bluuebunny/crossref_metadata_2025 huggingface dataset:
The cardiac cycle time effect revisited: Temporal dynamics of the central‐vagal modulation of heart rate in human reaction time tasks
Authors: Somsen Riek J. M. , Jennings J. Richard , Van der Molen Maurits W.
Published: November 2004

Lacey and Lacey (1974) suggested that during reaction time tasks higher brain centers dynamically adjust efferent vagal nerve pulses to the sino‐atrial node of the heart, inducing phase‐dependent heart rate changes. Since then, animal and human neuro‐physiological results have provided evidence for this hypothesis. Higher subcortical and cortical brain centers may have reciprocal interactive pathways relating to autonomic control comparable to those at the level of peripheral autonomic changes and brain stem reflexes. In humans such central effects may be observed in the short latency vagal control of heart rate that has been studied mostly in reaction time (RT) tasks. RT task parameters modulate vagal pulses to the cardiac sino‐atrial node (SAN), which in turn exerts a phase‐dependent change in the ongoing cardiac interbeat interval. Simulations of human RT task effects in an animal model of heart rate change support this hypothesis. The current study examined evidence for vagal control of three human phasic heart rate responses in RT tasks. The evidence indicates that the initiation of an RT response triggers a reflexive shift from vagal activation to vagal inhibition. This shift is cardiac cycle phase dependent. Graded anticipatory cardiac deceleration during the warning interval of an RT task varies with task relevance and time uncertainty. This response may be part of a control process engaged in time keeping. Hence, temporal variables mediate the central‐autonomic‐vagal modulation of heart rate.
Current‐source density analysis of slow brain potentials during time estimation
Authors: Gibbons Henning , Rammsayer Thomas H.
Published: November 2004

Two event‐related potential studies were conducted to investigate differential brain correlates of temporal processing of intervals below and above 3–4 s. In the first experiment, 24 participants were presented with auditorily marked target durations of 2, 4, and 6 s that had to be reproduced. Timing accuracy was similar for all three target durations. As revealed by current‐source density analysis, slow‐wave components during both presentation and reproduction were independent of target duration. Experiment 2 examined potential modulating effects of type of interval (filled and empty) and presentation mode (randomized and blocked presentation of target durations). Behavioral and slow‐wave findings were consistent with those of Experiment 1. Thus, the present findings support the notion of a general timing mechanism irrespective of interval duration as proposed by scalar timing theory and pacemaker‐counter models of time estimation.
Posterior brain ERP patterns related to the go/no‐go task in children
Authors: Ciesielski Kristina T. , Harris Richard J. , Cofer Lynette F.
Published: November 2004

Event‐related potentials (ERPs) and performance correlates of inhibition of responses to no‐go stimuli were investigated in 6–12‐year‐old children and young adults. The percent of correct responses to go stimuli was high and similar in both groups; the percent of false alarm errors to no‐go was significantly higher in children. Effective inhibition of responses to no‐go stimuli was elucidated by a negative component of ERPs, the frontal‐central N2, with peak latency 230–430 ms after stimulus onset. In adults, N2 was larger to no‐go than to go stimuli, regardless of stimulus frequency. This effect was more prominent in tasks with high (25% no‐gos and 75% gos), than with low (75% no‐gos and 25% gos) inhibitory demand. In children, the parietal N2 was generally larger to rare than to frequent stimuli, and it was more specifically related to inhibition. The analysis of the relationship of no‐go N2 to the inhibitory content of stimuli, probability of stimuli, and the contextual task difficulty suggests that child/adult differences in behavioral responses and ERPs may be related to both the immaturity of the fronto‐parietal cortical‐cortical network and to different task strategies.
Magnetic resonance imaging of two suspected cases of global brain ischemia
Authors: Panarello Gregory L. , Dewey Curtis W. , Barone Georgina , Stefanacci Joseph D.
Published: November 2004

Objective: To describe the clinical findings and magnetic resonance imaging (MRI) characteristics of two animals with suspected global brain ischemia (GBI).

Series summary: Two brachycephalic animals (Persian cat and Boston terrier) presented for altered mentation, blindness, ataxia, and seizures after being anesthetized. Common to both anesthetic protocols was ketamine. Clinically, the neurological deficits developed rapidly and did not progress after the initial 24 hours. MRI findings were most consistent with GBI. MRI, including pre‐ and post‐T1 weighted, T2 weighted, and fluid‐attenuated inversion recovery (FLAIR) studies, was used to image these animals. Abnormalities were seen post‐T1, T2, and FLAIR. Both animals improved neurologically with supportive care over several weeks, but both remained blind.

New information: The use of MRI for diagnosing GBI.
Cooling Air Temperature Reduction in a Direct Transfer Preswirl System
Authors: Geis T. , Dittmann M. , Dullenkopf K.
Published: November 2004

This paper describes an experimental study of the cooling efficiency of a preswirl rotor-stator system equipped with a small number of preswirl nozzles of circular shape, located on a radius equal to that of the receiver disk holes. In the direct transfer cooling air system, total air temperatures were measured in the relative frame, i.e., inside the receiver holes by means of small total temperature probes for different throughflow rates, rotational Reynolds numbers and swirl ratios. The experimental data were compared with a simple theoretical model which predicts air temperatures in an “ideal” preswirl system. This comparison served to quantify the efficiency of this cooling scheme. In a subsequent one-dimensional analysis which took into account flow data obtained in an earlier experimental study by the same authors, two different mechanisms responsible for the elevated cooling air temperatures were determined. The new model considers in addition to the observed reduction of swirl due to viscous drag on the stator and mixing inside the rotor-stator cavity, the work put in by the rotor at high disk rotational Reynolds numbers and low cooling air flow rates.
Analysis of Cycle Configurations for the Modernization of Combined Heat and Power Plant by Fitting a Gas Turbine System
Authors: Chmielniak Tadeusz , Kosman Gerard , Kosman Wojciech
Published: November 2004

