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T Köhler

Publications and source records attributed to T Köhler.

At least 19 recordsLinked to original sources
PubMed2001

Tetragonal crystalline carbon nitrides: theoretical predictions.

New tetragonal phases of crystalline carbon nitride (CN) and their atomic structures have been identified using a self-consistent-charge density-functional tight-binding method. A tetragonal rocksalt structure provides theoretical support to recent experimental evidence for a stoichiometric CN phase with tetragonal symmetry. A body-centered tetragonal CN phase with 1:1 stoichiometry is predicted to be highly stable and of interesting atomic structure with complicated C-C and N-N dimerizations along the c axis. The cubic-to-tetragonal transitions are carefully examined to understand the underlying mechanism.

Journal Article↗
PubMed2001

Differential selection of multidrug efflux mutants by trovafloxacin and ciprofloxacin in an experimental model of Pseudomonas aeruginosa acute pneumonia in rats.

The ability of trovafloxacin and ciprofloxacin to select efflux mutants in vivo was studied in a model of acute Pseudomonas aeruginosa pneumonia in rats. Twelve hours after intratracheal inoculation of 10(6) CFU of P. aeruginosa strain PAO1 enmeshed in agar beads, two groups of 12 rats were treated by three intraperitoneal injections of each antibiotic given every 5 h. Dosing regimens were chosen to obtain a comparable area under the concentration-time curve from 0 to infinity/MIC ratio of 27.9 min for trovafloxacin (75 mg/kg of body weight) and of 32.6 min for ciprofloxacin (12.5 mg/kg). Twelve rats were left untreated and served as controls. Rats were sacrificed 12 h after the last injection (34 h after infection) for lung bacteriological studies. Selection of resistant bacteria was determined by plating lung homogenates on Trypticase soy agar plates containing antibiotic. In untreated animals, the frequency of resistant colonies was 10-fold higher than in agar beads. Compared to controls, both treatment regimens resulted in a 2-log reduction of lung bacterial load. The frequency of resistant colonies was 10-fold less with trovafloxacin than with ciprofloxacin at twice the MIC (7.4 x 10(-5) versus 8.4 x 10(-4), respectively) (P < 0.05) and at four times the MIC (6.2 x 10(-4) versus 5.0 x 10(-5), respectively) (P < 0.05). A multidrug resistance phenotype typical of efflux mutants was observed in all 41 randomly tested colonies obtained from treated and untreated rats. In agreement with in vitro results, trovafloxacin and ciprofloxacin preferentially selected MexCD-OprJ and MexEF-OprN overproducers, respectively. These results demonstrate the differential ability of trovafloxacin and ciprofloxacin to select efflux mutants in vivo and highlight the rapid emergence of those mutants, even without treatment.

Acute Disease↗
PubMed2001

Inhibition of quorum sensing by a Pseudomonas aeruginosa dksA homologue.

The Pseudomonas aeruginosa las (lasR-lasI) and rhl (rhlR-rhlI) quorum-sensing systems regulate the expression of several virulence factors, including elastase and rhamnolipid. P. aeruginosa strain PR1-E4 is a lasR deletion mutant that contains a second, undefined mutation which allows production of elastase and rhamnolipid despite a nonfunctional las system. We have previously shown that this strain accomplishes this by increasing the expression of the autoinducer synthase gene rhlI. In this report, we show that the elastolytic phenotype of mutant PR1-E4 can be complemented with a P. aeruginosa homologue of the Escherichia coli dnaK mutation suppressor gene dksA. When supplied in trans on a multicopy plasmid, this gene completely suppressed elastase production by mutant PR1-E4. Cloning and Northern blot analysis revealed that dksA was neither mutated nor less transcribed in mutant PR1-E4. When overexpressed, dksA also reduced rhamnolipid production by both mutant PR1-E4 and the wild type, PAO1. Using Northern blot analysis and lacZ reporter fusions, we show that dksA inhibits rhlI, rhlAB, and lasB transcription. Exogenous N-butyryl-L-homoserine lactone overcame the reduced expression of rhlI and restored rhlAB and lasB expression, as well as elastase production. Our results suggest that the overproduction of the P. aeruginosa DksA homologue inhibits quorum-sensing-dependent virulence factor production by downregulating the transcription of the autoinducer synthase gene rhlI.

