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Biomedical subjects

D Haas

Publications and source records attributed to D Haas.

At least 55 records · Page 3Linked to original sources
PubMed2000

A new syndrome of optic nerve colobomas and renal abnormalities associated with arthrogryposis multiplex.

Renal-coloboma syndrome is a developmental disorder involving optic nerve colobomas and renal hypoplasia/insufficiency, which exhibits autosomal dominant inheritance and a highly variable phenotype (OMIM:120330). Mutation in the PAX2 gene was found to result in the renal-coloboma phenotype. We report on an Arab family with autosomal dominant inheritance of a syndrome characterized by a variable combination of optic nerve colobomas, renal abnormalities, vesicoureteral reflux, lax joints and arthrogryposis multiplex. Apart from the arthrogryposis multiplex which has not been described in the renal-coloboma syndrome, the features of the syndrome in this family are very similar to the renal-coloboma syndrome. However sequencing of all 12 axons of PAX2 gene revealed no mutation in this family. The disorder in this family is likely to represent a new syndrome with features overlapping with the renal-coloboma syndrome.

Abnormalities, Multiple↗
PubMed2000

Transcriptional control of the hydrogen cyanide biosynthetic genes hcnABC by the anaerobic regulator ANR and the quorum-sensing regulators LasR and RhlR in Pseudomonas aeruginosa.

Virulence factors of Pseudomonas aeruginosa include hydrogen cyanide (HCN). This secondary metabolite is maximally produced at low oxygen tension and high cell densities during the transition from exponential to stationary growth phase. The hcnABC genes encoding HCN synthase were identified on a genomic fragment complementing an HCN-deficient mutant of P. aeruginosa PAO1. The hcnA promoter was found to be controlled by the FNR-like anaerobic regulator ANR and by the quorum-sensing regulators LasR and RhlR. Primer extension analysis revealed two transcription starts, T1 and T2, separated by 29 bp. Their function was confirmed by transcriptional lacZ fusions. The promoter sequence displayed an FNR/ANR box at -42.5 bp upstream of T2 and a lux box centered around -42.5 bp upstream of T1. Expression of the hcn genes was completely abolished when this lux box was deleted or inactivated by two point mutations in conserved nucleotides. The lux box was recognized by both LasR [activated by N-(oxododecanoyl)-homoserine lactone] and RhlR (activated by N-butanoyl-homoserine lactone), as shown by expression experiments performed in quorum-sensing-defective P. aeruginosa mutants and in the N-acyl-homoserine lactone-negative heterologous host P. fluorescens CHA0. A second, less conserved lux box lying 160 bp upstream of T1 seems to account for enhanced quorum-sensing-dependent expression. Without LasR and RhlR, ANR could not activate the hcn promoter. Together, these data indicate that expression of the hcn promoter from T1 can occur under quorum-sensing control alone. Enhanced expression from T2 appears to rely on a synergistic action between LasR, RhlR, and ANR.

Amino Acid Sequence↗
PubMed2000

Autoinduction of 2,4-diacetylphloroglucinol biosynthesis in the biocontrol agent Pseudomonas fluorescens CHA0 and repression by the bacterial metabolites salicylate and pyoluteorin.

