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P Müller

Publications and source records attributed to P Müller.

At least 73 records · Page 4Linked to original sources
PubMed2001

Specific detection of Bradyrhizobium and Rhizobium strains colonizing rice (Oryza sativa) roots by 16S-23S ribosomal DNA intergenic spacer-targeted PCR.

In addition to forming symbiotic nodules on legumes, rhizobial strains are members of soil or rhizosphere communities or occur as endophytes, e.g., in rice. Two rhizobial strains which have been isolated from root nodules of the aquatic legumes Aeschynomene fluminensis (IRBG271) and Sesbania aculeata (IRBG74) were previously found to promote rice growth. In addition to analyzing their phylogenetic positions, we assessed the suitability of the 16S-23S ribosomal DNA (rDNA) intergenic spacer (IGS) sequences for the differentiation of closely related rhizobial taxa and for the development of PCR protocols allowing the specific detection of strains in the environment. 16S rDNA sequence analysis (sequence identity, 99%) and phylogenetic analysis of IGS sequences showed that strain IRBG271 was related to but distinct from Bradyrhizobium elkanii. Rhizobium sp. (Sesbania) strain IRBG74 was located in the Rhizobium-Agrobacterium cluster as a novel lineage according to phylogenetic 16S rDNA analysis (96.8 to 98.9% sequence identity with Agrobacterium tumefaciens; emended name, Rhizobium radiobacter). Strain IRBG74 harbored four copies of rRNA operons whose IGS sequences varied only slightly (2 to 9 nucleotides). The IGS sequence analyses allowed intraspecies differentiation, especially in the genus Bradyrhizobium, as illustrated here for strains of Bradyrhizobium japonicum, B. elkanii, Bradyrhizobium liaoningense, and Bradyrhizobium sp. (Chamaecytisus) strain BTA-1. It also clearly differentiated fast-growing rhizobial species and strains, albeit with lower statistical significance. Moreover, the high sequence variability allowed the development of highly specific IGS-targeted nested-PCR assays. Strains IRBG74 and IRBG271 were specifically detected in complex DNA mixtures of numerous related bacteria and in the DNA of roots of gnotobiotically cultured or even of soil-grown rice plants after inoculation. Thus, IGS sequence analysis is an attractive technique for both microbial ecology and systematics.

Bradyrhizobium↗
PubMed2001

Is the Zona pellucida thickness of metaphase-II oocytes in an IVF/ICSI program influenced by the patient's age?

Zona pellucida (Z.p.) thickness is believed to be considerably influenced by hormonal stimulation. Assessments of the influence of the patient's age on Z.p. thickness have been at variance. In our study we wanted to investigate, among several other parameters, the age of the patient as a possible factor influencing Z.p. thickness. Prior to intracytoplasmatic sperm injection (ICSI) we determined the mean thickness of the Z.p. of 843 metaphase-II oocytes from 100 patients, which had been obtained after ovarian stimulation in an in vitro fertilization (IVF)/ICSI program. The mean thickness of the Z.p. was significantly positively correlated with the age of the patient (p = 0.000, r = 0.129). When the intraindividual mean values of Z.p. thickness (n = 100) were used, a positive correlation with age (p = 0.174, r = 0.137) remained. However, this correlation was no longer significant. It is concluded that, beside other factors, a thicker Z.p. could be responsible for the possibly lower fertilization rate of older patients in a conventional IVF program.

Adult↗
PubMed2001

A shot in the dark: the silent quest of a free-flying phonotactic fly.

To reproduce, females of the parasitoid fly Ormia ochracea detect and localise a calling male cricket upon which they deposit their endoparasitic larvae. Calling male crickets are therefore subject to both sexual and natural selection by simultaneously attracting mates and phonotactic parasitoids. The possible strategy of song interruption employed by the cricket host to reduce his attractiveness to acoustic parasitoids was tested in the laboratory by examining the fly's phonotactic quest in response to synthetic cricket songs. Phonotactic flight trajectories were recorded in three dimensions with a stereo infrared video tracking system while the sound stimulus was controlled on-line as a function of the fly's position in space. Within a single flight, three distinct phases could be observed: a take-off phase, a cruising phase, during which course and altitude were rather constant, and a landing phase characterised by a spiralling descent towards the sound source. The flies showed remarkable phonotactic accuracy in darkness; they landed at a mean distance of 8.2 cm from the centre of the loudspeaker after a flight distance of approximately 4 m. The present data illustrate the fly's surprising ability to gauge the direction and distance of a sound source in three dimensions and, subsequently, to find it in darkness and silence.

