bluuebunny/crossref_metadata_2025 huggingface dataset:
Diffusion-Mediated Production of Protein Gradients by Way of Variable Depth Hydrogel Microstamps
Authors: Thompson Dewayne L. , Madon Ashley E. , Trinkle Christine A.
Published: June 2010

The precise application of proteins and other biomolecules to create patterned surfaces is an important step in many processes, including the creation of biosensors and directed cell growth for tissue engineering [1, 2]. While traditional poly-dimethylsiloxane (PDMS) elastomeric stamps have been used to successfully transfer proteins with sub-micron resolution, the resulting patterns are limited to a single, uniform protein concentration. The technique presented here utilizes varied-topography hydrophilic stamps as a diffusion medium, allowing protein gradients to be easily applied and accurately reproduced. Specifically, stamps of a 2% agarose hydrogel are used to demonstrate variable-concentration patterning of a fluorescently-labeled protein.
Analysis of Flow and Wall Shear Stress in Curvature Induced Dorsal Aneurysms: Effect of Arterial Curvature
Authors: Ramu Arun , Wang Guo-Xiang
Published: June 2010

Intracranial aneurysms are abnormal enlargement in the walls of cerebral arteries. The rupture of aneurysms is the leading cause of subarachnoid hemorrhage (SAH), with a high mortality and morbidity rate. A majority of saccular cerebral aneurysms occur at sites of arterial bifurcations. However, a good percentage of aneurysms are curvature induced and are found along the cavernous arterial segment. The occurrence of such non branching aneurysms, clinically called dorsal aneurysms, can be related to the increased wall shear stress at the curved arteries. The rupture of aneurysms usually occurs at the dome region, which is subjected to reduced wall shear stress (wss) owing to low re-circulating flow. Hence it is important to understand the impact of arterial curvature on the WSS distribution along the dome of aneurysms. Previously, studies have not taken into account the aspect of low WSS along the dome region. In the present 3-d computational fluid dynamic approach, we investigate the impact of varying arterial curvature on spherical dorsal aneurysms. The primary velocity patterns, the WSS distribution along the dome of the aneurysm and the area of increased WSS have been quantified for steady flow conditions.
Mechanical Properties of Amorphous Metal Honeycombs for Ballistic Applications
Authors: Hanan Jay C. , Jayakumar Balaji , Bhat Advait
Published: June 2010

Body armor technology continually improves in performance and endurance with the introduction of new designs and materials. Composite layered armor systems comprised of ceramics and fabric based materials are at the leading edge. Over the years, these systems have seen preference over their metal counterparts due to reduced weight. High performance polyethylene (Spectra, Dyneema), Aramids (Kevlar, Twaron, Zylon) and other composite materials, [1] have further improved dynamic resistance in body armor with reasonable strength-to-weight ratio.
Effects of Scaffold Design Parameters and Local Environment on Biopolymer Scaffold Degradation
Authors: Yan Karen Chang , Roros Christina , Swift Maria , Sun Wei
Published: June 2010

Biodegradable polymers have been extensively used in tissue engineering as temporary porous scaffolding structures to host or guide tissue/organ regeneration. The advantage lies in that the polymers degrade away harmlessly in the body. One key requirement in tissue scaffold design is to control the scaffolding degradation and erosion that synchronize the new cell/tissue growth. The degradation and/or erosion for biodegradable polymers are influenced by a multitude of factors. This paper presents an in vitro experimental study to examine effects of scaffold design parameters and local environment on the degradation rate of biopolymer scaffolds. Parameters considered are scaffold pore size, material composition and cellular activity.
Numerical Simulation of Blood Flow in Patient-Specific Stenotic Carotid Bifurcation Geometries
Authors: Cummins Megan , Rossmann Jenn S.
Published: June 2010

The hemodynamics and fluid mechanical forces in blood vessels have long been implicated in the deposition and growth of atherosclerotic plaque. Detailed information about the hemodynamics in vessels affected by significant plaque deposits can provide insight into the mechanisms and likelihood of plaque weakening and rupture. In the current study, the governing equations are solved in their finite volume formulation in several patient-specific geometries. Recirculation zones, vortex shedding, and secondary flows are captured. The forces on vessel walls are shown to correlate with unstable plaque deposits. The results of these simulations suggest morphological features that may usefully supplement percent stenosis as a predictor of plaque vulnerability.
Viscoelastic Properties of Brain Tissue Under High-Rate Large Deformation
Authors: Shafieian Mehdi , Darvish Kurosh
Published: June 2010

The nonlinearity of brain tissue material behavior for large deformations at high strain rates was investigated. The viscoelastic properties of brain tissue under high rate ramp- and hold shear strains were determined and nonlinearity in the elastic and time dependent properties of the tissue were examined based on modeling the experimental data. The results revealed that the elastic response of brain tissue is linear from 10% to 50% shear strain, but the time dependent part of the properties in short times shows nonlinear behavior.
Multidisciplinary Design and Optimization for Oscillating Flow Polymerase Chain Reaction Microfluidics Device
Authors: Suwa Tohru , Hadim Hamid , Shi Yong
Published: June 2010

