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M Hildebrand

Publications and source records attributed to M Hildebrand.

At least 19 recordsLinked to original sources
PubMed2006

Characterization of hns genes from Erwinia amylovora.

The small basic histone-like protein H-NS is known for bacteria to attenuate virulence of several animal pathogens. An hns homologue from E. amylovora was identified by complementing an E. coli hns-mutant strain with a cosmid library from E. amylovora. A 1.6 kb EcoRI-fragment complemented the mucoid phenotype and repressed the ss-glucosidase activity of E. coli PD32. The open reading frame encoding an H-NS-like protein of 134 amino acid was later shown to be located on plasmid pEA29 (McGhee and Jones 2000). A chromosomal hns gene was amplified with PCR consensus primers and localized near galU of E. amylovora. E. amylovora mutants were created by insertion of a resistance cassette, and the intact gene was inserted into a high copy number plasmid for constitutive expression. Purified chromosomal H-NS protein preferentially bound to a DNA fragment from the lsc region and bending was predicted for an adjacent fragment with the rlsB-promoter. Levan production was significantly increased by hns mutations. Synthesis of the capsular exopolysaccharide amylovoran and of levan were reduced, when hns from the E. amylovora plasmid was overexpressed. A mutation in chromosomal hns of E. amylovora increased amylovoran synthesis, and both mutations retarded symptom formation on immature pears.

Base Sequence↗
PubMed2003

Instability of short-sequence DNA repeats of pear pathogenic Erwinia strains from Japan and Erwinia amylovora fruit tree and raspberry strains.

An array of short-sequence DNA repeats (SSRs) occurs in the plasmid pEA29 of the fire blight pathogen Erwinia amylovora. A large number of "fruit tree" strains, mainly from Central and Western Europe, were screened for their SSR numbers, and the analyses were extended to five raspberry strains from North America and six pear pathogenic Erwinia strains from Japan. The repeat ATTACAGA present in all E. amylovorastrains was found to be reiterated 3 to 15 times. The Japanese strains contained the major repeat sequence GGATTCTG, which was reiterated 16 to 24 times. ATTACAGG, which resembles the SSR of E. amylovora, was reiterated two or three times. In a novel approach, sequencing gels were used to visualize the rare occurrence of shorter arrays (down to three repeats) in E. amylovoraand the Japanese Erwinia strains. Changes in the repeat numbers in E. amylovora were observed repeatedly when the bacteria had been exposed to stress conditions. The repeat structures of homo- and heteroduplices of PCR-amplified repeats were also analyzed by cleavage of annealed molecules with the single-strand-specific endonuclease from bacteriophage T4. Not only heteroduplexes, but also homoduplexes showed non-matching regions in the SSRs, which could arise from transient formation of loops due to strand slippage during the assays.

Base Sequence↗
PubMed2001

Spatiotemporal self-organization in a surface reaction: from the atomic to the mesoscopic scale.

Scanning tunneling microscopy data revealed the atomic processes in propagating reaction fronts that occur in the catalytic oxidation of hydrogen on Pt(111). The fronts were also characterized on mesoscopic length scales with respect to their velocity and width. Simulations on the basis of a reaction-diffusion model reproduce the experimental findings qualitatively well. The quantitative comparison reveals the limitations of this traditional approach to modeling spatiotemporal pattern formation in nonlinear dynamics.

Journal Article↗
PubMed2001

Spatial symmetry breaking in the Belousov-Zhabotinsky reaction with light-induced remote communication.

Domains containing spiral waves form on a stationary background in a photosensitive Belousov-Zhabotinsky reaction with light-induced alternating nonlocal feedback. Complex behavior of colliding and splitting wave fragments is found with feedback radii comparable to the spiral wavelength. A linear stability analysis of the uniform stationary states in an Oregonator model reveals a spatial symmetry breaking instability. Numerical simulations show behavior in agreement with that found experimentally and also predict a variety of other new patterns.

Journal Article↗
PubMed2001

Molecular comparison of pathogenic bacteria from pear trees in Japan and the fire blight pathogen Erwinia amylovora.