Bacterial Proteins↗
PubMed2001

Different domains of the essential GTPase Cdc42p required for growth and development of Saccharomyces cerevisiae.

In budding yeast, the Rho-type GTPase Cdc42p is essential for cell division and regulates pseudohyphal development and invasive growth. Here, we isolated novel Cdc42p mutant proteins with single-amino-acid substitutions that are sufficient to uncouple functions of Cdc42p essential for cell division from regulatory functions required for pseudohyphal development and invasive growth. In haploid cells, Cdc42p is able to regulate invasive growth dependent on and independent of FLO11 gene expression. In diploid cells, Cdc42p regulates pseudohyphal development by controlling pseudohyphal cell (PH cell) morphogenesis and invasive growth. Several of the Cdc42p mutants isolated here block PH cell morphogenesis in response to nitrogen starvation without affecting morphology or polarity of yeast form cells in nutrient-rich conditions, indicating that these proteins are impaired for certain signaling functions. Interaction studies between development-specific Cdc42p mutants and known effector proteins indicate that in addition to the p21-activated (PAK)-like protein kinase Ste20p, the Cdc42p/Rac-interactive-binding domain containing Gic1p and Gic2p proteins and the PAK-like protein kinase Skm1p might be further effectors of Cdc42p that regulate pseudohyphal and invasive growth.

Actins↗
PubMed2000

Asymmetrically localized Bud8p and Bud9p proteins control yeast cell polarity and development.

Diploid strains of the budding yeast Saccharomyces cerevisiae change the pattern of cell division from bipolar to unipolar when switching growth from the unicellular yeast form (YF) to filamentous, pseudohyphal (PH) cells in response to nitrogen starvation. The functions of two transmembrane proteins, Bud8p and Bud9p, in regulating YF and PH cell polarity were investigated. Bud8p is highly concentrated at the distal pole of both YF and PH cells, where it directs initiation of cell division. Asymmetric localization of Bud8p is independent of the Rsr1p/Bud1p GTPase. rsr1/bud1 mutations are epistatic to bud8 mutations, placing Rsr1p/Bud1p downstream of Bud8p. In YF cells, Bud9p is also localized at the distal pole, yet deletion of BUD9 favours distal bud initiation. In PH cells, nutritional starvation for nitrogen efficiently prevents distal localization of Bud9p. Because Bud8p and Bud9p proteins associate in vivo, we propose Bud8p as a landmark for bud initiation at the distal cell pole, where Bud9p acts as inhibitor. In response to nitrogen starvation, asymmetric localization of Bud9p is averted, favouring Bud8p-mediated cell division at the distal pole.

Cell Polarity↗
PubMed2000

3D cone-beam CT reconstruction for circular trajectories.

3D reconstruction from 2D projections obtained along a single circular source trajectory is most commonly done using an algorithm due to Feldkamp, Davis and Kress. In this paper we propose an alternative approach based on a cone-beam to parallel-beam rebinning step, a corresponding rebinning step into a rectangular virtual detector plane and a filtered backprojection. This approach yields an improved image quality reflected by a decreased low-intensity drop which is well known for 3D reconstruction from projection data obtained along circular trajectories. At the same time the computational complexity is lower than in Feldkamp's original approach. Based on this idea, a hybrid 3D cone-beam reconstruction method is formulated that enlarges the reconstruction volume in its dimension along the rotation axis of the cone-beam CT system. This enlargement is achieved by applying different reconstruction conditions for each voxel. An optimal ratio between the reconstructible and irradiated volume of the scanned object is achieved.

Algorithms↗
PubMed2000

The n-PI-method for helical cone-beam CT.