The application of a gas turbine generally allows to increase the number of possible configurations of cogenerated heat and electrical power systems, which became a significant substitute for classic, coal-fired power plants. They are characterized by better thermodynamical, economical, ecological, and operating indexes. Gas turbine units are also the best option for the modernization of existing power plants. This paper discusses the effectiveness of various technological configurations with gas turbines, which are to be applied during modernization projects of already existing conventional combined heat and power plants. In the analysis enthalpy and entropy methods were applied. Algorithms of the entropy method allow to determine the entropy generation in each section of a combined heat and power (CHP) plant. Several criteria were taken into consideration while analyzing the effectiveness of technological cycle configurations with gas turbines. These include the energy effectiveness, the efficiency of the HRSG and the steam cycle, the efficiency of the whole thermal electric power station, the exergetic efficiency of the HRSG and the steam cycle, and the fuel efficiency index. It was assumed that gas turbines operate under their nominal conditions. The composite curves were also taken into consideration while choosing the type of the turbine. The modernization project tends not to eliminate those existing power plant sections (machines and equipment), which are able to operate further. The project suggests that those units should remain in the system, which satisfy the applied durability criterion. The last phase of the optimization project focuses on the sensibility verification of several steam-gas CHP plant parameters and their influence on the whole system.
A Thermodynamic Analysis of Different Options to Break 60% Electric Efficiency in Combined Cycle Power Plants
Authors: Chiesa Paolo , Macchi Ennio
Published: November 2004

All major manufacturers of large size gas turbines are developing new techniques aimed at achieving net electric efficiency higher than 60% in combined cycle applications. An essential factor for this goal is the effective cooling of the hottest rows of the gas turbine. The present work investigates three different approaches to this problem: (i) the most conventional open-loop air cooling; (ii) the closed-loop steam cooling for vanes and rotor blades; (iii) the use of two independent closed-loop circuits: steam for stator vanes and air for rotor blades. Reference is made uniquely to large size, single shaft units and performance is estimated through an updated release of the thermodynamic code GS, developed at the Energy Department of Politecnico di Milano. A detailed presentation of the calculation method is given in the paper. Although many aspects (such as reliability, capital cost, environmental issues) which can affect gas turbine design were neglected, thermodynamic analysis showed that efficiency higher than 61% can be achieved in the frame of current, available technology.
Discharge Coefficients of Rotating Short Orifices With Radiused and Chamfered Inlets
Authors: Dittmann M. , Dullenkopf K. , Wittig S.
Published: November 2004

The secondary air system of modern gas turbine engines consists of numerous stationary or rotating passages to transport the cooling air, taken from the compressor, to thermally high loaded components that need cooling. Thereby the cooling air has to be metered by orifices to control the mass flow rate. Especially the discharge behavior of rotating holes may vary in a wide range depending on the actual geometry and the operating point. The exact knowledge of the discharge coefficients of these orifices is essential during the design process in order to guarantee a well adapted distribution of the cooling air inside the engine. This is crucial not only for a safe and efficient operation but also fundamental to predict the component’s life and reliability. In this paper two different methods to correlate discharge coefficients of rotating orifices are described and compared, both in the stationary and rotating frame of reference. The benefits of defining the discharge coefficient in the relative frame of reference will be pointed out. Measurements were conducted for two different length-to-diameter ratios of the orifices with varying inlet geometries. The pressure ratio across the rotor was varied for rotational Reynolds numbers up to ReΦ=8.6×105. The results demonstrate the strong influence of rotation on the discharge coefficient. An analysis of the complete data shows significant optimizing capabilities depending on the orifice geometry.
Robust Optimal Design in Multistage Expansion of a Gas Turbine Cogeneration Plant Under Uncertain Energy Demands
Authors: Yokoyama Ryohei , Ito Koichi , Murata Tatsuhiro
Published: November 2004

In designing cogeneration plants, the estimation of energy demands is an important work. However, many conditions under which energy demands are estimated have some uncertainty at the design stage. Therefore, designers should consider that energy demands have some uncertainty, evaluate the robustness in the performance under uncertain energy demands, and design plants rationally in consideration of the robustness. The authors have developed a robust optimal design method based on the minimax regret criterion for the single-stage planning of energy supply plants. In this paper, the method is extended for the multistage expansion planning. Under uncertain energy demands increasing stepwise, equipment capacities and utility contract demands as well as energy flow rates for each expansion period are determined in consideration of their sequential relationships to minimize the maximum regret in the annual total cost and satisfy all the possible energy demands for all the expansion periods. Through a case study on a gas turbine cogeneration plant for district energy supply, features of the economic robustness and the robust optimal design are clarified in relation to the uncertainty in energy demands and the numbers of years for the expansion periods.
Cycle Analysis of Gas Turbine–Fuel Cell Cycle Hybrid Micro Generation System
Authors: Uechi Hideyuki , Kimijima Shinji , Kasagi Nobuhide
Published: November 2004