Several strains of the genus Erwinia, which were isolated in Japan from pear trees with necrotic symptoms that resembled fire blight, and tentatively identified as Erwinia amylovora, were reinvestigated for their relationship to the fire blight pathogen. These isolates produced ooze on slices of immature pears and were mucoid on MM2Cu agar plates, but did not synthesize levan and did not give the expected PCR signals with several primer pairs specific for Erwinia amylovora. The isolates tested positive with PCR primers designed to detect the novel pear pathogen Erwinia pyrifoliae, which was isolated from Nashi pear trees in South Korea. The nucleotide sequence analysis of a DNA fragment preceding the gene cluster for exopolysaccharide synthesis revealed a closer relationship to Erwinia pyrifoliae than to Erwinia amylovora. Plasmid profiles, protein patterns and genomic DNA analysed by PFGE after XbaI and SpeI digestion were different than Erwinia amylovora. Experiments with strains of Erwinia amylovora isolated from raspberry (Rubus sp.), Erwinia mallotivora and Enterobacter pyrinus also did not reveal a relationship between these bacteria and the Japanese Erwinia strains. The latter are not identical to Erwinia pyrifoliae, but possess many similar features to this pathogen that causes Asian pear blight. It is concluded that pathogenic bacteria isolated in Japan from pear trees with symptoms resembling fire blight are possibly different from Erwinia amylovora.

DNA Primers↗
PubMed2000

Pharmacokinetics of oral iloprost in patients with Raynaud's phenomenon secondary to systemic sclerosis.

Iloprost is a chemically stable, pharmacologically highly potent prostacyclin-minietic. The therapeutic efficacy of the intravenous preparation was proven in patients with peripheral arterial occlusive disease or with Raynaud's phenomenon (RP). Recently, a sustained release oral preparation was developed for outpatient therapy. The purpose of the current study was to investigate whether the oral drug has a different pharmacokinetic profile in patients with RP secondary to systemic sclerosis (SSc) in comparison with healthy volunteers. Ten patients with RP secondary to SSc and 10 healthy volunteers (matched for age and sex) participated. Oral iloprost 50 microg was given twice daily for 8 days with dosing intervals of 5 h and plasma levels were taken over 10 h on Day 1 and 8. Plasma levels of iloprost were determined by a validated specific and sensitive radio-immunoassay. Compared with healthy volunteers, patients with SSc exhibited higher AUC values (by mean factors of 2. 1 and 2.0 on Day 1 and 8) and maximum plasma levels (by mean factors of 1.6 and 1.8 on Day 1 and 8). The increased systemic iloprost exposure was observed after both daily doses and on both monitored study days. Mean AUC values did not show accumulation over the 8 days. These findings are in agreement with a reduced total clearance of iloprost given by i.v. route in SSc patients compared to healthy volunteers, although no participant with severe renal impairment was included. A weak but significant correlation was found between individual creatinine clearance and AUC values. In conclusion, RP secondary to SSc is associated with an increased systemic iloprost exposure which is probably caused by changes of the metabolic clearance of iloprost. These effects cannot be explained by changes of renal function alone.

Administration, Oral↗
PubMed1998

Characterization of a silicon transporter gene family in Cylindrotheca fusiformis: sequences, expression analysis, and identification of homologs in other diatoms.

The transport of silicon is an integral part of the synthesis of the silicified cell wall of diatoms, yet knowledge of the number, features, and regulation of silicon transporters is lacking. We report the isolation and sequence determination of five silicon transporter (SIT) genes from Cylindrotheca fusiformis, and examine their expression patterns during cell wall synthesis. The encoded SIT amino acid sequences are highly conserved in their putative transmembrane domains. Nine conserved cysteines in this domain may account for the sensitivity of silicon uptake to sulfhydryl blocking agents. A less conserved C-terminal domain is predicted to form coiled-coil structures, suggesting that the SITs interact with other proteins. We show that SIT gene expression is induced just prior to, and during, cell wall synthesis. The genes are expressed at very different levels, and SIT1 is expressed in a different pattern from SIT 2-5. Hybridization experiments show that multiple SIT gene copies are present in all diatom species tested. From the data we infer that individual transporters play specific roles in silicon uptake, and propose that the cell regulates uptake by controlling the amount or location of each. The identification of all SIT genes in C. fusiformis will enhance our understanding of the mechanism and control of silicon transport in diatoms.

Amino Acid Sequence↗
PubMed1998

Tumor metastasis inhibition with the prostacyclin analogue cicaprost depends on discontinuous plasma peak levels.