The antimicrobial metabolite 2,4-diacetylphloroglucinol (2,4-DAPG) contributes to the capacity of Pseudomonas fluorescens strain CHA0 to control plant diseases caused by soilborne pathogens. A 2, 4-DAPG-negative Tn5 insertion mutant of strain CHA0 was isolated, and the nucleotide sequence of the 4-kb genomic DNA region adjacent to the Tn5 insertion site was determined. Four open reading frames were identified, two of which were homologous to phlA, the first gene of the 2,4-DAPG biosynthetic operon, and to the phlF gene encoding a pathway-specific transcriptional repressor. The Tn5 insertion was located in an open reading frame, tentatively named phlH, which is not related to known phl genes. In wild-type CHA0, 2, 4-DAPG production paralleled expression of a phlA'-'lacZ translational fusion, reaching a maximum in the late exponential growth phase. Thereafter, the compound appeared to be degraded to monoacetylphloroglucinol by the bacterium. 2,4-DAPG was identified as the active compound in extracts from culture supernatants of strain CHA0 specifically inducing phlA'-'lacZ expression about sixfold during exponential growth. Induction by exogenous 2,4-DAPG was most conspicuous in a phlA mutant, which was unable to produce 2, 4-DAPG. In a phlF mutant, 2,4-DAPG production was enhanced severalfold and phlA'-'lacZ was expressed at a level corresponding to that in the wild type with 2,4-DAPG added. The phlF mutant was insensitive to 2,4-DAPG addition. A transcriptional phlA-lacZ fusion was used to demonstrate that the repressor PhlF acts at the level of transcription. Expression of phlA'-'lacZ and 2,4-DAPG synthesis in strain CHA0 was strongly repressed by the bacterial extracellular metabolites salicylate and pyoluteorin as well as by fusaric acid, a toxin produced by the pythopathogenic fungus Fusarium. In the phlF mutant, these compounds did not affect phlA'-'lacZ expression and 2, 4-DAPG production. PhlF-mediated induction by 2,4-DAPG and repression by salicylate of phlA'-'lacZ expression was confirmed by using Escherichia coli as a heterologous host. In conclusion, our results show that autoinduction of 2,4-DAPG biosynthesis can be countered by certain bacterial (and fungal) metabolites. This mechanism, which depends on phlF function, may help P. fluorescens to produce homeostatically balanced amounts of extracellular metabolites.

Anti-Bacterial Agents↗
PubMed1999

Global GacA-steered control of cyanide and exoprotease production in Pseudomonas fluorescens involves specific ribosome binding sites.

The conserved two-component regulatory system GacS/GacA determines the expression of extracellular products and virulence factors in a variety of Gram-negative bacteria. In the biocontrol strain CHA0 of Pseudomonas fluorescens, the response regulator GacA is essential for the synthesis of extracellular protease (AprA) and secondary metabolites including hydrogen cyanide. GacA was found to exert its control on the hydrogen cyanide biosynthetic genes (hcnABC) and on the aprA gene indirectly via a posttranscriptional mechanism. Expression of a translational hcnA'-'lacZ fusion was GacA-dependent whereas a transcriptional hcnA-lacZ fusion was not. A distinct recognition site overlapping with the ribosome binding site appears to be primordial for GacA-steered regulation. GacA-dependence could be conferred to the Escherichia coli lacZ mRNA by a 3-bp substitution in the ribosome binding site. The gene coding for the global translational repressor RsmA of P. fluorescens was cloned. RsmA overexpression mimicked partial loss of GacA function and involved the same recognition site, suggesting that RsmA is a downstream regulatory element of the GacA control cascade. Mutational inactivation of the chromosomal rsmA gene partially suppressed a gacS defect. Thus, a central, GacA-dependent switch from primary to secondary metabolism may operate at the level of translation.

Amino Acid Sequence↗
PubMed1999

Morphological, immunophenotypical and electrophysiological properties of resting microglia in vitro.

Morphological, immunophenotypical and electrophysiological properties were investigated in isolated cultured murine microglia before and after exposure to astrocyte-conditioned medium (ACM). Following application of ACM, microglial cells underwent a dramatic shape transformation from an amoeboid appearance to a ramified morphology. In parallel to morphological changes, a downregulation of macrophage surface antigens was observed in microglia exposed to ACM. Staining intensities for major histocompatibility complex (MHC) class II molecules and for the adhesion molecules leukocyte function-associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) were significantly decreased in ramified microglia 5 days after exposure to ACM. In microglial cells treated daily with ACM over a period of 5 days, the smallest staining intensities for all surface antigens as well as the smallest ramification index as a measure for the highest degree of ramification were determined. In addition, upregulation of delayed rectifier K + currents was observed in microglia exposed to ACM for 1 day or treated daily with ACM for 5 days. In contrast, untreated amoeboid microglia or ramified microglia analysed 5 days after exposure to ACM did not express delayed rectifier K + currents. Analyses of the resting membrane potential and expression levels and properties of inward rectifier K + currents did not reveal any differences between untreated and ACM-treated microglia. It is suggested that electrophysiological properties of microglia do not strongly correlate with the morphology or the immunophenotype of microglial cells.

Animals↗
PubMed1999

Oxygen-sensing reporter strain of Pseudomonas fluorescens for monitoring the distribution of low-oxygen habitats in soil.

