Glutamate optimizes enzymatic activity under high hydrostatic pressure in Desulfovibrio species: effects on the ubiquitous thioredoxin system.

In piezophilic microorganisms, enzymes are optimized to perform under high hydrostatic pressure. The two major reported mechanisms responsible for such adaptation in bacterial species are changes in amino acids in the protein structure, favoring their activity and stability under high-pressure conditions, and the possible accumulation of micromolecular co-solutes in the cytoplasm. Recently, the accumulation of glutamate in the cytoplasm of piezophilic Desulfovibrio species has been reported under high-pressure growth conditions. In this study, analysis of the effect of glutamate on the enzymatic activity of the thioredoxin reductase/thioredoxin enzymatic complex of either a piezosensitive or a piezophilic microorganism confirms its role as a protective co-solute. Analysis of the thioredoxin structures suggests an adaptation both to the presence of glutamate and to high hydrostatic pressure in the enzyme from the piezophilic strain. Indeed, the presence of large surface pockets could counterbalance the overall compression that occurs at high hydrostatic pressure to maintain enzymatic activity. A lower isoelectric point and a greater dipolar moment than that of thioredoxin from the piezosensitive strain would allow the protein from the piezophilic strain to compensate for the presence of the charged amino acid glutamate to interact with its partner.

Laser-driven x-ray and proton micro-source and application to simultaneous single-shot bi-modal radiographic imaging.

Radiographic imaging with x-rays and protons is an omnipresent tool in basic research and applications in industry, material science and medical diagnostics. The information contained in both modalities can often be valuable in principle, but difficult to access simultaneously. Laser-driven solid-density plasma-sources deliver both kinds of radiation, but mostly single modalities have been explored for applications. Their potential for bi-modal radiographic imaging has never been fully realized, due to problems in generating appropriate sources and separating image modalities. Here, we report on the generation of proton and x-ray micro-sources in laser-plasma interactions of the focused Texas Petawatt laser with solid-density, micrometer-sized tungsten needles. We apply them for bi-modal radiographic imaging of biological and technological objects in a single laser shot. Thereby, advantages of laser-driven sources could be enriched beyond their small footprint by embracing their additional unique properties, including the spectral bandwidth, small source size and multi-mode emission.

[Lutetium-177-PSMA radioligand therapy : Consensus within the framework of GKV-funded care between the university hospitals in Aachen, Bonn, Düsseldorf, Essen, and Cologne and the MDK Nordrhein]. / Lutetium-177-PSMA-Radioligandentherapie : Konsensus im Rahmen der GKV-finanzierten Versorgung zwischen den Hochschulkliniken in Aachen, Bonn, Düsseldorf, Essen und Köln und dem MDK Nordrhein.

In the last 3 years, Lutetium-177 prostate-specific membrane antigen radioligand therapy (Lu-177-PSMA-RLT) has received increasing attention in nuclear medicine as a new form of treatment for castration-resistant metastatic prostate cancer. This therapy combines the radionuclide Lutetium-177, which has been therapeutically used in nuclear medicine for many years, with a molecular target of the transmembrane prostate-specific membrane antigen expressed by prostate cancer cells. Since there are no prospective randomized studies on Lu-177-PSMA-RLT and the question of reimbursement has repeatedly been the subject of review by the MDK Nordrhein (Medischenische Dienst der Krankenversicherung), there was a desire because of the increasing number of patients being treated to clarify under which circumstances Lu-177-PSMA-RLT can be reimbursed by German statutory health insurance. The goals of this article are to help treating physicians understand how this new therapy option works, to integrate it in the overall therapy concept for castration-resistant metastatic prostate cancer, and, above all, to use Lu-177-PSMA-RLT-based on the current data-at the right place in the therapy sequence of castration-resistant metastatic prostate cancer.

Enhanced Laser-Driven Ion Acceleration by Superponderomotive Electrons Generated from Near-Critical-Density Plasma.

We report on the experimental studies of laser driven ion acceleration from a double-layer target where a near-critical density target with a few-micron thickness is coated in front of a nanometer-thin diamondlike carbon foil. A significant enhancement of proton maximum energies from 12 to ∼30 MeV is observed when a relativistic laser pulse impinges on the double-layer target under linear polarization. We attributed the enhanced acceleration to superponderomotive electrons that were simultaneously measured in the experiments with energies far beyond the free-electron ponderomotive limit. Our interpretation is supported by two-dimensional simulation results.