A Polymerase Chain Reaction (PCR) process is almost always required prior to DNA (deoxyribonucleic acid) analysis to create multiple copies of DNA fragments. Using microfluidics technology, the PCR process requires much shorter process time and much less DNA samples than conventional PCR systems. Among existing microfluidics-based techniques, the oscillating flow PCR has advantages including faster analysis time than cavity PCR microfluidics, and smaller contact area between the sample and polymer channel wall compared to flow-through PCR. The smaller contact area reduces DNA adsorption and enhances DNA detection accuracy. In the proposed study, new design features of the oscillating flow PCR concept are evaluated including: (1) PDMS (polydimethylsiloxane) and glass are selected as the microfluidics chip material for realizing a disposable chip, (2) water impingement cooling is applied to effectively isolate the temperature zones, and (3) a copper layer is attached outside of the chip to enhance uniform temperature distribution within the temperature zones. When PDMS is used for PCR microfluidics devices, lower efficiency has been a disadvantage. The efficiency is lowered because the DNA fragments are trapped at the PDMS surface. This trapping can be reduced by minimizing the contact area between the sample and the PDMS surface. When the sample contact area is reduced, which can be achieved by increasing the flow channel cross-sectional area, thermal response is degraded. Optimal channel dimensions are determined by considering the trade-off between thermal response and sample contact area with PDMS channel wall. The resulting thermal response of the sample in the temperature zone is comparable to existing studies, which use silicon as the chip material. A transient FEM heat transfer analysis for the temperature zone is performed for more effective thermal design and optimization.
Some Boundary Value Problems of Elastic Media Containing Rigid Inclusions Under Compressive Mechanical Loads
Authors: Makaryan Vahagn , Sutton Michael , Chlingaryan Gurgen , Hasanyan Davresh , Deng Xiaomin
Published: June 2010

In this work we discuss some 2D boundary-value problems related to an elastic medium containing a thin rigid inclusion with general geometrical shape located in the interface between two separate elastic half-planes and subjected to compressive loading. Assuming perfect bonding between the inclusion and elastic medium, Fourier and Henkel integral transformation techniques are used to obtain the exact solution for the problem. Explicit forms are presented for arbitrary forms of thin inclusions, demonstrating that the tangent shear stress at the end-points of the inclusion has a square-root singularity. It is also shown that the normal stress has a logarithmic singularity when the end-points of the inclusion are approached from the inside of the inclusion and a square-root singularity when the end-points of the inclusion are approached from the outside of the inclusion. For special, extreme cases the solutions for anti-cracks are also presented.
An Effective Drug Delivery Process Using a Molecular Dynamics-Based Cylindrical Particle Model
Authors: Nagireddy Bharat , Lin Y. J.
Published: June 2010

This paper attacks the problem of drug particles modeling in their delivery process for better predictions of solubility and absorption rate. Most of the current models treat the particles to be spheres which may not be accurate. The drug particles formed by grinding and milling are usually not spheres but are irregular or spindles in shape. As such, in this paper modeling of drug particles as cylinders is proposed to rigorously study the impact on its solubility. Along the same line, we also delve into the effect on solubility of changing the aspect ratio of a cylindrical drug particle having a constant mass as well as the diffusion coefficient of the solute which dominates the control on the kinetics of drug release. Molecular Dynamics (MD) simulation is employed to calculate the diffusion coefficient of a given particle using one of Einstein’s fluctuation-dissipation equations that relates transport properties to time. The MD approach makes it possible to deal with parameters that are too difficult to be physically measured in a human body. The solubility analysis of a cylindrical drug particle is done using the Noyes-Whitney equation. The rates of change of radius and of mass are analyzed graphically for drug particles of different weights and sizes. Finally the solubility of a sphere versus that of a cylinder for the same mass is investigated. The data reveal that with the same given mass, the solubility of a spherical particle is less than that of the cylindrical shape. As to the effect of the diffusion coefficient, δ on the solubility of a particle, it is found that the increase in δ for a particle with fixed mass increases its solubility. However, the solubility decreases while δ is increased for a particle with fixed radius.
Biomechanics of Glaucoma: A Parametric Study of Flow in Schlemm’s Canal
Authors: Stolfi Carolyn , Rossmann Jenn S.
Published: June 2010

Glaucoma is one of the most common causes of blindness, affecting approximately seventy million people. Glaucoma is typically caused by high intraocular pressures due to increased outflow resistance, which is primarily concentrated within the trabeculae meshwork and the canal of Schlemm. Debris within trabecular spaces can complicate and limit formation and reabsorption of aqueous humor, leading to elevated interocular pressures. In this study, the biomechanics of Primary Open-Angle Glaucoma are investigated. Computational modeling in physiologically realistic geometries is performed to examine the relationship between intraocular pressure and downstream trabecular and canal resistance. The governing equations for fluid flow are solved, and the influence of such factors as tissue porosity is evaluated. Increased understanding of the biomechanics of glaucoma can improve diagnosis and treatment of this disease.
Dynamics of Human Spine in Jumps Using Lyapunov Exponents
Authors: Banu Eliza A. , Marghitu Dan , Raju P. K.
Published: June 2010

Characterizing and quantifying the local dynamics of the human spine during various athletic exercises is important for therapeutic and physiotherapy reasons or athletic related effects on the human spine. In this study we computed the largest finite-time Lyapunov exponents for the spine of a human subject during an athletic exercise for several volunteers. The kinematic data was collected using the 3D motion capture system (Motion Realty Inc.). Four healthy male subjects were asked to perform two sets of 30 jumps. In order to draw a conclusion about the chaotic nature of the dynamics of the lumbar spine Lyapunov exponent spectrum was calculated for each volunteer which included four Lyapunov exponents, one negative, one very close to zero and two positive. Subjects 1, 2 and 4 registered a significant increase in the largest Lyapunov exponent from the first set jumps to the next. Subject 3 registered no change in the values of Lyapunov exponents. Nonlinear analysis of the human spine demonstrates the chaotic nature of the system. The computation of the largest Lyapunov exponents enables a more precise characterization of the dynamics of the human spine.
A Computational Model of Scapulo-Humeral-Clavicle Complex via Multibody Dynamics
Authors: Duan Shanzhong Shawn , Baumgarten Keith M.
Published: June 2010