Hybrid systems, which are based on a micro gas turbine (μGT) and a solid oxide fuel cell (SOFC), are expected to achieve much higher efficiency than traditional μGT’s. In this paper, the effects of cycle design parameters on the performance and feasibility of a μGT-SOFC hybrid system of 30 kW power output are investigated. It is confirmed that the hybrid system is much superior to a recuperated gas turbine in terms of its power generation efficiency and aptitude for small distributed generation. General design strategy is found that less direct fuel input to a combustor as well as higher recuperator effectiveness leads to higher generation efficiency, while higher steam-carbon ratio moderates requirements for the material strength. The best possible conceptual design of a 30-kW μGT-SOFC hybrid system is shown to give power generation efficiency over 65% (lower heating value).
Three-Way Catalytic Converter Modeling as a Modern Engineering Design Tool
Authors: Pontikakis G. N. , Konstantas G. S. , Stamatelos A. M.
Published: November 2004

The competition to deliver ultra low emitting vehicles at a reasonable cost is driving the automotive industry to invest significant manpower and test lab resources in the design optimization of increasingly complex exhaust aftertreatment systems. Optimization can no longer be based on traditional approaches, which are intensive in hardware use and lab testing. This paper discusses the extents and limitations of applicability of state-of-the-art mathematical models of catalytic converter performance. In-house software from the authors’ lab, already in use during the last decade in design optimization studies, updated with recent, important model improvements, is employed as a reference in this discussion. Emphasis is on the engineering methodology of the computational tools and their application, which covers quality assurance of input data, advanced parameter estimation procedures, and a suggested performance measure that drives the parameter estimation code to optimum results and also allows a less subjective assessment of model prediction accuracy. Extensive comparisons between measured and computed instantaneous emissions over full cycles are presented, aiming to give a good picture of the capabilities of state of the art engineering models of automotive catalytic converter systems.
Flamelet Modeling of Pollutant Formation in a Gas Turbine Combustion Chamber Using Detailed Chemistry for a Kerosene Model Fuel
Authors: Riesmeier E. , Honnet S. , Peters N.
Published: November 2004

Combustion and pollutant formation in a gas turbine combustion chamber is investigated numerically using the Eulerian particle flamelet model. The code solving the unsteady flamelet equations is coupled to an unstructured computational fluid dynamics (CFD) code providing solutions for the flow and mixture field from which the flamelet parameters can be extracted. Flamelets are initialized in the fuel-rich region close to the fuel injectors of the combustor. They are represented by marker particles that are convected through the flow field. Each flamelet takes a different pathway through the combustor, leading to different histories for the flamelet parameters. Equations for the probability of finding a flamelet at a certain position and time are additionally solved in the CFD code. To model the chemical properties of kerosene, a detailed reaction mechanism for a mixture of n-decane and 1,2,4-trimethylbenzene is used. It includes a detailed NOx submechanism and the buildup of polycyclic aromatic hydrocarbons up to four aromatic rings. The kinetically based soot model describes the formation of soot particles by inception, further growth by coagulation, and condensation as well as surface growth and oxidation. Simulation results are compared to experimental data obtained on a high-pressure rig. The influence of the model on pollutant formation is shown, and the effect of the number of flamelets on the model is investigated.
The Oxidation of Metal Alloy Foils in the Presence of Water Vapor
Authors: Rakowski James M.
Published: November 2004

Water vapor can be detrimental to the elevated temperature oxidation resistance of alloys that rely on the formation of a protective chromium oxide layer. The resulting degradation can be significant, particularly when such alloys are in the form of light gauge sheet and strip. Long-term test results will be presented for commercially available wrought austenitic stainless steels and nickel-base superalloys exposed at 1300°F and 1400°F in environments containing various levels of water vapor.
Amoxicillin-Clavulanate Therapy Increases Childhood Nasal Colonization by Methicillin-Susceptible
Staphylococcus aureus
Strains Producing High Levels of Penicillinase
Authors: Guillemot Didier , Bonacorsi Stephane , Blanchard John S. , Weber Philippe , Simon Sylvie , Guesnon Bruno , Bingen Edouard , Carbon Claude
Published: November 2004

We examined factors associated with penicillinase production by nasal carriage
Staphylococcus aureus
strains in 648 children aged 3 to 6 years attending 20 randomly sampled playschools. The children were prospectively monitored for drug use and medical events for 6 months and were then screened for
S. aureus
carriage. Isolates were tested for their susceptibility to penicillin G and methicillin, and penicillinase production by methicillin-susceptible, penicillin-resistant strains was quantified.
S. aureus
was isolated from 166 children (25.6%). Exposure to amoxicillin-clavulanate during the previous 3 months was associated with higher penicillinase production by penicillin-resistant, methicillin-susceptible strains (odds ratio, 3.6;
P
= 0.03). These results suggest that use of the amoxicillin-clavulanate combination could induce a herd selection process of
S. aureus
strains producing higher levels of penicillinase.
Pharmacokinetics and Effects of Ribavirin following Intraventricular Administration for Treatment of Subacute Sclerosing Panencephalitis
Authors: Hosoya Mitsuaki , Mori Shuichi , Tomoda Akemi , Mori Kenji , Sawaishi Yukio , Kimura Hiroshi , Shigeta Shiro , Suzuki Hitoshi
Published: November 2004