Acoustic Stimulation↗
PubMed2001

Interleukin 3 improves the ex vivo expansion of primitive human cord blood progenitor cells and maintains the engraftment potential of scid repopulating cells.

In umbilical cord blood (UCB) transplantation, the number of nucleated cells per kilogram is a major predictive and critical factor of hematopoietic recovery. Thus, ex vivo expansion of hematopoietic UCB progenitors could potentially accelerate engraftment. Whereas Flt-3 ligand (FL), stem cell factor (SCF), and thrombopoietin (TPO) are considered indispensable, the role of interleukin 3 (IL-3) is still controversial: it has been reported either to support or abrogate the reconstituting ability of stem cells. By adding IL-3 we aimed to enhance the amplification of early and committed progenitor cells without impairing the long-term engraftment of stem cells. Demonstrating a positive impact of IL-3 on the proliferation of all progenitor subsets, the amplification of CD34+ UCB cells was increased 20.9-fold +/- 5.4 (mean +/- standard error) in serum-free culture with FL, SCF, TPO, and IL-3 as opposed to 9.3-fold +/- 3.2 without IL-3 after 7 days. If IL-3 was included, primitive long-term culture-initiating cells and committed colony-forming cells were expanded 16.3-fold +/- 5.5 and 18.1-fold +/- 2.4, respectively, compared to 12.6-fold +/- 5.6 and 9.1-fold +/- 2.0 without IL-3. Analysis of cultured CD34+ UCB cells in sublethally irradiated nonobese diabetic/severe combined immunodeficient mice confirmed that cultured cells had preserved their repopulating potential. After 6 weeks, all mice showed multilineage engraftment with their bone marrow containing an average of 45% human CD45+ cells of the unmanipulated sample, 43% of cells after culture in the presence of IL-3, and 27% of cells after culture without IL-3. In combination with early acting cytokines, IL-3 therefore improves the ex vivo expansion of UCB stem and progenitor cells without impairing their engraftment potential.

Animals↗
PubMed2001

Disseminated cytokeratin positive tumor cells in the bone marrow of patients with prostate cancer: detection and prognostic value.

PURPOSE: Previous investigations have shown that cytokeratin 18 positive bone marrow cells in localized and lymphatically spread prostate cancer correlates with neither established prognostic factors nor with the biochemical and clinical course after radical prostatectomy. Since the well-known down-regulation of cytokeratin 18 in tumor cells may lead to false-negative results, we asked whether staining with a pan-cytokeratin antibody recognizing a common epitope of cytokeratins 8, 18 and 19 would result in different data. MATERIALS AND METHODS: Preoperative bone marrow aspirates of 82 patients with localized (N0) and lymphatically spread (N1) prostate cancer were examined using the monoclonal antibody cytokeratin 2 and the pan-cytokeratin antibody A 45-B/B3, called A 45. RESULTS: In contrast to findings with the cytokeratin 18 antibody, those with the pan-cytokeratin antibody correlated with the biochemical course. At a median followup of 1,477 days (4 years) patients with pan-cytokeratin positive cells in the preoperative bone marrow aspirate had biochemical progression significantly earlier than those with pan-cytokeratin negative results (mean time to prostate specific antigen relapse 886 versus 1,409 days, p < or =0.004). Compared with other parameters, such as prostate specific antigen, pathological stage and Gleason score, preoperative pan-cytokeratin findings proved to be an independent prognostic factor. CONCLUSIONS: Cytokeratin positive cells in the bone marrow also have prognostic relevance in prostate cancer. The comprehensive analysis of these cells, studies of the individual course of these findings and sufficiently long followup allow us to discuss whether and under what conditions metastasis may develop from 1 or several cytokeratin positive cells.