A novel approach to electron data background treatment in an online wide-angle spectrometer for laser-accelerated ion and electron bunches.

Laser-based ion acceleration is driven by electrical fields emerging when target electrons absorb laser energy and consecutively leave the target material. A direct correlation between these electrons and the accelerated ions is thus to be expected and predicted by theoretical models. We report on a modified wide-angle spectrometer, allowing the simultaneous characterization of angularly resolved energy distributions of both ions and electrons. Equipped with online pixel detectors, the RadEye1 detectors, the investigation of this correlation gets attainable on a single shot basis. In addition to first insights, we present a novel approach for reliably extracting the primary electron energy distribution from the interfering secondary radiation background. This proves vitally important for quantitative extraction of average electron energies (temperatures) and emitted total charge.

A transportable Paul-trap for levitation and accurate positioning of micron-scale particles in vacuum for laser-plasma experiments.

We report on a Paul-trap system with large access angles that allows positioning of fully isolated micrometer-scale particles with micrometer precision as targets in high-intensity laser-plasma interactions. This paper summarizes theoretical and experimental concepts of the apparatus as well as supporting measurements that were performed for the trapping process of single particles.

Proton acceleration by irradiation of isolated spheres with an intense laser pulse.

We report on experiments irradiating isolated plastic spheres with a peak laser intensity of 2-3×10^{20}Wcm^{-2}. With a laser focal spot size of 10 µm full width half maximum (FWHM) the sphere diameter was varied between 520 nm and 19.3 µm. Maximum proton energies of ∼25 MeV are achieved for targets matching the focal spot size of 10 µm in diameter or being slightly smaller. For smaller spheres the kinetic energy distributions of protons become nonmonotonic, indicating a change in the accelerating mechanism from ambipolar expansion towards a regime dominated by effects caused by Coulomb repulsion of ions. The energy conversion efficiency from laser energy to proton kinetic energy is optimized when the target diameter matches the laser focal spot size with efficiencies reaching the percent level. The change of proton acceleration efficiency with target size can be attributed to the reduced cross-sectional overlap of subfocus targets with the laser. Reported experimental observations are in line with 3D3V particle in cell simulations. They make use of well-defined targets and point out pathways for future applications and experiments.

Ion Acceleration Using Relativistic Pulse Shaping in Near-Critical-Density Plasmas.

Ultraintense laser pulses with a few-cycle rising edge are ideally suited to accelerating ions from ultrathin foils, and achieving such pulses in practice represents a formidable challenge. We show that such pulses can be obtained using sufficiently strong and well-controlled relativistic nonlinearities in spatially well-defined near-critical-density plasmas. The resulting ultraintense pulses with an extremely steep rising edge give rise to significantly enhanced carbon ion energies consistent with a transition to radiation pressure acceleration.

Bright subcycle extreme ultraviolet bursts from a single dense relativistic electron sheet.

Double-foil targets separated by a low density plasma and irradiated by a petawatt-class laser are shown to be a copious source of coherent broadband radiation. Simulations show that a dense sheet of relativistic electrons is formed during the interaction of the laser with the tenuous plasma between the two foils. The coherent motion of the electron sheet as it transits the second foil results in strong broadband emission in the extreme ultraviolet, consistent with our experimental observations.

Dependence of laser-driven coherent synchrotron emission efficiency on pulse ellipticity and implications for polarization gating.

The polarization dependence of laser-driven coherent synchrotron emission transmitted through thin foils is investigated experimentally. The harmonic generation process is seen to be almost completely suppressed for circular polarization opening up the possibility of producing isolated attosecond pulses via polarization gating. Particle-in-cell simulations suggest that current laser pulses are capable of generating isolated attosecond pulses with high pulse energies.

Note: Thickness determination of freestanding ultra-thin foils using a table top laboratory extreme ultraviolet source.

We present a versatile and handy method allowing a thickness determination of freestanding thin plastic foils by its transmission characteristics in the extreme ultraviolet (EUV) spectrum. The method is based on a laser induced plasma source, emitting light in the EUV region, a compact double-mirror EUV monochromator operating at a fixed wavelength of 18.9 nm, and a CCD camera. The measurement delivers transmission values with a standard deviation of ΔT = 0.005 enabling foils thickness characterization with nm-accuracy at a given foil density and stoichiometric composition. Well characterized freestanding ultra-thin foils can be directly implemented in, e.g., high intensity laser matter experiments without further manipulation.