The shoulder-upper arm complex has the most mobile joint in the body and is composed of three main bones: the collarbone (clavicle), the shoulder blade (scapula), and the upper arm bone (humerus). The shoulder joint is a non-concentric ball and socket joint. It differs from the hip, a highly stabilized, concentric ball and socket joint, that is constrained mostly by its osseous anatomy. Thus, the shoulder has more flexibility and less inherent stability than the hip because it is mainly stabilized by muscles, tendons, and ligaments. The relative decrease in stability of the shoulder compared to other joints puts the shoulder at increase risk of damage by disease or injury. The constraints added by muscles, tendons, and ligaments make modeling of the shoulder a challenge task. This paper presents a multi rigid body model to describe dynamical properties of the scapulo-humeral-clavicle complex. The bones are represented by rigid bodies, and the soft tissues (tendons, ligaments and muscles) are represented by springs and actuators attached to the rigid bodies. The rigid bodies are connected by ideal kinematic joints and have fixed centers of gravity. Equations of motion of the multi rigid body model are derived via Kane’s methods. Combination of springs and actuators includes independent variables for both motion and constraint forces, the sum of which determine the activation level.
Exploring the Role of Lateral Bending Postures and Asymmetric Loading on Cervical Spine Compression Responses
Authors: Toomey Daniel E. , Mason Matthew J. , Hardy Warren N. , Yang King H. , Kopacz James M. , Van Ee Chris
Published: June 2010

In an effort to expand the understanding of head and neck injury dynamics in rollover type crashes, this investigation explores the influence of lateral bending postures and asymmetric compressive loads on the head and cervical spine. Drop testing of five male cadaver head-neck complexes was conducted with either an initial lateral bending posture onto a horizontal impact surface or with an initial neutral posture onto an obliquely oriented surface resulting in lateral bending. Five specimens were dropped from 0.45 and 0.53 m, with resulting impact speeds ranging from 2.9 to 3.25 m/s. Radiography of the specimens was performed pre- and post-testing to document any fractures. Three of the five specimens sustained compressive cervical vertebral fractures at lower neck loads ranging between 1518 N and 3472 N. Fracture patterns did suggest that the asymmetric postures and loading resulted in asymmetric fracture patterns. Overall compressive neck injury dynamics and tolerances appear similar to previous studies of purely sagittal plane dynamics based on these initial results. This study lays a foundation for quantifying the non-sagittal plane compressive response and tolerance of the cervical spine.
Geometry Governs Mechanics of Cardiovascular Stents
Authors: Douglas Graeham R. , Tan Tho Wei , Bond Tim , Phani A. Srikantha
Published: June 2010

Cardiovascular stents are tubular lattice structures implanted into a stenosed artery to provide adequate lumen support and promote circulation. Commonly encountered complications are stent migration, NeoIntimal Hyperplasia (NIH), and damage to the arterial wall. Central to all these problems is the mechanical response of a stent to forces operating in situ including stent-artery interaction. The influence of geometry or repetitive pattern of the stent upon its mechanical response is the subject of this study. We focus on damage to the arterial wall caused by the stent which can lead to eventual in-stent restenosis. Stent-artery compliance mismatch and longitudinal strain due to Poisson effect are hypothesized as the main contributing factors to restenosis. Finite Element Analysis (FEA) is employed to compare radial compliance and longitudinal strains of different stent geometries. Existing geometrical calculations in the literature [1] are applied to stents of different geometries to compute a non-dimensional NIH index. The main finding is that hybrid lattice stent designs exhibit negligible longitudinal strains (Poisson effect) as the stent expands/contracts during each Cardiac cycle. Wall stresses can be minimized though a careful tailoring of stent geometry.
High-Fidelity and Compact Modeling for Bone Conduction Communication Systems
Authors: Przekwas Andrzej , Zhou Xianlian , Tan X. G. , Chen Z. J. , Reeves Debbie , Wilkerson Patrick , Yang H. Q. , Harrand Vincent , Chancey Valeta Carol
Published: June 2010

Bone conduction (BC) hearing is the process of transmitting sound energy through vibrations of the skull, cerebrospinal fluid (CSF) and brain, which results in an auditory sensation (Stenfelt and Goode 2005). BC communication is attractive for military operations because the transducers are lightweight, inconspicuous, and easily integrated into military headgear. Bone conduction (BC) headsets can present audio when ambient sounds must be either blocked by hearing protection or preserved to maintain situational awareness, and they can provide necessary radio communication in quiet and high noise environments, especially when combined with an appropriate hearing protection system (McBride et al. 2005, Henry and Letowski, 2007). The overall objective of this research is to develop, validate, and deliver anatomy and physics based modeling tools and experimental procedures for analysis and design of cranial bone conduction (BC) communication systems. The modeling tools will be used to optimize the design, attachment, and anatomical location of BC speakers and microphones for best communication clarity in various military environments.
CPG-Controlled Frontal Biped Model for the Evaluation of Pathological Gait
Authors: Hunter Betsy V. , Dhaher Yasin Y.
Published: June 2010