Stable prostacyclin analogues exert a strong inhibitory effect on lymphogenous as well as haematogenous tumor metastasis in a series of tumor lines. The strong inhibition of metastasis was achieved by repeated once-daily i.g. applications. The mechanism of antimetastatic action is related to the expression of functional IP-receptors (PGI-receptors). As cellular assay systems indicated that the IP-receptor mediated signalling is down-regulated upon continuous exposure to prostacyclin or stable derivatives, it has been questioned whether a mode of drug application with constant plasma drug levels may potentially result in a decrease of the antimetastatic effect. We addressed this question using the stable prostacyclin analogue cicaprost in a disease model by comparing i.g. applications given once daily with a continuous administration of equivalent doses via drinking water. Very similar to our previous investigations in the 13762NF MTLn3 rat mammary carcinoma model, cicaprost administered by i.g. application strongly reduced lung and lymph node metastasis. In contrast, administration of equivalent doses via drinking water leading to lower but constant steady-state plasma levels failed to exert inhibitory effects. Plasma and urine levels of cicaprost were measured with a sensitive radioimmunoassay on the last treatment day. Pharmacokinetic evaluation demonstrated a similar bioavailability of cicaprost in both groups. This result first demonstrates a treatment failure of a prostacyclin derivative in a chronic disease model in association with a continuous drug administration leading to constant plasma levels. A desensitization of receptor signalling by constant plasma levels may be a possible mechanism for treatment failure.

Administration, Oral↗
PubMed1998

Patient-informant concordance on the Structured Clinical Interview for DSM-III-R personality disorders (SCID-II).

In the assessment of personality disorders, patients reports can be questioned for several reasons, such as lack of insight, shame, and acute psychiatric state. High concordance between patient-based and informant-based diagnoses would be an indication of the validity of patient reports (convergent validity). The present study examined the concordance between 42 psychotherapy outpatients and their informants (intimates) on the Structured Clinical Interview for DSM-III-R Personality Disorders (SCID-II). Similar to prior studies, low or only modest levels of agreement were found. In comparison with evaluations of the personality of the patient by the therapist, patient interviews seemed to be more valid than informant interviews. Furthermore, couples with high intensity and intimacy in the contact generally showed higher agreement than couples with low intensity and intimacy. In conclusion, the data slightly suggest that patient reports are more valid than informant reports. However, the lack of a golden standard forces us to find more evidence before concluding that patient reports on personality result in valid diagnoses.

Adult↗
PubMed1997

Pharmacokinetics and tolerability of oral iloprost in thromboangiitis obliterans patients.

OBJECTIVE: Iloprost is a potent PGI2 mimetic, which has been shown to be therapeutically effective in several vascular disorders. Due to its rapid clearance from the central compartment, iloprost is administered mainly by i.v. infusion, which limits its use to hospitalized patients. In order to improve pharmacotherapeutic use of this PGI2 mimetic, an oral extended-release (ER) dosage form has been developed, which should mimic plasma level profiles as observed after i.v. infusion and serve as a therapeutic equivalent. METHODS: This trial was performed to investigate the tolerability and pharmacokinetics of iloprost administered perorally, compared with i.v. infusion, in 12 patients suffering from thromboangiitis obliterans (TAO). A dose titration was carried out for 1 week with i.v. iloprost, followed by a p.o. titration and treatment phase of 3 weeks' duration. Pharmacokinetics was investigated at the individually tolerated dose levels; i.e., on days 5-7 (i.v. infusion at 2, 2.5 and 3 ng.kg-1.min-1), and twice during p.o. treatment after b.i.d. administration of 50, 100, 150, 200 or 300 micrograms. RESULTS: Individual tolerability of iloprost varied: 7 patients out of 12 tolerated the maximum i.v. dose of 3 ng.kg-1.min-1; six tolerated the maximum oral dose of 600 micrograms. No patients withdrew from the study due to adverse events. Flush and headache were the most common adverse events and seemed to be related to the study drug. After i.v. infusion of iloprost, dose-normalized (3 ng.kg-1.min-1), steady-state plasma levels were 260 pg.ml-1. Terminal half-life was 0.57 h. Total clearance ranged from 8 to 17 ml.min-1.kg-1. Peroral administration of the ER formulation resulted in dose-dependent Cmax and AUC values. AUC values of the first and second daily dose interval, i.e., 0-5 h and 5-11 h after first dosing, were almost identical. Absolute bioavailability was 24%, with the exception of two patients who tolerated only 50 micrograms b.i.d. and exhibited a bioavailability of approx. 60%. The AUC values observed in weeks 2 and 4 were identical, demonstrating low day-to-day variability of iloprost plasma level profiles in TAO patients. CONCLUSION: Based upon pharmacokinetic data, the ER formulation provides an equivalent to the i.v. infusion of iloprost and broadens the range of therapy to nonhospitalized patients. The availability of capsules with 50 and 100 micrograms iloprost enables individual dose titration and pharmacotherapy. Beneficial effects, as observed with i.v. iloprost in TAO patients, should therefore be achievable by peroral pharmacotherapy using the new ER formulation.