Ribavirin is a broad-spectrum antiviral drug with inhibitory activity against many RNA viruses, including measles virus. Five patients with subacute sclerosing panencephalitis (SSPE) were treated with ribavirin by intraventricular administration. Although there were transient side effects attributed to ribavirin, such as drowsiness, headache, lip and gingival swelling, and conjunctival hyperemia, intraventricular ribavirin therapy was generally safe and well tolerated. The cerebrospinal fluid (CSF) ribavirin concentration decreased, as described by a monoexponential function, after a single intraventricular dose. There was considerable interindividual variability, however, in the peak level and half-life. We aimed to adjust the individual dose and frequency of intraventricular administration based on the peak level and half-life of ribavirin in the CSF in order to maintain the CSF ribavirin concentration at the target level. Clinical effectiveness (significant neurologic improvement and/or a significant decrease in titers of hemagglutination inhibition antibodies against measles virus in CSF) was observed for four of five patients. For these four patients, CSF ribavirin concentrations were maintained at a level at which SSPE virus replication was almost completely inhibited in vitro and in vivo, whereas the concentration was lower in the patient without clinical improvement. These results suggest that intraventricular administration of ribavirin is effective against SSPE if the CSF ribavirin concentration is maintained at a high level. Intraventricular ribavirin therapy should be pursued further for its potential use for patients with SSPE and might be applied in the treatment of patients with encephalitis caused by other RNA viruses.
Pharmacodynamic Profiling of Piperacillin in the Presence of Tazobactam in Patients through the Use of Population Pharmacokinetic Models and Monte Carlo Simulation
Authors: Lodise Thomas P. , Lomaestro Ben , Rodvold Keith A. , Danziger Larry H. , Drusano George L.
Published: November 2004

The primary objectives of this analysis were to determine which pharmacokinetic model most accurately describes the elimination pathways for piperacillin in the presence of tazobactam through population pharmacokinetic modeling and to characterize its pharmacodynamic profile. Once the optimal pharmacokinetic model was identified, Monte Carlo simulation of 10,000 subjects with ADAPT II was performed to estimate the probability of attaining a target free-piperacillin concentration greater than the MIC for 50% of the dosing interval for 3.375 g every 6 h or every 4 h given as a 0.5-h infusion at each MIC between 0.25 and 32 μg/ml. In the population pharmacokinetic analysis, measurements of bias and precision, observed-predicted plots, andr2values were highly acceptable for all three models and all three models were appropriate candidates for the Monte Carlo simulation evaluation. Visual comparison of the distribution of the piperacillin concentrations at the pharmacodynamic endpoint—h 3 concentrations of a 6-h dosing interval—between the simulated populations and raw data revealed that the linear model was most reflective of the raw data at the pharmacodynamic endpoint, and the linear model was therefore selected for the target attainment analysis. In the target attainment analysis, administration of 3 g of piperacillin every 6 h resulted in a robust target attainment rate that exceeded 95% for MICs of ≤8 mg/liter. The 4-h piperacillin administration interval had a superior pharmacodynamic profile and provided target attainment rates exceeding 95% for MICs of ≤16 mg/liter. This study indicates that piperacillin-tazobactam should have utility for empirical therapy of hospital-onset infections.
Oral Activity of a Methylenecyclopropane Analog, Cyclopropavir, in Animal Models for Cytomegalovirus Infections
Authors: Kern Earl R. , Bidanset Deborah J. , Hartline Caroll B. , Yan Zhaohua , Zemlicka Jiri , Quenelle Debra C.
Published: November 2004

We reported previously that purine 2-(hydroxymethyl)methylenecyclopropane analogs have good activity against cytomegalovirus infection. A second-generation analog, (
Z
)-9-{[2,2-bis-(hydroxymethyl)cyclopropylidene]methyl}guanine (ZSM-I-62, cyclopropavir [CPV]), has particularly good activity against murine and human cytomegaloviruses (MCMV and HCMV) in vitro. To determine the oral activity of this compound in vivo, BALB/c or severe combined immunodeficient (SCID) mice infected with MCMV and two models using SCID mice implanted with human fetal tissue and subsequently infected with HCMV were used. In MCMV-infected normal mice, CPV at 10 mg/kg of body weight was highly effective in preventing mortality when administered at 24, 48, or 72 h post-viral inoculation and reduced titers of virus in tissues of SCID mice by 2 to 5 log
10
. In one HCMV model, human fetal retinal tissue was implanted into the anterior chamber of the mouse eye and inoculated with the Toledo strain of HCMV, and in the second, human fetal thymus and liver tissues were implanted under the kidney capsule of mice and then inoculated with HCMV. In general, replication of HCMV in both types of implant tissue increased from 7 through 21 to 28 days and then gradually decreased to undetectable levels by 8 weeks postinfection. Oral treatment with 45 or 15 mg of CPV/kg initiated 24 h after infection was highly effective in reducing replication to undetectable levels in both models and was generally more effective than ganciclovir. These data indicate that the methylenecyclopropane analog, CPV, was highly efficacious in these four animal models and should be evaluated for use in HCMV infections in humans.
Antistaphylococcal Activity of WCK 771, a Tricyclic Fluoroquinolone, in Animal Infection Models
Authors: Patel Mahesh V. , De Souza Noel J. , Gupte Shrikant V. , Jafri Mohammad A. , Bhagwat Sachin S. , Chugh Yati , Khorakiwala Habil F. , Jacobs Michael R. , Appelbaum Peter C.
Published: November 2004