The root-colonizing bacterium Pseudomonas fluorescens CHA0 was used to construct an oxygen-responsive biosensor. An anaerobically inducible promoter of Pseudomonas aeruginosa, which depends on the FNR (fumarate and nitrate reductase regulation)-like transcriptional regulator ANR (anaerobic regulation of arginine deiminase and nitrate reductase pathways), was fused to the structural lacZ gene of Escherichia coli. By inserting the reporter fusion into the chromosomal attTn7 site of P. fluorescens CHA0 by using a mini-Tn7 transposon, the reporter strain, CHA900, was obtained. Grown in glutamate-yeast extract medium in an oxystat at defined oxygen levels, the biosensor CHA900 responded to a decrease in oxygen concentration from 210 x 10(2) Pa to 2 x 10(2) Pa of O(2) by a nearly 100-fold increase in beta-galactosidase activity. Half-maximal induction of the reporter occurred at about 5 x 10(2) Pa. This dose response closely resembles that found for E. coli promoters which are activated by the FNR protein. In a carbon-free buffer or in bulk soil, the biosensor CHA900 still responded to a decrease in oxygen concentration, although here induction was about 10 times lower and the low oxygen response was gradually lost within 3 days. Introduced into a barley-soil microcosm, the biosensor could report decreasing oxygen concentrations in the rhizosphere for a 6-day period. When the water content in the microcosm was raised from 60% to 85% of field capacity, expression of the reporter gene was elevated about twofold above a basal level after 2 days of incubation, suggesting that a water content of 85% caused mild anoxia. Increased compaction of the soil was shown to have a faster and more dramatic effect on the expression of the oxygen reporter than soil water content alone, indicating that factors other than the water-filled pore space influenced the oxygen status of the soil. These experiments illustrate the utility of the biosensor for detecting low oxygen concentrations in the rhizosphere and other soil habitats.

Bacterial Proteins↗
PubMed1999

Target joining of duplicated insertion sequence IS21 is assisted by IstB protein in vitro.

Tandemly repeated insertion sequence IS21, located on a suicide plasmid, promoted replicon fusion with bacteriophage lambda in vitro in the presence of ATP. This reaction was catalyzed in a cell extract containing the 45-kDa IstA protein (cointegrase) and the 30-kDa IstB helper protein of IS21 after both proteins had been overproduced in Escherichia coli. Without IstB, replicon fusion was inefficient and did not produce the 4-bp target duplications typical of IS21.

Artificial Gene Fusion↗
PubMed1999

The ArgR regulatory protein, a helper to the anaerobic regulator ANR during transcriptional activation of the arcD promoter in Pseudomonas aeruginosa.

Pseudomonas aeruginosa, when deprived of oxygen, generates ATP from arginine catabolism by enzymes of the arginine deiminase pathway, encoded by the arcDABC operon. Under conditions of low oxygen tension, the transcriptional activator ANR binds to a site centered 41.5 bp upstream of the arcD transcriptional start. ANR-mediated anaerobic induction was enhanced two- to threefold by extracellular arginine. This arginine effect depended, in trans, on the transcriptional regulator ArgR and, in cis, on an ArgR binding site centered at -73.5 bp in the arcD promoter. Binding of purified ArgR protein to this site was demonstrated by electrophoretic mobility shift assays and DNase I footprinting. This ArgR recognition site contained a sequence, 5'-TGACGC-3', which deviated in only 1 base from the common sequence motif 5'-TGTCGC-3' found in other ArgR binding sites of P. aeruginosa. Furthermore, an alignment of all known ArgR binding sites confirmed that they consist of two directly repeated half-sites. In the absence of ANR, arginine did not induce the arc operon, suggesting that ArgR alone does not activate the arcD promoter. According to a model proposed, ArgR makes physical contact with ANR and thereby facilitates initiation of arc transcription.

Adenosine Triphosphate↗
PubMed1999

Peripheral T cell lymphoma in a patient with common variable immunodeficiency disease: case report and literature review.

This report documents the occurrence of a peripheral T cell lymphoma arising in the bone marrow and liver of a patient with common variable immunodeficiency disease. The T cell origin of this lymphoma was demonstrated by immunohistochemical phenotyping and gene rearrangement studies and was not associated with EBV infection of the lymphoma cells. The frequency and characteristics of lymphomas complicating CVID are reviewed.