Increased frontal-plane hip movement of the affected leg during the swing phase is a commonly observed gait adaptation in stroke patients. Recent evidence suggests that pathologically-induced torque coupling may contribute to asymmetric gait behaviors observed following stroke. This study proposes to use a CPG-controlled three-dimensional (3D) bipedal model to quantify the effects of abnormal torque coupling on frontal plane gait kinematics. Model dynamics have been evaluated using overground data collection observed under comparable in vivo experimental conditions. The CPG controller has demonstrated ability to provide sustained stable gait over an inclined surface in a simplified model. Preliminary results indicate that the proposed framework is feasible to control a 3D model for investigating the effects of torque coupling on the abnormal frontal plane kinematics of pathological gait.
High-Frequency Ultrasound Tissue Classification of Atherosclerotic Plaques in an APOE-KO Mouse Model Using Spectral Analysis
Authors: Mahmoud Ahmed M. , Teng Bunyen , Mustafa S. Jamal , Mukdadi Osama M.
Published: June 2010

Small animal models have been widely used in cardiovascular research when studying the development and treatment of different diseases. This kind of research has promoted the development of noninvasive techniques to assess cardiac tissue and blood vessels of small animals. Recently, we have developed a high-frequency ultrasound imaging system for small animals, in particular, mouse and rat models. In this work, we aim to elucidate the usefulness of using spectral analysis of the received radiofrequency (RF) ultrasound signals to extract quantitative parameters to assess mechanical properties of cardiac and vascular tissues. A custom system that employs high-frequency single-element ultrasound transducers (30–120 MHz) is used for scanning. Various signal and image processing techniques are applied on the received ultrasound signals to reconstruct high resolution B-mode and spectral images. In vitro imaging of isolated heart and vessels of APOE-KO “knock-out” mouse model with atherosclerosis was performed. Power spectral densities (PSD) of RF signals were evaluated within various regions of interests (ROI) including degassed water, normal cardiac tissue, and cardiac tissue with atheroma. Various parameters were extracted from the power spectrum such as the maximum power (Pmax), the frequency at maximum power (Fpeak), and the variance of power spectrum (Pvar). Results of the preliminary spectral analysis indicated larger values for the Pmax, Fpeak, and Pvar parameters for ROI contains atheroma than other regions. For example using the envelop data, the normalized maximum power (Pmax) value for cardiac tissue with atheroma was 0.0 ± 0.789 (dB), whereas for normal tissues it was about −13.71± 0.267 (dB). These results suggest the use spectral images as a quantitative method when assessing mouse hearts and blood vessels noninvasively.
Aerothermal and Trajectory Analysis of Small Payloads Launched to Low Earth Orbit From an Airborne Platform
Authors: Guillot Martin J.
Published: June 2010

In recent years there has been an ever increasing need to launch small payloads (∼1–100 kg) into low earth orbit (LEO); examples include the defense, telecommunications and other civilian industries. NASA’s stated mission of launching a manned mission to Mars requires many tons of raw materials to be economically launched into LEO and assembled there. Conventional rocket launch from earth is prohibitively expensive for small mass payloads. Estimates range from $7000–$20,000 to launch 1 kg of mass into low earth orbit. Several concepts have been proposed to economically launch small payloads from earth, including light gas guns, electromagnetic launchers and the so called “slingatron” concept. The goal of these concepts is to reduce the cost per kg (to under $1000) to achieve LEO. Each of these concepts involves launching small payloads that traverse the atmosphere and then placed into a circular low earth orbit. As the launch vehicle traverses the dense lower portion of the atmosphere it experiences thermal heating loads that must be absorbed by a thermal protection system (TPS) if the payload is to survive the transit. High launch angles are desirable from the standpoint of minimizing TPS mass. However, for ballistic trajectories, high launch angles require a large propellant mass to achieve a stable circular orbit. This effort performs aerothermal and trajectory analyses on a nominal 10 kg payload launched from 16 km altitude airborne platform into a 200 km circular orbit. The study focuses on two efforts: 1) computing ballistic trajectories of sphere cones with ablation assuming laminar and turbulent flow in order to quantify the total ablation and required propellant mass to circularize the orbit for given launch conditions and 2) study lifting trajectories without ablation by flying axisymmetric sphere-cone projectiles at small angles of attack and asymmetric projectiles (ellipsleds) that turn the velocity vector during atmospheric transit in an effort to reduce the ΔV needed to circularize the orbit. The TPS is assumed to be made of graphite. Total parasitic mass is reported for several launch angles. Even though ablation is not considered for the lifting trajectories, the study allows comparison of relative effectiveness of various lifting trajectories in reducing the ΔV required to circularize the orbit.
3D Numerical Simulation of Contaminant Distribution in an Aircraft
Authors: Tang Yandong , Ratko Daniel , Wang Xiuling
Published: June 2010

Indoor air quality studies are very important as they allow designers to verify that a comfortable and healthy environment is being provided before it is physically constructed. It can be especially important on the closed confines such as a commercial aircraft which may carry a high density of people from various parts of the world. In this study, 3-D contaminant dispersion simulation within an aircraft is conducted by using Finite Volume Methods. Effects of two types of air distribution systems on contaminant dispersion are analyzed. The preliminary results show that the Under Floor Air Distribution system performs better in the overall air quality provided to the passengers of the aircraft than the Ceiling Air Distribution system.
A Low-Cost, Two-Axis, Precision Robot for Automated Fluorescence In-Situ Hybridization Assays
Authors: Kulik M. J. , Shenoda D. S. , Forest C. R.
Published: June 2010