Adult↗
PubMed2000

Critical dynamics of gelation.

Shear relaxation and dynamic density fluctuations are studied within a Rouse model, generalized to include the effects of permanent random crosslinks. We derive an exact correspondence between the static shear viscosity and the resistance of a random resistor network. This relation allows us to compute the static shear viscosity exactly for uncorrelated crosslinks. For more general percolation models, which are amenable to a scaling description, it yields the scaling relation k=straight phi-beta for the critical exponent of the shear viscosity. Here beta is the thermal exponent for the gel fraction, and straight phi is the crossover exponent of the resistor network. The results on the shear viscosity are also used in deriving upper and lower bounds on the incoherent scattering function in the long-time limit, thereby corroborating previous results.

Journal Article↗
PubMed2000

Systematic, immediate in-hospital initiation of lipid-lowering drugs during acute coronary events improves lipid control.

Background: Patients who have had a coronary heart attack often go completely untreated for hypercholesterolemia. We investigated whether immediate initiation of lipid-lowering drugs during hospitalization for acute coronary events increases the proportion of correctly treated patients compared to referred treatment as recommended by current guidelines. Methods: This prospective, multicenter study randomized 57 hypercholesterolemic patients hospitalized for acute coronary events to immediate in-hospital initiation or to referred initiation of lipid-lowering drugs by primary care physicians 3 months after unsuccessful nutritional intervention. Results: After 6 months, 53 patients were available for follow-up. More patients in the immediate initiation group (26/30 patients, 87%) were treated with lipid-lowering drugs than in the referred initiation-group (13/23 patients, 57%, P=0.03). Twenty-seven patients (87%) in the immediate initiation group versus 17 patients (65%) in the referred initiation group had a 10% or greater decrease in total cholesterol or a 15% or greater decrease in LDL-cholesterol (P=0.18). Although statistically not significant, there was a trend to improved lipid values in the immediate initiation group compared to the referred initiation group (TC, -21.1 vs. -13.8% (P=0.08); LDL-C, -28.2 vs. -18.9% (P=0.13); HDL-C, +10.8 vs. +5% (P=0.44); TC/HDL-C ratio, -24.7 vs. -15.1% (P=0.22)), and the LDL-C/HDL-C ratio was -34.1 vs. -19.1% (P=0.04, P=NS after Bonferroni correction). Conclusion: The immediate initiation of lipid-lowering drugs in hypercholesterolemic patients hospitalized for acute coronary events increases the rate of correctly treated patients and has the potential to improve lipid control.

Journal Article↗
PubMed2000

Conservation of the sequence and temporal expression of let-7 heterochronic regulatory RNA.

Two small RNAs regulate the timing of Caenorhabditis elegans development. Transition from the first to the second larval stage fates requires the 22-nucleotide lin-4 RNA, and transition from late larval to adult cell fates requires the 21-nucleotide let-7 RNA. The lin-4 and let-7 RNA genes are not homologous to each other, but are each complementary to sequences in the 3' untranslated regions of a set of protein-coding target genes that are normally negatively regulated by the RNAs. Here we have detected let-7 RNAs of approximately 21 nucleotides in samples from a wide range of animal species, including vertebrate, ascidian, hemichordate, mollusc, annelid and arthropod, but not in RNAs from several cnidarian and poriferan species, Saccharomyces cerevisiae, Escherichia coli or Arabidopsis. We did not detect lin-4 RNA in these species. We found that let-7 temporal regulation is also conserved: let-7 RNA expression is first detected at late larval stages in C. elegans and Drosophila, at 48 hours after fertilization in zebrafish, and in adult stages of annelids and molluscs. The let-7 regulatory RNA may control late temporal transitions during development across animal phylogeny.

Adult↗
PubMed2000

Light-induced reorganization of phospholipids in rod disc membranes.