WCK 771, the arginine salt ofS-(−)-nadifloxacin, was evaluated in animal models of staphylococcal infection and in vitro. For 302 methicillin-susceptible strains the MIC at which 50% of isolates are inhibited (MIC50) and the MIC90of WCK 771 were 0.03 and 0.03 μg/ml, respectively, and for 198 methicillin-resistant strains the MIC50and the MIC90were 0.5 and 1.0 μg/ml, respectively. All methicillin-susceptible staphylococci were quinolone susceptible, and almost all methicillin-resistant staphylococci were quinolone resistant. WCK 771 was more potent than moxifloxacin, trovafloxacin, levofloxacin, and ciprofloxacin and had potency comparable to that of clinafloxacin. Only WCK 771 and clinafloxacin demonstrated strong potencies against vancomycin-intermediateStaphylococcus aureusstrains (MICs = 1 μg/ml). WCK 771 is not a substrate of the NorA pump, as evident from the lack of an effect of reserpine on the MICs and similar protective doses against infections caused by efflux-positive and -negative staphylococci. WCK 771 was effective by both the oral and the subcutaneous routes in mice infected intraperitoneally with quinolone-susceptible methicillin-susceptibleS. aureus(MSSA) strains. For infections caused by quinolone-resistant methicillin-resistantS. aureus(MRSA) strains, the activity of WCK 771 administered subcutaneously was superior to those of trovafloxacin and sparfloxacin, with a 50% effective dose range of 27.8 to 46.8 mg/kg of body weight. The activity of WCK 771 was superior to those of moxifloxacin, vancomycin, and linezolid in a mouse cellulitis model of infection caused by one MSSA and two MRSA strains, with effective doses of 2.5 and 5 mg/kg for the MSSA strain and 10-fold higher effective doses for MRSA strains. WCK 771, like vancomycin and linezolid, eradicated MRSA from mouse liver, spleen, kidney, and lung when it was administered subcutaneously at a dose of 50 mg/kg for four doses. These studies have demonstrated the effectiveness of WCK 771, administered orally and parenterally, for the treatment of diverse staphylococcal infections in mice, including those caused by quinolone-resistant strains.
Derivatives of a Vancomycin-ResistantStaphylococcus aureusStrain Isolated at Hershey Medical Center
Authors: Bozdogan Bülent , Ednie Lois , Credito Kim , Kosowska Klaudia , Appelbaum Peter C.
Published: November 2004

Antimicrobial susceptibilities and genetic relatedness of the vancomycin-resistantStaphylococcus aureusstrain (VRSA) isolated at Hershey, Pa. (VRSA Hershey), and its vancomycin-susceptible and high-level-resistant derivatives were studied and compared to 32 methicillin-resistantS. aureusstrains (MRSA) isolated from patients and medical staff in contact with the VRSA patient. Derivatives of VRSA were obtained by subculturing six VRSA colonies from the original culture with or without vancomycin. Ten days of drug-free subculture caused the loss ofvanAin two vancomycin-susceptible derivatives for which vancomycin MICs were 1 to 4 μg/ml. Multistep selection of three VRSA clones with vancomycin for 10 days increased vancomycin MICs from 32 to 1,024 to 2,048 μg/ml. MICs of teicoplanin, dalbavancin, and oritavancin were also increased from 4, 0.5, and 0.12 to 64, 1, and 32 μg/ml, respectively. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing analysis indicated that VRSA Hershey was thevanA-acquired variety of a common MRSA clone in our hospital with sequence type 5 (ST5). Three of five vancomycin-intermediateS. aureusstrains tested from geographically different areas were also ST5, and the Michigan VRSA was ST371, a one-allele variant of ST5. Derivatives of VRSA Hershey had differences in PFGE profiles and the size of SmaI fragment that carries thevanAgene cluster, indicating instability of this cluster in VRSA Hershey. However induction with vancomycin increased glycopeptide MICs and stabilized the resistance.
Pharmacokinetics in Animals and Humans of a First-in-Class Peptide Deformylase Inhibitor
Authors: Ramanathan-Girish Sandhya , McColm Juliet , Clements John M. , Taupin Phil , Barrowcliffe Sue , Hevizi John , Safrin Sharon , Moore Clive , Patou Gary , Moser Heinz , Gadd Alison , Hoch Ute , Jiang Vernon , Lofland Denene , Johnson Kirk W.
Published: November 2004

BB-83698, a potent and selective inhibitor of peptide deformylase, was the first compound of this novel antibacterial class to progress to clinical trials. Single- and/or multiple-dose studies with doses ranging from 10 to 50 mg of BB-83698/kg of body weight were done with mice, rats, and dogs. Intravenous pharmacokinetics were characterized by low to moderate clearances and moderate volumes of distribution for all species. In dogs, but not in rodents, central nervous system (CNS) effects were dose limiting for intravenously administered BB-83698 and were suspected to be related to a high maximum concentration of the agent in plasma (
C
max
) rather than to total systemic exposure. Controlled infusion studies with dogs demonstrated that CNS effects could be avoided without compromising systemic exposure by reducing the
C
max
. A randomized, double-blind, placebo-controlled, five-way-crossover, single-dose-escalation, phase I study to explore the safety, tolerability, and pharmacokinetics of intravenous BB-83698 at doses ranging from 10 to 475 mg was performed with healthy male volunteers. Systemic exposures were generally in linear relationships with administered doses in animals and humans. Pharmacokinetics were consistent, predictable, and exhibited good allometric scaling among all species (
r
2
>0.98). Moreover, BB-83698 dosing in humans proceeded to a predicted efficacious exposure (the area under the concentration-time curve/MIC ratio, up to 184) without any clinically significant adverse effects.
Antimycoplasmal Activity of Hydroxytyrosol
Authors: Furneri Pio Maria , Piperno Anna , Sajia Antonella , Bisignano Giuseppe
Published: November 2004