Genetics research often relies on experiments that require repetitive, time-consuming handling of small volumes of liquid (1 mL) and biomass (10–20 μL) such as fluorescence in-situ hybridization (FISH), β-galactosidase staining, immunohisto chemistry, skeletal and tunel assays. Often manual, these experiments are time intensive and error-prone. We report on the design, fabrication, and testing of a low-cost, two-axis, precision robot for FISH assays on whole mice embryos. The robot can complete 20 successive embryo immersions in unique isothermal solutions in minutes for 6 samples. Repeatability of the orthogonal axes is 66 and 214 μm, near the measurement uncertainty limit and sufficient for operation. Accuracy is achieved by systematic error compensation. Low-cost and precision are obtained using design and manufacturing techniques and processes, resulting in a cost of 15% of comparable instruments (e.g., InsituStain, Intavis Bioanalytical Instruments). This design demonstrates a simple, automated platform to perform a typically manual experimental genetics technique.
Use of Pristinamycin for Infections by Gram-Positive Bacteria: Clinical Experience at an Australian Hospital
Authors: Reid A. B. , Daffy J. R. , Stanley P. , Buising K. L.
Published: June 2010

Thirty-six patients were treated with pristinamycin for 46 different microbiological isolates between April 2007 and July 2009. Pathogens included 9 methicillin-resistant
Staphylococcus aureus
isolates, 13 methicillin-resistant coagulase negative staphylococci, and 9 vancomycin-resistant enterococci. Sites of infections included 12 osteomyelitis cases, 10 prosthetic joints, 4 other prostheses, and 1 epidural abscess. Five patients ceased treatment due to side effects. Ten patients were cured of their infections, and 21 patients had infections successfully suppressed.
Adaptive Resistance to the “Last Hope” Antibiotics Polymyxin B and Colistin in
Pseudomonas aeruginosa
Is Mediated by the Novel Two-Component Regulatory System ParR-ParS
Authors: Fernández Lucía , Gooderham W. James , Bains Manjeet , McPhee Joseph B. , Wiegand Irith , Hancock Robert E. W.
Published: June 2010

As multidrug resistance increases alarmingly, polymyxin B and colistin are increasingly being used in the clinic to treat serious
Pseudomonas aeruginosa
infections. In this opportunistic pathogen, subinhibitory levels of polymyxins and certain antimicrobial peptides induce resistance toward higher, otherwise lethal, levels of these antimicrobial agents. It is known that the modification of lipid A of lipopolysaccharide (LPS) is a key component of this adaptive peptide resistance, but to date, the regulatory mechanism underlying peptide regulation in
P. aeruginosa
has remained elusive. The PhoP-PhoQ and PmrA-PmrB two-component systems, which control this modification under low-Mg
2+
conditions, do not appear to play a major role in peptide-mediated adaptive resistance, unlike in
Salmonella
where PhoQ is a peptide sensor. Here we describe the identification and characterization of a novel
P. aeruginosa
two-component regulator affecting

p

olymyxin-

a

daptive

r

esistance, ParR-ParS (PA1799-PA1798). This system was required for activation of the
arnBCADTEF
LPS modification operon in the presence of subinhibitory concentrations of polymyxin, colistin, or the bovine peptide indolicidin, leading to increased resistance to various polycationic antibiotics, including aminoglycosides. This study highlights the complexity of the regulatory network controlling resistance to cationic antibiotics and host peptides in
P. aeruginosa
, which has major relevance in the development and deployment of cationic antimicrobials.
Amodiaquine Resistance in
Plasmodium falciparum
Malaria in Afghanistan Is Associated with the
pfcrt
SVMNT Allele at Codons 72 to 76
Authors: Beshir Khalid , Sutherland Colin J. , Merinopoulos Ioannis , Durrani Naeem , Leslie Toby , Rowland Mark , Hallett Rachel L.
Published: June 2010

Mutations in the
Plasmodium falciparum
genes
pfcrt
and
pfmdr1
are selected by amodiaquine treatment in Africa. To examine the importance of these mutations in amodiaquine-treated Asian parasites, we determined pre- and posttreatment genotypes for amodiaquine treatment failures from a clinical trial in Afghanistan. The
pfcrt
codon 72 to 76 haplotype SVMNT was present in all samples tested, both before and after treatment. Amodiaquine did not clearly select for any
pfmdr1
genotype, but a novel mutation,
pfmdr1
N86F, was detected in four samples. We provide
in vivo
data to support the
in vitro
correlation between
pfcrt
SVMNT and increased resistance to the metabolite of amodiaquine.
Structure-Activity Analysis of Vinylogous Urea Inhibitors of Human Immunodeficiency Virus-Encoded Ribonuclease H
Authors: Chung Suhman , Wendeler Michaela , Rausch Jason W. , Beilhartz Greg , Gotte Matthias , O'Keefe Barry R. , Bermingham Alun , Beutler John A. , Liu Shixin , Zhuang Xiaowei , Le Grice Stuart F. J.
Published: June 2010