A new class of acquisition schemes for helical cone-beam computed tomography (CB-CT) scanning is introduced, and their effect on the reconstruction methods is analyzed. These acquisition schemes are based on a new detector shape that is bounded by the helix. It will be shown that the data acquired with these schemes are compatible with exact reconstruction methods, and the adaptation of exact reconstruction algorithms to the new acquisition geometry is described. At the same time, the so-called PI-sufficiency condition is fulfilled. Moreover, a good fit to the acquisition requirements of the various medical applications of cone-beam CT is achieved. In contrast to other helical cone-beam acquisition and reconstruction methods, the n-PI-method introduced in this publication allows for variable pitches of the acquisition helix. This additional feature will introduce a higher flexibility into the acquisition protocols of future medical cone-beam scanners. An approximative n-PI-filtered backprojection (n-PI-FBP) reconstruction method is presented and verified. It yields convincing image quality.

Image Processing, Computer-Assisted↗
PubMed2000

Swarming of Pseudomonas aeruginosa is dependent on cell-to-cell signaling and requires flagella and pili.

We describe swarming in Pseudomonas aeruginosa as a third mode of surface translocation in addition to the previously described swimming and twitching motilities. Swarming in P. aeruginosa is induced on semisolid surfaces (0.5 to 0.7% agar) under conditions of nitrogen limitation and in response to certain amino acids. Glutamate, aspartate, histidine, or proline, when provided as the sole source of nitrogen, induced swarming, while arginine, asparagine, and glutamine, among other amino acids, did not sustain swarming. Cells from the edge of the swarm were about twice as long as cells from the swarm center. In both instances, bacteria possessing two polar flagella were observed by light and electron microscopy. While a fliC mutant of P. aeruginosa displayed slightly diminished swarming, a pilR and a pilA mutant, both deficient in type IV pili, were unable to swarm. Furthermore, cells with mutations in the las cell-to-cell signaling system showed diminished swarming behavior, while rhl mutants were completely unable to swarm. Evidence is presented for rhamnolipids being the actual surfactant involved in swarming motility, which explains the involvement of the cell-to-cell signaling circuitry of P. aeruginosa in this type of surface motility.

Aspartic Acid↗
PubMed2000

Improvement of nitric oxide-dependent vasodilatation by HMG-CoA reductase inhibitors through attenuation of endothelial superoxide anion formation.

Three 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (HCRIs), atorvastatin, pravastatin, and cerivastatin, inhibited phorbol ester-stimulated superoxide anion (O(2)(-)) formation in endothelium-intact segments of the rat aorta in a time- and concentration-dependent manner (maximum inhibition of 70% after 18 hours at 1 to 10 micromol/L). The HMG-CoA reductase product mevalonic acid (400 micromol/L) reversed the inhibitory effect of the HCRIs, which, conversely, was mimicked by inactivation of p21 Rac with Clostridium sordellii lethal toxin but not by inactivation of p21 Rho with Clostridium botulinum exoenzyme (C3). A mevalonate-sensitive inhibition of phorbol ester-stimulated O(2)(-) formation by atorvastatin was also observed in porcine cultured endothelial cells and in a murine macrophage cell line. In the rat aorta, no effect of the HCRIs on protein kinase C, NADPH oxidase, or superoxide dismutase (SOD) activity and expression was detected, whereas that of endothelial nitric oxide (NO) synthase was enhanced approximately 2-fold. Moreover, exposure of the segments to atorvastatin resulted in a significant improvement of endothelium-dependent NO-mediated relaxation, and this effect was abolished in the presence of SOD. Taken together, these findings suggest that in addition to augmenting endothelial NO synthesis, HCRIs inhibit endothelial O(2)(-) formation by preventing the isoprenylation of p21 Rac, which is critical for the assembly of NADPH oxidase after activation of protein kinase C. The resulting shift in the balance between NO and O(2)(-) in the endothelium improves endothelial function even in healthy blood vessels and therefore may provide a reasonable explanation for the beneficial effects of HCRIs in patients with coronary heart disease in addition to or as an alternative to the reduction in serum LDL cholesterol.

Animals↗
PubMed2000

Cytokine-driven differentiation of blasts from patients with acute myelogenous and lymphoblastic leukemia into dendritic cells.