The aim of this study was to investigate the in vitro antimycoplasmal activity of hydroxytyrosol. Twenty strains ofMycoplasma hominis, three strains ofMycoplasma fermentans, and one strain ofMycoplasma pneumoniaewere used. ForM. pneumoniae,M. hominis, andM. fermentans, the MICs were 0.5, 0.03 (for 90% of the strains tested), and 0.25 μg/ml, respectively.
Emergence in Asian Countries of
Staphylococcus aureus
with Reduced Susceptibility to Vancomycin
Authors: Song Jae-Hoon , Hiramatsu Keiichi , Suh Ji Yoeun , Ko Kwan Soo , Ito Teruyo , Kapi Maria , Kiem Sungmin , Kim Yeon-Sook , Oh Won Sup , Peck Kyong Ran , Lee Nam Yong
Published: November 2004

To investigate the prevalence of
Staphylococcus aureus
with reduced susceptibility to vancomycin among methicillin-resistant
S. aureus
(MRSA) strains in Asian countries, a total of 1,357 clinical isolates of MRSA collected from 12 Asian countries were screened by using brain heart infusion agar plates containing 4 mg of vancomycin per liter. The presence of strains that were heterointermediately resistant to vancomycin (hVISA) was confirmed by population analysis. Of 347 (25.6%) MRSA isolates that grew on the screening agar plates, 58 isolates (4.3%) were hVISA. hVISA strains were found in India, South Korea, Japan, the Philippines, Singapore, Thailand, and Vietnam. However, neither vancomycin-intermediate
S. aureus
nor vancomycin-resistant
S. aureus
isolates were found among MRSA isolates from Asian countries in this survey.
Replicase-Binding Sites on Plus- and Minus-Strand Brome Mosaic Virus RNAs and Their Roles in RNA Replication in Plant Cells
Authors: Choi S.-K. , Hema M. , Gopinath K. , Santos J. , Kao C.
Published: November 2004

The
cis
-acting elements for
Brome mosaic virus
(BMV) RNA synthesis have been characterized primarily for RNA3. To identify additional replicase-binding elements, nested fragments of all three of the BMV RNAs, both plus- and minus-sense fragments, were constructed and tested for binding enriched BMV replicase in a template competition assay. Ten RNA fragments containing replicase-binding sites were identified; eight were characterized further because they were more effective competitors. All eight mapped to noncoding regions of BMV RNAs, and the positions of seven localized to sequences containing previously characterized core promoter elements (C. C. Kao, Mol. Plant Pathol. 3:55-62, 2001), thus suggesting the identities of the replicase-binding sites. Three contained the tRNA-like structures that direct minus-strand RNA synthesis, three were within the 3′ region of each minus-strand RNA that contained the core promoter for genomic plus-strand initiation, and one was in the core subgenomic promoter. Single-nucleotide mutations known previously to abolish RNA synthesis in vitro prevented replicase binding. When tested in the context of the respective full-length RNAs, the same mutations abolished BMV RNA synthesis in transfected barley protoplasts. The eighth site was within the intercistronic region (ICR) of plus-strand RNA3. Further mapping showed that a sequence of 22 consecutive adenylates was responsible for binding the replicase, with 16 being the minimal required length. Deletion of the poly(A) sequence was previously shown to severely debilitate BMV RNA replication in plants (E. Smirnyagina, Y. H. Hsu, N. Chua, and P. Ahlquist, Virology 198:427-436, 1994). Interestingly, the B box motif in the ICR of RNA3, which has previously been determined to bind the 1a protein, does not bind the replicase. These results identify the replicase-binding sites in all of the BMV RNAs and suggest that the recognition of RNA3 is different from that of RNA1 and RNA2.
Heptad Repeat 2-Based Peptides Inhibit Avian Sarcoma and Leukosis Virus Subgroup A Infection and Identify a Fusion Intermediate
Authors: Netter Robert C. , Amberg Sean M. , Balliet John W. , Biscone Mark J. , Vermeulen Arwen , Earp Laurie J. , White Judith M. , Bates Paul
Published: November 2004

Fusion proteins of enveloped viruses categorized as class I are typified by two distinct heptad repeat domains within the transmembrane subunit. These repeats are important structural elements that assemble into the six-helix bundles characteristic of the fusion-activated envelope trimer. Peptides derived from these domains can be potent and specific inhibitors of membrane fusion and virus infection. To facilitate our understanding of retroviral entry, peptides corresponding to the two heptad repeat domains of the avian sarcoma and leukosis virus subgroup A (ASLV-A) TM subunit of the envelope protein were characterized. Two peptides corresponding to the C-terminal heptad repeat (HR2), offset from one another by three residues, were effective inhibitors of infection, while two overlapping peptides derived from the N-terminal heptad repeat (HR1) were not. Analysis of envelope mutants containing substitutions within the HR1 domain revealed that a single amino acid change, L62A, significantly reduced sensitivity to peptide inhibition. Virus bound to cells at 4°C became sensitive to peptide within the first 5 min of elevating the temperature to 37°C and lost sensitivity to peptide after 15 to 30 min, consistent with a transient intermediate in which the peptide binding site is exposed. In cell-cell fusion experiments, peptide inhibitor sensitivity occurred prior to a fusion-enhancing low-pH pulse. Soluble receptor for ASLV-A induces a lipophilic character in the envelope which can be measured by stable liposome binding, and this activation was found to be unaffected by inhibitory HR2 peptide. Finally, receptor-triggered conformational changes in the TM subunit were also found to be unaffected by inhibitory peptide. These changes are marked by a dramatic shift in mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, from a subunit of 37 kDa to a complex of about 80 kDa. Biotinylated HR2 peptide bound specifically to the 80-kDa complex, demonstrating a surprisingly stable envelope conformation in which the HR2 binding site is exposed. These experiments support a model in which receptor interaction promotes formation of an envelope conformation in which the TM subunit is stably associated with its target membrane and is able to bind a C-terminal peptide.
Induction of Id1 and Id3 by Latent Membrane Protein 1 of Epstein-Barr Virus and Regulation of p27/Kip and Cyclin-Dependent Kinase 2 in Rodent Fibroblast Transformation
Authors: Everly David N. , Mainou Bernardo A. , Raab-Traub Nancy
Published: November 2004