Vinylogous ureas 2-amino-5,6,7,8-tetrahydro-4
H
-cyclohepta[
b
]thiophene-3-carboxamide and
N
-[3-(aminocarbonyl)-4,5-dimethyl-2-thienyl]-2-furancarboxamide (compounds 1 and 2, respectively) were recently identified to be modestly potent inhibitors of the RNase H activity of HIV-1 and HIV-2 reverse transcriptase (RT). Both compounds shared a 3-CONH
2
-substituted thiophene ring but were otherwise structurally unrelated, which prevented a precise definition of the pharmacophore. We have therefore examined a larger series of vinylogous ureas carrying amide, amine, and cycloalkane modifications of the thiophene ring of compound 1. While cycloheptane- and cyclohexane-substituted derivatives retained potency, cyclopentane and cyclooctane substitutions eliminated activity. In the presence of a cycloheptane ring, modifying the 2-NH
2
or 3-CONH
2
functions decreased the potency. With respect to compound 2, vinylogous ureas whose dimethylthiophene ring contained modifications of the 2-NH
2
and 3-CONH
2
functions were investigated. 2-NH
2
-modified analogs displayed potency equivalent to or enhanced over that of compound 2, the most active of which, compound 16, reflected intramolecular cyclization of the 2-NH
2
and 3-CONH
2
groups. Molecular modeling was used to define an inhibitor binding site in the p51 thumb subdomain, suggesting that an interaction with the catalytically conserved His539 of the p66 RNase H domain could underlie inhibition of RNase H activity. Collectively, our data indicate that multiple functional groups of vinylogous ureas contribute to their potencies as RNase H inhibitors. Finally, single-molecule spectroscopy indicates that vinylogous ureas have the property of altering the reverse transcriptase orientation on a model RNA-DNA hybrid mimicking initiation plus-strand DNA synthesis.
Microarray and Molecular Analyses of the Azole Resistance Mechanism in
Candida glabrata
Oropharyngeal Isolates
Authors: Tsai Huei-Fung , Sammons Lindsay R. , Zhang Xiaozhen , Suffis Sara D. , Su Qin , Myers Timothy G. , Marr Kieren A. , Bennett John E.
Published: June 2010

DNA microarrays were used to analyze
Candida glabrata
oropharyngeal isolates from seven hematopoietic stem cell transplant recipients whose isolates developed azole resistance while the recipients received fluconazole prophylaxis. Transcriptional profiling of the paired isolates revealed 19 genes upregulated in the majority of resistant isolates compared to their paired susceptible isolates. All seven resistant isolates had greater than 2-fold upregulation of
C. glabrata PDR1
(Cg
PDR1
), a master transcriptional regulator of the pleiotropic drug resistance (PDR) network, and all seven resistant isolates showed upregulation of known Cg
PDR1
target genes. The altered transcriptome can be explained in part by the observation that all seven resistant isolates had acquired a single nonsynonymous mutation in their Cg
PDR1
open reading frame. Four mutations occurred in the regulatory domain (L280P, L344S, G348A, and S391L) and one in the activation domain (G943S), while two mutations (N764I and R772I) occurred in an undefined region. Association of azole resistance and the Cg
PDR1
mutations was investigated in the same genetic background by introducing the Cg
PDR1
sequences from one sensitive isolate and five resistant isolates into a laboratory azole-hypersusceptible strain (Cg
pdr1
strain) via integrative transformation. The Cg
pdr1
strain was restored to wild-type fluconazole susceptibility when transformed with Cg
PDR1
from the susceptible isolate but became resistant when transformed with Cg
PDR1
from the resistant isolates. However, despite the identical genetic backgrounds, upregulation of Cg
PDR1
and Cg
PDR1
target genes varied between the five transformants, independent of the domain locations in which the mutations occurred. In summary, gain-of-function mutations in Cg
PDR1
contributed to the clinical azole resistance, but different mutations had various degrees of impact on the Cg
PDR1
target genes.
Nationwide Investigation of Extended-Spectrum β-Lactamases, Metallo-β-Lactamases, and Extended-Spectrum Oxacillinases Produced by Ceftazidime-Resistant
Pseudomonas aeruginosa
Strains in France
Authors: Hocquet Didier , Plésiat Patrick , Dehecq Barbara , Mariotte Pierre , Talon Daniel , Bertrand Xavier
Published: June 2010

A nationwide study aimed to identify the extended-spectrum β-lactamases (ESBLs), metallo-β-lactamases (MBLs), and extended-spectrum oxacillinases (ES-OXAs) in a French collection of 140 clinical
Pseudomonas aeruginosa
isolates highly resistant to ceftazidime. Six ESBLs (PER-1,
n
= 3; SHV-2a,
n
= 2; VEB-1a,
n
= 1), four MBLs (VIM-2,
n
= 3; IMP-18,
n
= 1), and five ES-OXAs (OXA-19,
n
= 4; OXA-28,
n
= 1) were identified in 13 isolates (9.3% of the collection). The prevalence of these enzymes is still low in French clinical
P. aeruginosa
isolates but deserves to be closely monitored.
Voriconazole Pharmacokinetics and Safety in Immunocompromised Children Compared to Adult Patients
Authors: Michael Claudia , Bierbach Uta , Frenzel Katrin , Lange Thoralf , Basara Nadezda , Niederwieser Dietger , Mauz-Körholz Christine , Preiss Rainer
Published: June 2010

The aim of this study was to investigate the pharmacokinetics and safety of voriconazole after intravenous (i.v.) administration in immunocompromised children (2 to 11 years old) and adults (20 to 60 years old) who required treatment for the prevention or therapy of systemic fungal infections. Nine pediatric patients were treated with a dose of 7 mg/kg i.v. every 12 h for a period of 10 days. Three children and 12 adults received two loading doses of 6 mg/kg i.v. every 12 h, followed by a maintenance dose of 5 mg/kg (children) or 4 mg/kg (adults) twice a day during the entire study period. Trough voriconazole levels in blood over 10 days of therapy and regular voriconazole levels in blood for up to 12 h postdose on day 3 were examined. Wide intra- and interindividual variations in plasma voriconazole levels were noted in each dose group and were most pronounced in the children receiving the 7-mg/kg dose. Five (56%) of them frequently had trough voriconazole levels in plasma below 1 μg/ml or above 6 μg/ml. The recommended dose of 7 mg/kg i.v. in children provides exposure (area under the concentration-time curve) comparable to that observed in adults receiving 4 mg/kg i.v. The children had significantly higher
C
max
values; other pharmacokinetic parameters were not significantly different from those of adults. Voriconazole exhibits nonlinear pharmacokinetics in the majority of children. Voriconazole therapy was safe and well tolerated in pediatric and adult patients. The European Medicines Agency-approved i.v. dose of 7 mg/kg can be recommended for children aged 2 to <12 years.
Image-Based High-Throughput Drug Screening Targeting the Intracellular Stage of
Trypanosoma cruzi
, the Agent of Chagas' Disease
Authors: Engel Juan C. , Ang Kenny K. H. , Chen Steven , Arkin Michelle R. , McKerrow James H. , Doyle Patricia S.
Published: June 2010