The transbilayer redistribution of spin-labeled phospholipid analogues (SL-PL) with choline, serine, and ethanolamine head groups (PC, PS, and PE, respectively) was studied on intact disc vesicles of bovine rod outer segment membranes in the dark and after illumination. Redistribution was measured by the extraction of spin-labeled lipid analogues from the outer leaflet of membrane using the bovine serum albumin back-exchange assay. In the dark, PS was distributed asymmetrically, favoring the outer leaflet, whereas PC and PE showed small if any asymmetry. Green illumination for 1 min caused lipid head group-specific reorganization of SL-PL. Extraction of SL-PS by bovine serum albumin showed a fast transient (<10 min) enhancement, which was further augmented by a peptide stabilizing the active metarhodopsin II conformation. The data suggest a direct release of 1 molecule of bound PS per rhodopsin into the outer leaflet and subsequent redistribution between the two leaflets. SL-PE and SL-PC showed more complex kinetics, in both cases consistent with a prolonged period of reduced extraction (2 phospholipids per rhodopsin in each case). The different phases of SL-PL reorganization after illumination may be related to the formation and decay of the active rhodopsin species and to their subsequent regeneration process.

Animals↗
PubMed2000

Sparteine-mediated asymmetric nucleophilic substitution at prochiral, sp(3)-hybridized carbon.

[reaction: see text] ++-Phenyl-1,3-dioxolanes (1) react with organolithium reagents (2), associated with (-)-sparteine, in the presence of BF(3).OEt(2) to afford chiral monosubstitution products 3. Enantioselectivity is highest if both 1 and 2 carry alkyl substituents in the ortho position. However, the enantioselectivity decreases in the case of very bulky substituents such as tert-butyl or phenyl.

Journal Article↗
PubMed2000

Tracking of flying insects using pan-tilt cameras.

Potent and affordable video and computer systems for automatic data acquisition are becoming increasingly important in behavioural neuroscience. It has remained challenging, however, to acquire data from small and fast-moving animals, such as insects in flight, due to the limited spatial and temporal resolution of the systems currently available. Our research on free-flying insects motivated the development of new methods in the context of two different experimental settings. First, the position and precise body axis direction of honey bees approaching a food source were automatically measured. Second, the flight trajectories of a phonotactic parasitoid fly homing in on its cricket host were recorded in 3D. We used pan-tilt cameras, i.e. cameras with moveable optics, to follow the animal's path with a close up image. Novel methods were developed for image acquisition and position measurement using pan-tilt cameras, as well as calibration and data evaluation in 3D world coordinates. The innovations of this system comprise: (1) Acquisition of images in high spatial detail over large observation areas. (2) Image acquisition at a field rate of 50 Hz PAL. (3) Free positioning of the cameras for 3D acquisition. (4) Computation of the flight path in 3D world coordinates. We illustrate the capabilities of the system with data obtained from a calibration object as well as from the behaviour of unrestricted, free-flying flies and bees. Potential applications in behavioural neuroscience and the psychophysics of sensory perception are briefly discussed.

Animals↗
PubMed2000

Transmembrane movement of diether phospholipids in human erythrocytes and human fibroblasts.

We have synthesized spin-labeled (SL) and fluorescently labeled diacyl, 1-alkyl-2-acyl-, and di-alkyl glycerophospholipids. The sn-2 chain was a short chain with either a nitroxide group or a 7-nitro-2, 1,3-benzoxadiazol-4-yl (NBD). After incorporation in the exoplasmic leaflet of human erythrocytes, we found that SL-phosphatidylcholine (PC) redistributed very slowly across the plasma membrane, less than 20% reaching the cytoplasmic leaflet in 3 h at 37 degrees C. In contrast, SL-phosphatidylserine (PS) accumulated on the cytoplasmic leaflet with the same plateau corresponding to 90% of the probes inside. The characteristic times for inward redistribution were different for the three PS analogues: at 37 degrees C, the t(1/2) for the diacyl, alkyl-acyl, and dialkyl compounds were 2.3, 3.5, and 41 min, respectively. ATP depletion or incubation with N-ethylmaleimide inhibited the rapid translocation of the PS derivatives. The diether PS bearing an NBD group translocated very slowly in human erythrocytes and no acceleration by ATP could be measured. On the other hand, in human fibroblasts, the diether NBD-PS and SL-PS were both transported from the exoplasmic to the cytoplasmic monolayer of the plasma membrane as it is the case for the transport of the respective diester PS analogues. These results prove that the ether bonds do not prevent completely PS binding and translocation by the aminophospholipid translocase despite a probable hindrance due to the ether linkage on the sn-2 chain. Because of the high stability of the ether linkage, SL and NBD diether analogues should be useful to investigate lipid traffic in cultured cells.