We investigated the ability of both acute myelogenous leukemia (AML) and acute lymphoblastic leukemia (ALL) blasts to differentiate into dendritic cells (DC) in vitro. Cytokine-supplemented suspension cultures of leukemic blasts in 98 patients with AML and five patients with ALL (normal karyotype, n = 2; BCR/ABL, n = 3) were performed. Mononuclear cells out of peripheral blood or bone marrow containing between 60% and 90% leukemic blasts were cultured for eight days using different growth factor combinations. The highest yield of CD1a(+)/CD14(-) cells could be obtained with stem cell factor, transforming growth factor-beta, tumor necrosis factor-alpha, GM-CSF, and FLT-3-ligand. In the AML samples the median content of CD1a(+)/CD14(-) cells after eight days of culture was 3.5% (r = 0%-82%). In five informed patients CD1a(+)/CD14(-) cells were sorted by fluorescence-activated cell sorting or immunomagnetic separation. Cytogenetic and polymerase chain reaction analyses showed known primary chromosomal aberrations (monosomy 7 and inversion 16) in the sorted fractions, respectively. Dendritic cells (DC) could be generated out of leukemic blasts in 68% of AML patients. Leukemic DC showed no phagocytosis of latex beads, but stimulated allogeneic naive cord blood-derived T cells more efficiently than did uncultured blasts. In ALL patients the median percentage of CD1a(+)/CD14(-) cells was 1.2% (r = 0.7%-3.8%) after culture. The sorted CD1(+)/CD14(-) fractions were BCR/ABL-negative when analyzed with fluorescence in situ hybridization, indicating their nonleukemic origin. Leukemic DC can be generated out of leukemic progenitors in patients with AML. These cells might become relevant for autologous and allogeneic immunotherapy in selected patients. BCR/ABL-positive lymphoblasts could not be transformed into cells with an early dendritic phenotype with the cytokines used in our experiments.

Adolescent↗
PubMed2000

Effect of avilamycin, tylosin and ionophore anticoccidials on Clostridium perfringens enterotoxaemia in chickens.

In order to study the prophylactic and metaphylactic effect of antomicrobial growth promoters and ionophorous anticoccidials on the incidence of Cl. perfringens enterotoxaemia in chickens, experimental attempts were performed with 675 chickens in 27 trials. The birds were intraduodenally infected with Cl. perfringens type A (ATCC 3624). The following antimicrobial growth promoters and ionophore anticoccidials were used either on their own or in combination: avilamycin, narasin, monensin and tylosin. While infected and non-medicated trials showed an average incubation period of 1 week, clinical symptoms occurred 2-4 days later in infected and medicated birds. Avilamycin medicated birds had the longest incubation period. In the infected and non-medicated trials, a mortality rate of 16%-36% was noted within 3 weeks post infection. The avilamycin trials showed a mortality rate of 0-8% (0-2 birds died) and the narasin and monensin a mortality rate of 0-8%, respectively. In the combination groups (monensin + avilamycin or narasin + avilamycin), the mortality rate ranged from 0 to 4%. Tylosin showed a very good metaphylactic/therapeutic effect against Cl. perfringens enterotoxaemia. Following infection, medicated birds showed a significantly better bodyweight gain than the chickens, whose feeds had not been supplemented. From epidemiological point of view, the systematic prevention of coccidiosis is a key in the control of Cl. perfringens enterotoxaemia in chickens.

Animals↗
PubMed1999

Bacterial antibiotic efflux systems of medical importance.

Multidrug efflux systems endow on bacterial cells the ability to limit the access of antimicrobial agents to their targets. By actively pumping out antibiotic molecules, these systems prevent the intracellular accumulation necessary for antibiotics to exert their lethal activity. Drug efflux appears to be one of the most widespread antibiotic resistance mechanisms among microorganisms, since it has been demonstrated to occur in many Gram-positive and Gram-negative bacteria including medically important species like staphylococci, streptococci, enterobacteria and opportunistic pathogens like Pseudomonas aeruginosa. Efflux pumps can be specific for only one substrate or accommodate a more or less wide range of noxious products. Export of structurally unrelated compounds confers a multidrug-resistance phenotype on bacterial cells. Therapeutically critical levels of resistance can be achieved by overexpression of efflux systems, especially in those species such as P. aeruginosa which possess a low outer membrane permeability. It is suspected that the dual physiological function of active efflux systems is both the secretion of intracellular metabolites and the protection against a variety of harmful substances that the microorganism may encounter in its natural environment.