Adult↗
PubMed1999

Anterior segment anomalies of the eye associated with multiple skeletal abnormalities and early lethality: confirmation of an autosomal recessive syndrome.

We report two sibs from a complexly consanguineous family with joint contractures, skeletal abnormalities, anterior segment anomalies of the eye and early lethality. The features of these sibs are similar to the features of the sibs reported by Al-Gazali et al. (Clin Dysmorphol 3: 238-244, 1994). This report confirms the identity of this new autosomal recessive syndrome.

Abnormalities, Multiple↗
PubMed1999

Comparative investigations of airborne culturable microorganisms in selected waste treatment facilities and in neighbouring residential areas.

The evaluation of airborne microorganisms in waste treatment facilities is complicated by different measuring systems, a lack of measuring standards and large variations between individual counts. In the present study, different sectors of the waste management industry were compared by determining median values of airborne culturable microorganisms from numerous parallel counts over a prolonged time period. The samples were taken during the warm season using the six-stage Andersen volumetric sampler in a large composting plant and its immediate vicinity, in an agricultural composting plant, a waste disposal site, and a sorting facility for recyclable materials. Control samples were taken at a site not influenced by the waste management industry in an open and largely uninhabited area. The highest median values for culturable bacteria (37 degrees C) found were 1.1 x 10(5) CFU/m3, for moulds (25 degrees C) 1.4 x 10(5) CFU/m3, and for A. fumigatus (37 degrees C) 1.7 x 10(4) CFU/m3 in the sorting cabins of the sorting facility (p < 0.001). The highest median values for thermophilic bacteria (actinomycetes and bacillaceae, 50 degrees C) were 7.3 x 10(3) CFU/m3 in the large composting facility. In all other facilities as well as in the neighbouring residential areas of all facilities investigated, all median values were significantly lower and corresponded to the naturally occurring levels: approx. 10(2) CFU/m3 for bacteria, approx. 10(3) CFU/m3 for moulds and approx. 10(1) CFU/m3 for A. fumigatus and thermophilic bacteria. Only in the neighbouring residential area of the large composting plant, the median values for thermophilic bacteria were approx. 10(2) CFU/m3, but an additional impact from farms cannot be excluded in this case. These results show permanent increased loads of the investigated microorganisms inside large composting facilities and especially in the sorting cabins for recyclable materials. The increasing number of reports on potential health hazards in these areas require adequate measures on the part of occupational medicine in order to limit the health risk to a minimum. The most important task is the automatization of the sorting process for recyclable materials.

Air Microbiology↗
PubMed1999

[Waste management--health: a longitudinal study].

Altogether 256 (148 men and 108 women) employees from manual sorting facilities and 120 (98 men and 22 women) employees from compost facilities were examined The main focus of the investigations was in the assessment of the lung-function and the immune-system. A part of employees (87 in sorting and 34 in compost facilities) was observed over a period of 3 years. The lung-function was in the area of the expected values, decreased however from year to year. The concentration of total-IgE increased over the period of 3 years. The observation is from great interest that the difference of the concentration of the total-IgE fluctuated within the 3 years at the employees of manual sorting facilities from +200% to -100% while it amounted to 0% at employees of the compost facilities.

Adult↗
PubMed1999

[Comparative studies of airborne, cultivatable microorganisms at selected sites of waste managements, domestic animal husbandry and in the surrounding residential area].

During the last years, sampling of airborne microorganisms (including mesophilic bacteria, moulds, thermophilic bacteria and A. fumigatus) in waste disposal, recycling industry and control sampling locations in the urban and rural districts of Graz was conducted using Andersen 6-stage viable cascade impactors. In the present study additional sampling in livestock buildings has been conducted. Except mesophilic bacteria the emission in the area of waste disposal and recycling industry was significantly higher than in livestock buildings. In the surroundings of the livestock buildings the number of microorganisms was not increased, while the normal background level in the surroundings of the waste proceeding plants was reached from a distance of 150 to 300 meters.

Air Microbiology↗
PubMed1998

Cointegrase, a naturally occurring, truncated form of IS21 transposase, catalyzes replicon fusion rather than simple insertion of IS21.