Biological Transport↗
PubMed2000

Cytokeratin-positive cells in the bone marrow and survival of patients with stage I, II, or III breast cancer.

BACKGROUND: Cytokeratins are specific markers of epithelial cancer cells in bone marrow. We assessed the influence of cytokeratin-positive micrometastases in the bone marrow on the prognosis of women with breast cancer. METHODS: We obtained bone marrow aspirates from both upper iliac crests of 552 patients with stage I, II, or III breast cancer who underwent complete resection of the tumor and 191 patients with nonmalignant disease. The specimens were stained with the monoclonal antibody A45-B/B3, which binds to an antigen on cytokeratins. The median follow-up was 38 months (range, 10 to 70). The primary end point was survival. RESULTS: Cytokeratin-positive cells were detected in the bone marrow specimens of 2 of the 191 control patients with nonmalignant conditions (1 percent) and 199 of the 552 patients with breast cancer (36 percent). The presence of occult metastatic cells in bone marrow was unrelated to the presence or absence of lymph-node metastasis (P=0.13). After four years of follow-up, the presence of micrometastases in bone marrow was associated with the occurrence of clinically overt distant metastasis and death from cancer-related causes (P<0.001), but not with locoregional relapse (P=0.77). Of 199 patients with occult metastatic cells, 49 died of cancer, whereas of 353 patients without such cells, 22 died of cancer-related causes (P<0.001). Among the 301 women without lymph-node metastases, 14 of the 100 with bone marrow micrometastases died of cancer-related causes, as did 2 of the 201 without bone marrow micrometastases (P<0.001). The presence of occult metastatic cells in bone marrow, as compared with their absence, was an independent prognostic indicator of the risk of death from cancer (relative risk, 4.17; 95 percent confidence interval, 2.51 to 6.94; P<0.001), after adjustment for the use of systemic adjuvant chemotherapy. CONCLUSIONS: The presence of occult cytokeratin-positive metastatic cells in bone marrow increases the risk of relapse in patients with stage I, II, or III breast cancer.

Bone Marrow↗
PubMed2000

MLV-derived retroviral vectors selective for CD4-expressing cells and resistant to neutralization by sera from HIV-infected patients.

Retroviral vectors derived from amphotropic murine leukemia viruses (MLV) mediate gene transfer into almost all human cells and are thus not suitable for in vivo applications in gene therapy in which cell-specific gene delivery is required. We and others recently reported the generation of MLV-derived vectors pseudotyped by variants of the envelope glycoproteins (Env) of human immunodeficiency virus type 1 (HIV-1), thus displaying the CD4-dependent tropism of the parental lentivirus (Mammano et al., 1997, J. Virol. 71, 3341-3345; Schnierle et al., 1997, Proc. Natl. Acad. Sci. USA 76, 8640-8645). However, because of their HIV-1-derived envelopes these vectors are neutralized by HIV-specific antibodies present in some infected patients. To circumvent this problem, we pseudotyped MLV capsid particles with variants of Env proteins derived from the apathogenic simian immunodeficiency virus (SIVagm) of African green monkeys (AGM; Chlorocebus pygerythrus). Truncation of the C-terminal domain of the transmembrane protein was found to be necessary to allow formation of infectious pseudotype vectors. These [MLV(SIVagm)] vectors efficiently transduced various human CD4-expressing cell lines using the coreceptors CCR5 and Bonzo to enter target cells. Moreover, they were resistant to neutralization by antibodies directed against HIV-1. Therefore, [MLV(SIVagm)] vectors will be useful to study the mechanisms of SIVagm cell entry and for the selective gene transfer into CD4+ T-cells of AIDS patients.