Latent membrane protein 1 (LMP1), the Epstein-Barr virus oncoprotein, activates NF-κB, phosphatidylinositol 3-kinase, mitogen-activated protein kinase, and c-Jun N-terminal kinase signaling. To determine global transcriptional changes induced by LMP1 in epithelial cells, genomic analysis of C33A cells stably expressing LMP1 was performed. Relatively few genes were induced by LMP1. Expression of two members of the Id (inhibitor of differentiation) family of proteins, Id1 and Id3, was induced in the presence of LMP1 and confirmed by mRNA and protein in C33A and Rat-1 cells. In Rat-1 foci transformed by LMP1, Id1 protein was also increased. Id proteins are known negative regulators of E-box proteins that positively regulate p16 and potentially other cyclin-dependent kinase inhibitors (cdki's). In LMP1-expressing Rat-1 cells, cdki p27 was specifically downregulated. Decreased p27 was correlated with increased levels of Cdk2 and increased levels of phosphorylated retinoblastoma protein. This study describes new properties of LMP1 that likely contribute to transformation and oncogenesis.
Coxsackievirus B3 Infection Induces
cyr61
Activation via JNK To Mediate Cell Death
Authors: Kim Sun-Mi , Park Jung-Hyun , Chung Sun-Ku , Kim Joo-Young , Hwang Ha-Young , Chung Kwang-Chul , Jo Inho , Park Sang-Ick , Nam Jae-Hwan
Published: November 2004

Coxsackievirus B3 (CVB3), an enterovirus in the
Picornavirus
family, is the most common human pathogen associated with myocarditis and idiopathic dilated cardiomyopathy. We found upregulation of the cysteine-rich protein gene (
cyr61
) after CVB3 infection in HeLa cells with a cDNA microarray approach, which is confirmed by Northern blot analysis. It is also revealed that the extracellular amount of Cyr61 protein was increased after CVB3 infection in HeLa cells.
cyr61
is an early-transcribed gene, and the Cyr61 protein is secreted into the extracellular matrix. Its function is related to cell adhesion, migration, and neuronal cell death. Here, we show that activation of the
cyr61
promoter by CVB3 infection is dependent on JNK activation induced by CVB3 replication and viral protein expression in infected cells. To explore the role of Cyr61 protein in infected HeLa cells, we transiently overexpressed
cyr61
and infected HeLa cells with CVB3. This increased CVB3 growth in the cells and promoted host cell death by viral infection, whereas down-expression of
cyr61
with short interfering RNA reduced CVB3 growth and showed resistance to cell death by CVB3 infection. In conclusion, we have demonstrated a new role for
cyr61
in HeLa cells infected with CVB3, which is associated with the cell death induced by virus infection. These data thus expand our understanding of the physiological functions of
cyr61
in virus-induced cell death and provide new insights into the cellular factors involved.
A Conserved Histidine in the ij Loop of the Semliki Forest Virus E1 Protein Plays an Important Role in Membrane Fusion
Authors: Chanel-Vos Chantal , Kielian Margaret
Published: November 2004

The enveloped alphavirus Semliki Forest virus (SFV) infects cells via a low pH-triggered membrane fusion reaction mediated by the E1 protein. E1 is a class II fusion protein that contains the hydrophobic fusion peptide loop and converts to a stable homotrimer during the fusion reaction. Intriguingly, the fusion loop is closely associated with a loop connecting the i and j β-strands. This ij loop plays a role in the cholesterol dependence of membrane fusion and is specifically susceptible to proteolysis in the protease-resistant E1 homotrimer. The SFV ij loop contains a histidine residue at position 230. Sequence comparisons revealed that an analogous histidine is completely conserved in all alphavirus and flavivirus fusion proteins. An E1 H230A mutant was constructed using the SFV infectious clone. Although cells infected with H230A RNA produced virus particles, these virions were completely noninfectious and were blocked in both cell-cell fusion and lipid mixing assays. The H230A virions efficiently bound to cell surface receptors and responded to low pH by undergoing acid-dependent conformational changes including dissociation of the E1/E2 dimer, exposure of the fusion loop, association with target liposomes, exposure of acid-conformation-specific epitopes, and formation of the stable E1 homotrimer. Studies with a soluble fragment of E1 showed that the mutant protein was defective in lipid-dependent conformational changes. Our results indicate that the E1 ij loop and the conserved H230 residue play a critical role in alphavirus-membrane fusion and suggest the presence of a previously undescribed late intermediate in the fusion reaction.
Furin-Mediated Cleavage of the Feline Foamy Virus Env Leader Protein
Authors: Geiselhart Verena , Bastone Patrizia , Kempf Tore , Schnölzer Martina , Löchelt Martin
Published: November 2004