Bacteria↗
PubMed1999

Defining optimum conditions for the ex vivo expansion of human umbilical cord blood cells. Influences of progenitor enrichment, interference with feeder layers, early-acting cytokines and agitation of culture vessels.

Ex vivo expansion of human umbilical cord blood cells (HUCBC) is explored by several investigators to enhance the repopulating potential of HUCBC. We performed experiments using either Ficoll-separated or CD34+-selected HUCBC from the same donation in serum-free medium. CD34-purified HUCBC were cultured on either human umbilical vein endothelial cells (HUVEC) or irradiated bone marrow-derived stroma cells (BMSC) with addition of different cytokines. In addition, we tested the expansion of HUCBC in culture vessels with continuous rotation. CD34 enrichment led to a significant increase in the expansion factor of CD34+ cells compared with unmanipulated HUCBC. BMSC were more efficient in amplifying early progenitors than HUVEC. Optimum results were reached by a combination of SCF, FLT-3L at 300 ng/ml and IL-3 at 50 ng/ml. No significant improvement in the expansion of CD34+/38- primitive progenitors could be obtained with other combinations. Addition of megakaryocyte-derived growth and development factor to each growth factor cocktail improved the expansion results. Continuous rotation of culture vessels did not ameliorate the expansion rate of the analyzed subsets. Culture conditions separating stroma and HUCBC by a semipermeable membrane improved the expansion factors of CD34+, CD34+/38-, and CD34+/41+ cells and CFU-GM compared with contact cultures. These data might be useful when designing culture systems for clinical scale ex vivo expansion of HUCBC.

Antigens, CD34↗
PubMed1999

Absolute levels of MDR-1, MRP, and BCL-2 MRNA and tumor remission in acute leukemia.

Mononuclear cells prepared from peripheral blood or bone marrow of 119 AML and 28 ALL patients prior and following therapy were analyzed for absolute transcript levels of the chemoresistance genes mdr-1 and MRP, and the proto-oncogene bcl-2, by validated contamination-protected quantitative RT-PCR. In newly diagnosed AML mainly tumors of the granulocytic lineage (FAB M1-M2) expressed increased mdr-1 mRNA amounts. The MRP gene was expressed in all investigated samples without relation to a particular FAB class. High initial expression of both genes did not confer a poor prognosis even at high number of CD34+ cells. Data compared prior to and after therapy start (paired samples) revealed that AML patients who did not respond to therapy (NR) expressed increased levels of mdr-1 mRNA, as well as MRP and bcl-2 cDNA normalized to GAPDH reference transcripts, when compared to patients achieving complete remission (CR; p = 0.003, 0.008 and 0.0005, respectively). In ALL-NR the mdr-1 and bcl-2 genes were entirely more active after induction chemotherapy. Arbitrary cut-off values were established in order to delimit pathological from non-pathological gene expression. 59% of studied AML and 33% of ALL-NR exceeded the arbitrary values (mdr-1: > 2 amol/microgram RNA, MRP: > 10 zmol/amol GAPDH, bcl-2: > 5 zmol/amol GAPDH) for one and 11% of AML-NR for two parameters. Only 17% of the AML-CR and none of the ALL-CR group were above these limits. The results indicate that high individual activity of usually one, rarely two of the investigated genes might be associated with poor clinical outcome in treated acute leukemia.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1999

Drug resistance testing of acute myeloid leukemia in adults using the MTT assay.

