wild-type values of these kinetic parameters. This missence mutation also confers an increase in virus stability in 2 to 3 M-urea. In contrast to this, inserting 163 bases into the length of DNA packaged within the phi X174 capsid does not lead to a detectable change in eclipse rate over the same temperature range. yet this insertion into the J--F intercistronic region imparts a significant decrease in virus stability in urea. These results suggest that a specific set of non-covalent interactions is involved in phi X174 DNA ejection. This is supported by the small (50%), but significant, increase in eclipse rate that occurs when 27 bases are deleted from the J--F intercistronic region. The latter effect must be base-sequence-specific since no change in rate is observed when only seven of the 27 bases are deleted. Thus, the kinetics of the phi X174 eclipse reaction can be used as a sensitive probe of quaternary structure by correlating the change in reaction rate with alterations in amino acid and base sequences in the structural components of the virus.

Adsorption↗
PubMed1984

Avian retrovirus pp32 DNA binding protein. Preferential binding to the promoter region of long terminal repeat DNA.

The avian retrovirus pp32 protein possesses DNA endonuclease activity and unique DNA binding properties. An improved purification procedure was developed for pp32, resulting in a severalfold increase in the yield of this virion protein. By use of the nitrocellulose filter binding assay, the protein retains approximately 2-fold more supercoiled (form I) DNA molecules than equivalent linear duplex DNA molecules. Single-stranded DNA is only slightly preferred over double-stranded DNA for pp32 binding. The pp32 DNA binding sites on form I pBR322 DNA which contained an insert of avian retrovirus long terminal repeat (LTR) DNA were determined. A preformed protein-DNA complex was digested with one of several different multicut restriction enzymes and filtered through nitrocellulose filters. Fragments containing viral LTR DNA sequences and plasmid DNA containing promoter sequences for the ampicillin and tetracycline genes, sequences for the "left-end" inverted repeat of transposon 3, and sequences encompassing the carboxyl terminus of the beta-lactamase gene were preferentially retained on the filter by pp32. Partial mapping of pp32 DNA binding sites on LTR DNA was accomplished by generation of deletions in LTR DNA sequences. The pp32 protein preferentially bound viral DNA fragments which contain the viral promoter (TATTTAA) and the adjacent "R" repeat sequences. Computer analysis revealed that three of the four plasmid DNA fragments retained by pp32 contained LTR DNA promoter-like sequences (one mismatch only) which were part of statistically significant and thermodynamically stable hairpin structures.

Animals↗
PubMed1984

Studies of coexisting honeybee and vespid-venom sensitivity.

Honeybee and vespid venom-specific IgE were measured by RAST in randomly selected sera of 87 patients who had had anaphylactic reactions after insect stings. Overall there was a poor correlation between the titers of honeybee venom and yellow jacket or hornet venom-specific IgE. Sera from nine patients with high titers of both honeybee venom and yellow jacket venom-specific IgE were selected for RAST-inhibition studies, with these two venoms as coupling and inhibiting antigens. Three patterns of IgE-antibody specificity were detected. Four patients had unique antibody activity with no cross-reactivity between the yellow jacket and honeybee venom-specific IgE. This is probably the most common pattern in patients with dual sensitivity. Three patients reacted to a major allergen in yellow jacket venom cross-reacting with a minor allergen in honeybee venom. Their RAST-inhibition patterns demonstrated that the yellow jacket-venom RAST was inhibited by yellow jacket venom only and the honeybee-venom RAST was inhibited by both yellow jacket venom and honeybee venom. Two patients had the opposite pattern with honeybee-venom RAST inhibited by honeybee venom only and the yellow jacket RAST inhibited by both honeybee venom and yellow jacket venom. These latter patients reacted to a major allergen in honeybee venom that was cross-reacting with a minor allergen in yellow jacket venom. Studies with rabbit antisera raised to vespid and honeybee venoms demonstrated major antigens that were unique to each family that did not cross-react and several minor cross-reacting antigens.

Adolescent↗