The molecular biology of spuma or foamy retroviruses is different from that of the other members of the
Retroviridae
. Among the distinguishing features, the N-terminal domain of the foamy virus Env glycoprotein, the 16-kDa Env leader protein Elp, is a component of released, infectious virions and is required for particle budding. The transmembrane protein Elp specifically interacts with N-terminal Gag sequences during morphogenesis. In this study, we investigate the mechanism of Elp release from the Env precursor protein. By a combination of genetic, biochemical, and biophysical methods, we show that the feline foamy virus (FFV) Elp is released by a cellular furin-like protease, most likely furin itself, generating an Elp protein consisting of 127 amino acid residues. The cleavage site fully conforms to the rules for an optimal furin site. Proteolytic processing at the furin cleavage site is required for full infectivity of FFV. However, utilization of other furin proteases and/or cleavage at a suboptimal signal peptidase cleavage site can partially rescue virus viability. In addition, we show that FFV Elp carries an N-linked oligosaccharide that is not conserved among the known foamy viruses.
C-Terminal nsP1a Protein of Human Astrovirus Colocalizes with the Endoplasmic Reticulum and Viral RNA
Authors: Guix Susana , Caballero Santiago , Bosch Albert , Pintó Rosa M.
Published: November 2004

Computational and biological approaches were undertaken to characterize the role of the human astrovirus nonstructural protein nsP1a/4, located at the C-terminal fragment of nsP1a. Computer analysis reveals sequence similarities to other nonstructural viral proteins involved in RNA replication and/or transcription and allows the identification of a glutamine- and proline-rich region, the prediction of many phosphorylation and O-glycosylation sites, and the occurrence of a KKXX-like endoplasmic reticulum retention signal. Immunoprecipitation analysis with an antibody against a synthetic peptide of the nsP1a/4 sequence detected polyprotein precursors of 160, 75, and 38 to 40 kDa as well as five smaller proteins in the range of 21 to 27 kDa. Immunofluorescence labeling showed that the nsP1a/4 protein is accumulated at the perinuclear region, in association with the endoplasmic reticulum and the viral RNA. These results suggest the involvement of nsP1a/4 protein in the RNA replication process in endoplasmic reticulum-derived intracellular membranes.
Oligomerization of Hantavirus N Protein: C-Terminal α-Helices Interact To Form a Shared Hydrophobic Space
Authors: Kaukinen Pasi , Kumar Vibhor , Tulimäki Kirsi , Engelhardt Peter , Vaheri Antti , Plyusnin Alexander
Published: November 2004

The structure of the nucleocapsid protein of bunyaviruses has not been defined. Earlier we have shown that Tula hantavirus N protein oligomerization is dependent on the C-terminal domains. Of them, the helix-loop-helix motif was found to be an essential structure. Computer modeling predicted that oligomerization occurs via helix protrusions, and the shared hydrophobic space formed by amino acids residues 380-IILLF-384 in the first helix and 413-LI-414 in the second helix is responsible for stabilizing the interaction. The model was validated by two approaches. First, analysis of the oligomerization capacity of the N protein mutants performed with the mammalian two-hybrid system showed that both preservation of the helix structure and formation of the shared hydrophobic space are crucial for the interaction. Second, oligomerization was shown to be a prerequisite for the granular pattern of transiently expressed N protein in transfected cells. N protein trimerization was supported by three-dimensional reconstruction of the N protein by electron microscopy after negative staining. Finally, we discuss how N protein trimerization could occur.
Identification of Residues in the Dengue Virus Type 2 NS2B Cofactor That Are Critical for NS3 Protease Activation
Authors: Niyomrattanakit Pornwaratt , Winoyanuwattikun Pakorn , Chanprapaph Santad , Angsuthanasombat Chanan , Panyim Sakol , Katzenmeier Gerd
Published: November 2004

Proteolytic processing of the dengue virus polyprotein is mediated by host cell proteases and the virus-encoded NS2B-NS3 two-component protease. The NS3 protease represents an attractive target for the development of antiviral inhibitors. The three-dimensional structure of the NS3 protease domain has been determined, but the structural determinants necessary for activation of the enzyme by the NS2B cofactor have been characterized only to a limited extent. To test a possible functional role of the recently proposed Φx
3
Φ motif in NS3 protease activation, we targeted six residues within the NS2B cofactor by site-specific mutagenesis. Residues Trp62, Ser71, Leu75, Ile77, Thr78, and Ile79 in NS2B were replaced with alanine, and in addition, an L75A/I79A double mutant was generated. The effects of these mutations on the activity of the NS2B(H)-NS3pro protease were analyzed in vitro by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of autoproteolytic cleavage at the NS2B/NS3 site and by assay of the enzyme with the fluorogenic peptide substrate GRR-AMC. Compared to the wild type, the L75A, I77A, and I79A mutants demonstrated inefficient autoproteolysis, whereas in the W62A and the L75A/I79A mutants self-cleavage appeared to be almost completely abolished. With exception of the S71A mutant, which had a
k
cat
/
K

m

value for the GRR-AMC peptide similar to that of the wild type, all other mutants exhibited drastically reduced
k
cat
values. These results indicate a pivotal function of conserved residues Trp62, Leu75, and Ile79 in the NS2B cofactor in the structural activation of the dengue virus NS3 serine protease.