Chagas' disease, caused by infection with the parasite
Trypanosoma cruzi
, is the major cause of heart failure in Latin America. Classic clinical manifestations result from the infection of heart muscle cells leading to progressive cardiomyopathy. To ameliorate disease, chemotherapy must eradicate the parasite. Current drugs are ineffective and toxic, and new therapy is a critical need. To expedite drug screening for this neglected disease, we have developed and validated a cell-based, high-throughput assay that can be used with a variety of untransfected
T. cruzi
isolates and host cells and that simultaneously measures efficacy against the intracellular amastigote stage and toxicity to host cells.
T. cruzi
-infected muscle cells were incubated in 96-well plates with test compounds. Assay plates were automatically imaged and analyzed based on size differences between the DAPI (4′,6-diamidino-2-phenylindole)-stained host cell nuclei and parasite kinetoplasts. A reduction in the ratio of
T. cruzi
per host cell provided a quantitative measure of parasite growth inhibition, while a decrease in count of the host nuclei indicated compound toxicity. The assay was used to screen a library of clinically approved drugs and identified 55 compounds with activity against
T. cruzi
. The flexible assay design allows the use of various parasite strains, including clinical isolates with different biological characteristics (e.g., tissue tropism and drug sensitivity), and a broad range of host cells and may even be adapted to screen for inhibitors against other intracellular pathogens. This high-throughput assay will have an important impact in antiparasitic drug discovery.
Immunomodulatory Effects of Serotype B Glucuronoxylomannan from
Cryptococcus gattii
Correlate with Polysaccharide Diameter
Authors: Fonseca Fernanda L. , Nohara Lilian L. , Cordero Radames J. B. , Frases Susana , Casadevall Arturo , Almeida Igor C. , Nimrichter Leonardo , Rodrigues Marcio L.
Published: June 2010

Glucuronoxylomannan (GXM), the major capsular component in the
Cryptococcus
complex, interacts with the immune system in multiple ways, which include the activation of Toll-like receptors (TLRs) and the modulation of nitric oxide (NO) production by phagocytes. In this study, we analyzed several structural parameters of GXM samples from
Cryptococcus neoformans
(serotypes A and D) and
Cryptococcus gattii
(serotypes B and C) and correlated them with the production of NO by phagocytes and the activation of TLRs. GXM fractions were differentially recognized by TLR2/TLR1 (TLR2/1) and TLR2/6 heterodimers expressed on TLR-transfected HEK293A cells. Higher NF-κB luciferase reporter activity induced by GXM was observed in cells expressing TLR2/1 than in cells transfected with TLR2/6 constructs. A serotype B GXM from
C. gattii
was the most effective polysaccharide fraction activating the TLR-mediated response. This serotype B polysaccharide, which was also highly efficient at eliciting the production of NO by macrophages, was similar to the other GXM samples in monosaccharide composition, zeta potential, and electrophoretic mobility. However, immunofluorescence with four different monoclonal antibodies and dynamic light-scattering analysis revealed that the serotype B GXM showed particularities in serological reactivity and had the smallest effective diameter among the GXM samples analyzed in this study. Fractionation of additional serotype B GXMs, followed by exposure of these fractions to macrophages, revealed a correlation between NO production and reduced effective diameters. Our results demonstrate a great functional diversity in GXM samples from different isolates and establish their abilities to differentially activate cellular responses. We propose that serological properties as well as physical chemical parameters, such as the diameter of polysaccharide molecules, may potentially influence the inflammatory response against
Cryptococcus
spp. and may contribute to the differences in granulomatous inflammation between cryptococcal species.
E3 Ubiquitin Ligase Activity and Targeting of BAT3 by MultipleLegionella pneumophilaTranslocated Substrates
Authors: Ensminger Alexander W. , Isberg Ralph R.
Published: June 2010

The intracellular bacterial pathogenLegionella pneumophilamodulates a number of host processes during intracellular growth, including the eukaryotic ubiquitination machinery, which dictates the stability, activity, and/or localization of a large number of proteins. A number ofL. pneumophilaproteins contain eukaryotic-like motifs typically associated with ubiquitination. Central among these is a family of five F-box-domain-containing proteins ofLegionella pneumophila. Each of these five proteins is translocated to the host cytosol by the Dot/Icm type IV protein translocation system during infection. We show that three of these proteins, LegU1, LegAU13, and LicA, interact with components of the host ubiquitination machineryin vivo. In addition, LegU1 and LegAU13 are integrated into functional Skp-Cullin-F-box (SCF) complexes that confer E3 ubiquitin ligase activity. LegU1 specifically interacts with and can direct the ubiquitination of the host chaperone protein BAT3. In a screen for additionalL. pneumophilaproteins that associate with LegU1 in mammalian cells, we identified the bacterial protein Lpg2160. We demonstrate that Lpg2160 also associates with BAT3 independently of LegU1. We show that Lpg2160 is a translocated substrate of the Dot/Icm system and contains a C-terminal translocation signal. We propose a model in which LegU1 and Lpg2160 may function redundantly or in concert to modulate BAT3 activity during the course of infection.
Optical and voltammetric studies on tetraphenylethylene salts
Authors: Mohammad Mahboob , Mahmood Tariq , Jamil Tahir , Khan Ather Yaseen
Published: June 2010