The bacterial insertion sequence IS21 contains two genes, istA and istB, which are organized as an operon. IS21 spontaneously forms tandem repeats designated (IS21)2. Plasmids carrying (IS21)2 react efficiently with other replicons, producing cointegrates via a cut-and-paste mechanism. Here we show that transposition of a single IS21 element (simple insertion) and cointegrate formation involving (IS21)2 result from two distinct non-replicative pathways, which are essentially due to two differentiated IstA proteins, transposase and cointegrase. In Escherichia coli, transposase was characterized as the full-length, 46 kDa product of the istA gene, whereas the 45 kDa cointegrase was expressed, in-frame, from a natural internal translation start of istA. The istB gene, which could be experimentally disconnected from istA, provided a helper protein that strongly stimulated the transposase and cointegrase-driven reactions. Site-directed mutagenesis was used to express either cointegrase or transposase from the istA gene. Cointegrase promoted replicon fusion at high frequencies by acting on IS21 ends which were linked by 2, 3, or 4 bp junction sequences in (IS21)2. By contrast, cointegrase poorly catalyzed simple insertion of IS21 elements. Transposase had intermediate, uniform activity in both pathways. The ability of transposase to synapse two widely spaced IS21 ends may reside in the eight N-terminal amino acid residues which are absent from cointegrase. Given the 2 or 3 bp spacing in naturally occurring IS21 tandems and the specialization of cointegrase, the fulminant spread of IS21 via cointegration can now be understood.

Base Sequence↗
PubMed1998

Activated microglial cells migrate towards sites of excitotoxic neuronal injury inside organotypic hippocampal slice cultures.

The aim of this study was to analyse microglial reactions to excitotoxic N-methyl-D-aspartic acid (NMDA)-induced degeneration of rat dentate and hippocampal neurons in vitro. We used a migration model combining the techniques of microglial single cell culture and organotypic hippocampal slice culture (OHSC). Site-specific oxidative damage in OHSCs was induced by pretreatment with 50 microM NMDA. Neuronal injury determined by propidium iodide (PI) uptake included the hippocampal cell layers of the dentate gyrus (DG) and the cornu ammonis (CA). Fluorescence-prelabelled microglial cells with ameboid morphology were transferred onto the OHSC and migrated predominantly to the prelesioned cell layers of DG and CA when compared with unlesioned areas of the OHSC. In NMDA pretreated slices, microglial cells clustered around degenerating granule cells in the DG and pyramidal cells in the CA. This effect was significantly inhibited in unlesioned slice cultures and in NMDA-exposed cultures that were pretreated with the NMDA-antagonist MK-801. Our observations suggest that microglia -- attracted by the presence of stimuli provided by NMDA-induced neuronal death -- migrate specifically towards these lesioned neurons.

Animals↗
PubMed1998

Astrocytic factors deactivate antigen presenting cells that invade the central nervous system.

We hypothesized that CNS tissue has the potential to deactivate invading monocytes/macrophages in order to maintain the immune privilege of the brain, and furthermore, that astrocytes are the cells that initiate monocyte/macrophage deactivation. To test this hypothesis, fluorescent prelabeled rat spleen macrophages with typical amoeboid morphology were transferred into organotypic hippocampal slice cultures (OHSCs), where they gradually developed a ramified morphology similar to the appearance of resting microglial cells. This morphological transformation also occurred if macrophages or monocytes were co-cultured with mixed glial cultures or with astrocytoma cells, and ramification was accompanied by reduced expression of adhesion molecules leukocyte function antigen (LFA)-1, intercellular adhesion molecule (ICAM)-1, and major histocompatibility complex (MHC)-class-II molecules. Moreover, treatment of macrophages with astrocyte culture supernatant effectively down-regulated the LPS-induced expression of adhesion- and MHC-class-II-molecules. Astrocyte supernatant-induced inhibition of adhesion and MHC-class-II-molecule expression was mimicked by transforming growth factor (TGF)-beta1, furthermore, this inhibitory effect was diminished by simultaneous treatment with neutralizing anti-TGF-beta-antibodies. In conclusion, our results suggest that astrocyte-derived, soluble factors that are present in the CNS microenvironment deactivate invading macrophages, thus contributing to the maintenance of CNS immune-privilege following impairment of blood-brain-barrier (BBB) integrity.

Animals↗