OBJECTIVE: We want to evaluate the MTT in vitro assay for newly diagnosed AML in adults, in particular its prognostic significance. METHODS: MTT tests were performed according to Pieters et al., Blood 76: 2327, 1990. Up to 18 cytostatic drugs were tested in a wide range of concentrations over 4 days and compared with controls. If necessary, blasts were enriched > 80% by negative selection with dynabeads (Kaspers et al., Br. J. Cancer 70: 1047, 1994). Percent S-phase was measured at begin of assay and after 4 days. RESULTS: Technical success rate was 80-85% and the assay took about one day of work. IC50 values as a measure for drug resistance were highly reproducible with an interassay CV (range) of 28% (13-47) between days. Median cell viability of controls after 4 days was 80% (44-97);% S-phase at begin of assay was 3% (0.1-15) and after 4 days 9% (1-39). Leukemic blasts from 57 adult AML patients were prospectively tested in vitro at diagnosis. Large interindividual differences in drug resistance were observed (over 1000 fold on average). There was a trend for greater IC50 with higher cytogenetic risk and nonresponders to induction. Other clinical prognostic factors such as patient age, initial WBC, FAB subtype, prior MDS and % S-phase did not show a relationship with the in vitro data. There was a high correlation between the topoisomerase II related drugs regarding in vitro resistance. CONCLUSION: In vitro sensitivity tests like the MTT assay may provide a valuable tool for prediction of individual therapy outcome in combination with cytogenetics. However, longer follow-up is required.

Adolescent↗
PubMed1999

[Histological biomorphosis of human heart valve. II. Morphometric studies].

Previous polarization-microscopical measurements combined with silver-staining methods at histological sections of the bicuspid valve and the tricuspid valve in the region of fibrosa showed (similar to that of the corresponding tendinous cords) for middle-aged persons a distinct difference in the collagen composition in comparison between high pressure vs. low pressure system. The aim of the current study was the morphometric investigation of the collagen composition, of the relative content of total collagen (measured as relative number of total collagen fibers per measuring area), and of the cell-collagen relation in dependence of age and possibly on sex in the high and low pressure system, respectively. Tissue samples of atrioventricular valves of 67 probands of both sexes were available. The probands were healthy with regard to heart and circulation and distributed in 3 age groups (1st to 2nd decade, 3rd to 5th decade, and 6th to 9th decade). After corresponding preliminary treatment, the histological sections of tissue samples were evaluated by a suitable combination of polarization-microscopical, immuno-histochemical, and morphometrical methodology. The thickness of the heart valves was measured by microscopic image analysis. A rise of the number of fibers per measuring area (ma) for both sexes and both valves was observed with increasing age. The slopes of the linear regression curves and the mean values of the numbers of collagenous fibers/ma (for each age group) were different between both sexes. They depend upon the valve considered. Analogous to these results, the differentiated statements were possible to the number of fibers/ma and to the percentages of the fiber species by the distinction between the collagen types I and III in the fibrosa of the heart valves. Whereas the relative numbers of fibers/ma rose with increasing age, the number of cell (fibroblasts, fibrocytes) nuclei for both valves and sexes slightly decreased. For the latter, there was no difference between high and low pressure system in contrast to the collagenous fibers. The opposite age-dependent behavior of the collagenous fibers and of their producing cells during the biomorphosis of the heart valves could be explained with the collagen turn-over.

Adolescent↗
PubMed1999

Linear and nonlinear current density reconstructions.

Minimum norm algorithms for EEG source reconstruction are studied in view of their spatial resolution, regularization, and lead-field normalization properties, and their computational efforts. Two classes of minimum norm solutions are examined: linear least squares methods and nonlinear L1-norm approaches. Two special cases of linear algorithms, the well known Minimum Norm Least Squares and an implementation with Laplacian smoothness constraints, are compared to two nonlinear algorithms comprising sparse and standard L1-norm methods. In a signal-to-noise-ratio framework, two of the methods allow automatic determination of the optimum regularization parameter. Compensation methods for the different depth dependencies of all approaches by lead-field normalization are discussed. Simulations with tangentially and radially oriented test dipoles at two different noise levels are performed to reveal and compare the properties of all approaches. Finally, cortically constrained versions of the algorithms are applied to two epileptic spike data sets and compared to results of single equivalent dipole fits and spatiotemporal source models.

Algorithms↗