Spatiotemporal evolution of femtosecond laser pulses guided in air-clad fused-silica nanoweb.

We investigate nonlinear propagation and self-focusing of femtosecond Ti:sapphire laser pulses in an 800-nm-thick silica nanoweb fiber. Different dispersion regimes are accessible by launching TE- or TM-polarized light. Excitation in the anomalous dispersion regime (TM) results in pulse splitting and spectral broadening, which lead to supercontinuum generation, whereas, for normal dispersion (TE, excited close to a zero dispersion wavelength), self-phase modulation causes spectral broadening, which leads at higher power to beam collapse and the creation of a damage track.

Effect of saquinavir/ritonavir on P-glycoprotein activity in healthy volunteers using digoxin as a probe.

BACKGROUND: Saquinavir and ritonavir, both human immunodeficiency virus-1 protease inhibitors, also inhibit the adenosine triphosphate-dependent efflux pump P-glycoprotein (P-gp), which is located at a variety of anatomic sites, including the human intestine. P-gp plays an important role in the absorption, distribution and elimination of numerous drugs. This study investigated the inhibitory potential of multiple administrations of ritonavir-boosted saquinavir at the target therapeutic dose of 1,000 mg saquinavir/100 mg ritonavir twice daily on the pharmacokinetics of oral digoxin, a model P-gp substrate that is predominantly excreted as unchanged drug in the urine. METHODS: In an open-label, 1-sequence, 2-period crossover study, a single digoxin dose of 0.5 mg was administered orally on Day 1. From Days 11 through 26, participants received oral administration of saquinavir/ritonavir 1,000/100 mg twice daily. A second dose of digoxin was administered on Day 24. Blood and urine sampling for pharmacokinetic analyses of digoxin was performed at scheduled time points on Days 1 - 4 and Days 24 - 27. Serial blood samples were drawn to determine plasma levels of saquinavir and ritonavir on Days 21 - 24. Adverse event reports were collected. RESULTS: Of the 17 enrolled participants (9 males and 8 females) who received at least one dose of study medication, 16 completed the study. Two weeks of pretreatment with ritonavir and saquinavir resulted in a 1.27-fold increase in digoxin Cmax (90% confidence interval (1.05 - 1.54)) and a 1.49-fold increase in AUC0-72 (90% CI (1.32 - 1.69)). Renal clearance decreased by a factor 0.88 from 111 to 97.3 ml/min while digoxin half-life increased from 37.0 to 45.3 h. The unbound fraction of digoxin was almost unaffected. The changes in digoxin renal clearance and exposure (AUC0-72) following 2 weeks of treatment with saquinavir/ritonavir were found to be more pronounced among female participants compared with males. Plasma concentrations of saquinavir/ritonavir at trough and at 4 h postdose were within the expected ranges for each gender, with female participants showing higher concentrations than male participants. All three treatments were well tolerated, with no serious adverse events noted. Despite the higher digoxin exposure among females compared to males following saquinavir/ritonavir administration, overall safety profiles were similar. On electrocardiographic readings, a trend of a longer PR interval was noted with triple combination of agents. CONCLUSIONS: Pretreatment with saquinavir/ritonavir 1,000/100 mg twice daily increased digoxin exposure most likely via P-gp-inhibition. Given the relatively narrow therapeutic window of digoxin, caution should be exercised when these three drugs are administered together. It is recommended to reduce digoxin doses and to monitor digoxin serum concentrations.

[Transient periventricular echodensities (TPE) in preterm infants under 1500 gramms: an analysis of the outcome of the last 10 years]. / Transiente periventrikuläre Echodensitäten (TPE) bei Frühgeborenen unter 1500 Gramm: Eine Analyse des Outcome der letzten 10 Jahre.