Amino Acid Sequence↗
PubMed2000

Bone marrow "micrometastases" of epithelial tumors: detection and clinical relevance.

The detection and elimination of "micrometastases" or, more precisely, isolated disseminated tumor cells or minimal residual disease is one of the main current topics in clinical oncology. Immunocytochemical and molecular, polymerase chain reaction (PCR) based methods are the preferred methods. Bone marrow as a mesenchymal organ and a frequent location for distant metastases is very suitable to study isolated disseminated tumor cells. Under optimal conditions one tumor cell among one million mononuclear bone marrow cells can be detected by immunocytochemistry or molecular methods. The specificity, however, varies significantly depending on the assay conditions used by each individual group, with false positive rates below 1% and over 80% reported. Immunocytochemistry with antibodies against different epithelial markers has shown that the presence of isolated disseminated tumor cells in bone marrow is an independent prognostic variable in breast, colorectal, gastric, and non-small cell lung cancer. Due to the lack of a standardized assay, however, comparison of the results between different groups is very difficult. Before the assessment for isolated disseminated tumor cells in bone marrow or other organs can be transferred into routine clinical practice, standardized methods have to be developed which must be tested in multicenter prospective trials.

Bone Marrow↗
PubMed2000

Rapid flip-flop of phospholipids in endoplasmic reticulum membranes studied by a stopped-flow approach.

The transbilayer movement of short-chain spin-labeled and fluorescent 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) phospholipid analogs in rat liver microsomes is measured by stopped-flow mixing of labeled microsomes with bovine serum albumin (BSA) solution. Extraction of analogs from the outer leaflet of microsomes to BSA can be directly monitored in conjunction with electron paramagnetic resonance or fluorescence spectroscopy by taking advantage of the fact that the signal of spin-labeled or fluorescent analogs bound to BSA is different from that of the analogs inserted into membranes. From the signal kinetics, the transbilayer movement and the distribution of analogs in microsomal membranes can be derived provided the extraction of analogs by BSA is much faster in comparison to the transbilayer movement of analogs. Half-times of the back-exchange for spin-labeled and fluorescent analogs were <3.5 and <9.5 s, respectively. The unprecedented time resolution of the assay revealed that the transbilayer movement of spin-labeled analogs is much faster than previously reported. The half-time of the movement was about 16 s or even less at room temperature. Transmembrane movement of NBD-labeled analogs was six- to eightfold slower than that of spin-labeled analogs.

4-Chloro-7-nitrobenzofurazan↗
PubMed2000

Continuous measurement of rapid transbilayer movement of a pyrene-labeled phospholipid analogue.

The excimer forming capacity of the fluorescent moiety pyrene is employed to measure continuously the transbilayer (re)distribution of a pyrene-labeled phosphatidylcholine analogue (pyPC) in liposomal membranes. pyPC with a lauroyl residue (sn-1 position) and a short (butyroyl) fatty acid chain (sn-2 position) bearing the pyrene moiety incorporates rapidly into the outer leaflet of liposomes. The fluorescence intensities of excimers (I(E)) and of monomers (I(M)) of pyPC depend on the concentration of the analogue in a membrane leaflet. Therefore, the redistribution of pyPC from the outer to the inner leaflet can be followed by changes of the ratio I(E)/I(M). The transbilayer movement of pyPC in pure phospholipid vesicles is very slow indicated by a constant I(E)/I(M). However, addition of membrane active peptides (melittin, magainin 2 amide or a mutant of magainin 2 amide) induced a rapid translocation of pyPC from the outer to the inner leaflet. An approach is presented which allows estimating the transbilayer distribution of pyPC from the measured ratio I(E)/I(M).

Antimicrobial Cationic Peptides↗