Administration, Oral↗
PubMed1997

Pharmacokinetics and pharmacodynamics of intra-graft iloprost in femorodistal bypass surgery.

Intra-graft injection of the prostacyclin analogue, iloprost, was performed at the end of femorodistal bypass procedures in 12 patients with severe peripheral arterial occlusive disease. Iloprost plasma levels were measured and compared with changes in haemodynamics. There was a high initial iloprost plasma level (mean 625 pg/ml) which dropped to a mean of 50 pg/ml after 15 min. This correlated with an immediate reduction in systolic blood pressure which had returned to pretreatment levels after 15 min. In contrast, the vascular resistance distal to the graft showed a reduction after 5 min which was maintained for at least 20 min after iloprost injection and the mean blood flow through the graft increased steadily throughout the same period of measurement. The study showed an effect of iloprost on blood pressure which correlated with plasma levels, but the time course of the changes in distal vascular resistance and graft blood flow demonstrated an effect more prolonged than the half-life of iloprost.

Aged↗
PubMed1996

Identification and characterization of a gene cluster involved in manganese oxidation by spores of the marine Bacillus sp. strain SG-1.

The marine Bacillus sp. strain SG-1 forms spores that oxidize manganese(II) as a result of the activities of uncharacterized components of its spore coat. Nucleotide sequence analysis of chromosomal loci previously identified through insertion mutagenesis as being involved in manganese oxidation identified seven possible genes (designated mnxA to mnxG) in what appears to be an operon. A potential recognition site for the sporulation, mother-cell-specific, RNA polymerase sigma factor, sigmaK, was located just upstream of the cluster, and correspondingly, measurement of beta-galactosidase activity from a Tn917-lacZ insertion in mnxD showed expression at mid-sporulation to late sporulation (approximately stage IV to V of sporulation). Spores of nonoxidizing mutants appeared unaffected with respect to their temperature and chemical resistance properties and germination characteristics. However, transmission electron microscopy revealed alterations in the outermost spore coat. This suggests that products of these genes may be involved in the deposition of the spore coat structure and/or are spore coat proteins themselves. Regions of the deduced protein product of mnxG showed amino acid sequence similarity to the family of multicopper oxidases, a diverse group of proteins that use multiple copper ions to oxidize a variety of substrates. Similar regions included those that are involved in binding of copper, and the addition of copper at a low concentration was found to enhance manganese oxidation by the spores. This suggests that the product of this gene may function like a copper oxidase and that it may be directly responsible for the oxidation of manganese by the spores.

Amino Acid Sequence↗
PubMed1995

Stable expression of human cytochrome P450 3A4 in V79 cells and its application for metabolic profiling of ergot derivatives.

Expression of human cytochrome (CYP) in heterologous cells is a means of specifically studying the role of these enzymes in drug metabolism. The complete cDNA encoding CYP3A4 (PCN1) was inserted into an expression vector containing the strong myeloproliferative sarcoma virus promoter in combination with the enhancer of the cytomegalovirus and stably expressed in V79 Chinese hamster cells. The presence of genomically integrated CYP3A4 cDNA cell clones was confirmed by polymerase chain reaction analysis. Transcription was detected by reverse transcribed polymerase chain reaction analysis. Functional expression could be demonstrated by conversion of testosterone to the specific 6beta-hydroxylated product. In recombinant V79 cells expressing CYP3A4 about 6% of the substrate was converted to 6beta-hydroxytestosterone. The metabolism of two dopaminergic ergot derivatives was investigated in live recombinant V79 cells. Both lisuride and terguride were monodeethylated.

Animals↗