BACKGROUND: The aim of our study was to analyze neurodevelopmental outcome in preterm infants with TPE. PATIENTS: 46 preterm infants with a birthweight (BW) < or = 1500 g were included in the study. They all had TPE in parieto-occipital location (some combined with frontal flares). Exclusion criteria were PVL II-IV, intraventricular hemorrhage, asphyxia and isolated frontal TPE. METHOD: We analyzed ultrasound scans (1992 - 2001) of patients with TPE (diameter > 1 cm, visible in both sagittal and coronal planes, duration > 7 days), and neurological follow-up visits (date of demission, corrected age of 4, 8, 12 months, and after that every 2 years until 6 years of age) regarding the neurodevelopmental outcome. We devided the patients in group 1 (duration of TPE 8 - 14 days) and group 2 (duration of TPE > 14 days) and then analyzed the groups regarding neurodevelopmental disorders and factors increasing the risk of occurrence of TPE. RESULTS: From 1992 - 2001 578 children < or = 1500 g were admitted at our department. 48 patients (8,3 %) fulfilled the entry criteria, 2 of them died, so 46 were included in the study. In group 1 (10 patients) the median gestational age (GA) was 28 weeks (range 25 - 31), median BW 1075 g (range 685 - 1430), 6 children were neurologically unimpaired, one developed a motor handicap, and 3 a psychomotor retardation. In group 2 (36 patients) the median GA was 28 weeks (range 25 - 35), median BW 1034,5 g (range 540 - 1470), 24 patients were neurologically unimpaired, 7 developed a motor handicap, and 5 a psychomotor retardation. No statistical difference was found between the groups neither in neurodevelopmental outcome nor factors increasing the risk of the occurrence of TPE. CONCLUSIONS: Obviously, the finding of TPE is of clinical relevance already at a duration of only 7 days regarding later neurological development.

Inhibitory effect of the pancreatic lipase C-terminal domain on intestinal lipolysis in rats fed a high-fat diet: chronic study.

OBJECTIVE: Previous in vitro experiments, as well as acute assays in rat showed that the C-terminal domain (CT-domain) of porcine pancreatic lipase behaves as a potent specific noncovalent inhibitor of pancreatic lipase. Nevertheless, the potential use of the CT-domain as a therapeutic tool against obesity in humans requires further investigation and would be best achieved using the human CT-domain. In the present study, we investigated the inhibitory effects of the recombinant human CT-domain, in vivo, upon chronic administration to rats fed a high-fat diet. DESIGN AND MEASUREMENT: The long-term in vivo study requiring relatively high amounts of the human CT-domain, the domain was overexpressed in Escherichia coli as inclusion bodies and an efficient refolding protocol was designed. The inhibitory effect of the recombinant human CT-domain on the activity of pancreatic lipase from different species was first investigated in vitro. Then chronic assays were performed for 4 weeks in rats fed a high-fat diet with or without a daily dose of 1.2 mg of CT-domain per kilogram rat. The time course of food intake, body weight, plasma parameters, liver lipids, faecal output of fat and total cholesterol were measured. RESULTS: A high yield of correctly folded recombinant human CT-domain was obtained using our refolding process, as evidenced by the capability of the recombinant domain to inhibit human horse and porcine pancreatic lipases in vitro. The recombinant human CT-domain had no influence on the food intake, but significantly reduced the body weight gain. As compared to control rats, higher amounts of total fat (mainly triglycerides and monoglycerides) and total cholesterol were found in the faeces of the rats treated with the CT-domain. Finally, a decrease in liver triglycerides and nonesterified cholesterol was observed while no significant effect could be detected on the plasma parameters. CONCLUSIONS: These results demonstrated that the CT-domain efficiently reduces in vivo, lipolysis and subsequently body weight gain in rat fed a high-fat diet. The CT-domain could, therefore, be effective in preventing obesity.

Granulocyte colony-stimulating factor (G-CSF) downregulates its receptor (CD114) on neutrophils and induces gelatinase B release in humans.

Despite the increasing use of granulocyte colony-stimulating factor (G-CSF) for the mobilization of stem cells and neutrophils, its pharmacodynamic actions are not fully understood. Because of the roles of G-CSF and gelatinase B in leucokinetics, we set out to characterize the interaction of G-CSF with its receptor in humans and its effects on gelatinase B release. G-CSF was infused at bolus doses of 1 microg/kg and 5 microg/kg, and compared to placebo and dexamethasone (1 mg/kg b.i.d), which enhances the plasma levels of endogenous G-CSF. The study was randomized, double-blind, four-way crossover, in eight healthy male volunteers. G-CSF dose-independently induced profound neutropenia (> 95%) within minutes and downregulated its own receptor (CD114) on neutrophils by 75%. The G-CSF/CD114 interaction dose-independently induced degranulation of neutrophils as evidenced by a 300-400% increase in CD11b expression. Degranulation induced up to a 10-fold increase in plasma levels of gelatinase B, an enzyme known to precipitate neutropenia and subsequent neutrophilia in animals. In this study, it was shown that G-CSF downmodulates CD114 expression on the surface of neutrophils in humans and the consequent degranulation enhances gelatinase B release into plasma, which may contribute to mobilization of neutrophils or stem cells.