Optical spectra of tetraphenylethylene salts with lithium, potassium, rubidium, and tetrabutylammonium as counter ions, were recorded in tetrahydrofuran. Also the optical spectrum of sodium salt of tetraphenylethylene was recorded at very high dilution. On computer resolution of the optical spectra it was found that the observed bands were composed of three peaks, located around 385, 490, and 585 nm. Depending upon the counter ions and the environments these occur in different height ratios. These findings pointed to ion pairs formation and helped in elucidating their structure. The cyclic voltammetric studies on tetraphenylethylene helped in the evaluation of dissociation constant of T2-, M+ and the disproportination constant 2T.- ⇄ T2- + T. The former was found to be 1.2 . 10-5 mol 1-1, the latter 0.42 in dimethylformamide.
Synthesis and electrochemical behaviour of a new class of macrocyclic Li+-selective diamides
Authors: Hruška Zdeněk , Petránek Jaromír
Published: June 2010

A series of macrocyclic N-substituted oligomethylenediamides of 4,4,5,5-tetramethyl-3,6-dioxaoctanedioic acid, 5,5-dimethyl-3,7-dioxanonanedioic and trans-1,2-cyclohexanedioxydiacetic acid were prepared. Selectivity of these compounds with respect to the structure of the molecule was investigated electrochemically in polymeric (PVC) membranes of ion-selective electrodes.
Halogen substituted adamantanones. Structural studies with lanthanide shift reagents
Authors: Hájek Milan , Horská Alena , Trška Petr , Vodička Luděk , Raber Douglas J. , Podlogar Brent L. , Peters Joop A.
Published: June 2010

Europium shift reagents (both FOD and DPM) have been used to study a series of halogen substituted adamantanones by 1H NMR spectroscopy. Reliable correlations of molecular structure with the lanthanide induced shifts can be obtained only if certain precautions are taken. The source of atomic coordinates used to predict induced shifts with the pseudocontact equation is critical; coordinates by X-ray crystallography can be unreliable for this purpose, while those from empirical force field calculations yield satisfactory results. Determination of the europium-oxygen bond length by optimization of the agreement factor is shown to be inadequate. Even with a restricted Eu-O bond length, calculation of the best lanthanide position is shown to be dependent on the step size by which the lanthanide is moved in the optimization procedure. All of the compounds studied were found to give complexes with approximately linear C-O-Eu arrays, and the distortions from linearity are consistent with steric effects.
Reaction of 1-acetyladamantane with thionyl chloride
Authors: Burkhard Jiří , Janků Josef , Vodička Luděk
Published: June 2010

1-Acetyladamantane (I) reacts with thionyl chloride in the presence of pyridine to give 1-(1’-chloroetenyl)adamantane (IV). In the absence of pyridine, the same reaction affords a more complex mixture containing 1-adamantanecarboxylic acid (VI), its ethyl ester (VII), and 1-chloroacetyladamantane (V).
Photolysis of 19β,28-epoxy-18α-oleanan-2β-ol nitrites: Functionalization of 10β- and 8β-methyl groups
Authors: Sejbal Jan , Klinot Jiří , Vystrčil Alois
Published: June 2010

On photolysis, 19β,28-epoxy-18α-oleanan-2β-ol nitrite (I) undergoes functionalization of the 10β-methyl group (formation of oxime II) as well as of the rather distant 8β-methyl group (leading to oxime III). No products of attack at the 4β-methyl group have been found. The 2β-nitrolyloxy-3β-acetoxy derivative XX reacts similarly whereas its 2β-nitrosyloxy-3α-acetoxy isomer XIII is photolyzed to give diol XI. Oximes II and III were converted into two series of nitriles (a and b) with various functional groups on the ring A. Structure of all compounds was derived from their spectral data. Double radical transfer has been proposed for functionalization of the 8β-methyl group (position 26).
Potentional anxiolytics and hypnotics: 1-(Alkanesulfonamidoalkyl)-6-aryl-8-halogeno-s-triazolo[4,3-a]-1,4-benzodiazepines and related compounds
Authors: Vejdělek Zdeněk , Metyš Jan , Holubek Jiří , Buděšínský Miloš , Svátek Emil , Matoušová Oluše , Protiva Miroslav
Published: June 2010

7-Chloro-5-phenyl-1,3-dihydro-1,4-benzodiazepin-2-thione and its 5-(2-chlorophenyl) and 7-bromo-5-(2-chlorophenyl)analogues were reacted with N-(methanesulfonyl)- and N-(ethanesulfonyl)glycine and –alanine hydrazides (X-XIII) in boiling butanol to give the title compounds Iabc - IVabc. The alanine-derived substances IIIabc and IVabc were characterized by 1H NMR spectra as mixtures of two diastereoisomers. Similar reactions of 5-methylimidazole-4-carboxylic acid hydrazide (XIV) gave the s-triazolo[4,3-a]-1,4-benzodiazepines Vabc together with their ring-opened precursors XVI and XVII. The compounds prepared showed the activity profile of the anxiolytic and hypnotic 4H-s-triazolo[4,3-a]-1,4-benzodiazepine derivatives.