Interactions of bile salt micelles and colipase studied through intermolecular nOes.

Colipase is a small protein (10 kDa), which acts as a protein cofactor for the pancreatic lipase. Various models of the activated ternary complex (lipase-colipase-bile salt micelles) have been proposed using detergent micelles, but no structural information has been established with bile salt micelles. We have investigated the organization of sodium taurodeoxycholate (NaTDC) micelles and their interactions with pig and horse colipases by homonuclear nuclear magnetic resonance (NMR) spectroscopy. The NMR data supply evidence that the folding of horse colipase is similar to that already described for pig colipase. Intermolecular nuclear Overhauser effects have shown that two conserved aromatic residues interact with NaTDC micelles.

The formate dehydrogenase-cytochrome c553 complex from Desulfovibrio vulgaris Hildenborough.

The electron transfer between formate dehydrogenase and cytochrome c553 from the anaerobic bacteria Desulfovibrio vulgaris Hildenborough has been investigated. Parameters of the electron transfer kinetics are reported. The ionic strength dependence of the complex formation has been evidenced. Two mutants of cytochrome c553 have been obtained using site-directed mutagenesis with the substitutions K62E and K62E,K63E. According to one-dimensional and two-dimensional NMR analysis, the two variants were found to have the same folding pattern as that of the wild-type cytochrome. The replacements of the lysine residues by acidic groups have important effects on the affinity between the two oxidoreduction partners. K62 and K63 are essential for recognition between the formate dehydrogenase and the cytochrome c553. Previous structural studies of cytochrome c553 have demonstrated the involvement of the polypeptide chain in the modulation of the particular low oxidoreduction potential of this cytochrome. The present study provides evidence that, during the evolution of cytochromes from the anaerobic metabolism to aerobic respiration and photosynthesis, the electrostatic distribution at the recognised encounter surface around the heme is highly conserved in all cytochromes.

Tyrosine 64 of cytochrome c553 is required for electron exchange with formate dehydrogenase in Desulfovibrio vulgaris Hildenborough.

Replacement of tyrosine 64 by alanine in cytochrome c553 from Desulfovibrio vulgarisHildenborough prevents electron transfer with the formate dehydrogenase. Biophysical and biochemical studies show that the protein is correctly folded and that the oxidoreduction potential is not modified. The solution structure of the mutant cytochrome determined by two-dimensional (2D) NMR clearly establishes that the overall fold of the molecule is nearly identical to that of the wild-type cytochrome. The electrostatic surface charge distributions for the wild-type and mutant cytochrome are similar, suggesting that the interaction site of the physiological partners is not modified by the mutation. The lack of the aromatic ring induces slight destabilization of the hydrophobic core of the molecule and modifications of the hydrogen bond at position 64, as well as conformational disorder of the side chain of K63. The loss of the hydrogen bond from tyrosine 64 and the increase of the solvent exposure of the heme are probably responsible of the loss of electron transfer between formate dehydrogenase and cytochrome c553.

1H-NMR study of the structural influence of Y64 substitution in Desulfovibrio vulgaris Hildenborough cytochrome c553.

Y64 has been replaced in cytochrome c553 from Desulfovibrio vulgaris Hildenborough by phenylalanine, leucine, valine, serine and alanine residues. An NMR study of structural variation induced in both oxidoreduction states of the molecule has been carried out by analysing observed chemical-shift variations. Dynamic changes were evidenced using NH exchange. We have observed that the substitution has a drastic effect on the stability of the molecule in the reduced state, although there is no effect on the reduction potential of the cytochrome. Y64-->F substitution induces particular effects on the NH exchange at the N-terminal, C-terminal and central alpha-helices and increases the stability of the oxidized molecule.