bluuebunny/crossref_metadata_2025 huggingface dataset:
Amphetamine and methamphetamine reduce striatal dopamine transporter function without concurrent dopamine transporter relocalization
Authors: German Christopher L. , Hanson Glen R. , Fleckenstein Annette E.
Published: July 2012

J. Neurochem. (2012) 123, 288–297.

Amphetamine (AMPH) and methamphetamine (METH) alter dopamine transporter (DAT) function. In vitro heterologous cell line and synaptosome studies demonstrate AMPH‐induced DAT internalization, implicating relocalization in reduced DAT uptake following drug exposure. However, few studies have evaluated DAT localization following in vivo AMPH/METH administration. To determine DAT subcellular localization following drug administration, a centrifugation technique was developed to isolate striatal synaptosomal membrane and vesicle fractions. DAT was distributed between the synaptosomal membrane (60%) and endosomal vesicles (40%), and in vitro application of the protein kinase C activator phorbol 12‐myristate 13‐acetate to striatal synaptosomes caused DAT internalization into the vesicle fractions. In contrast, neither single nor repeated in vivo AMPH and/or METH administrations altered DAT localization 5, 15, 30, or 60 min post‐treatment, despite reduced DAT uptake. Importantly, repeated METH injections uniformly decreased total DAT immunoreactivity within all fractions 7 days post‐treatment. These findings suggest that factors other than internalization can contribute to the observed acute and persistent DAT dysfunction and dopaminergic deficits following in vivo AMPH or METH administration.
Adverse reactions from essential oil‐containing natural flea products exempted from Environmental Protection Agency regulations in dogs and cats
Authors: Genovese Allison G. , McLean Mary Kay , Khan Safdar A.
Published: July 2012

Objective

To describe adverse effects in dogs and cats exposed to Environmental Protection Agency exempted plant‐derived flea preventatives containing mixtures of essential oils.

Design

Retrospective study from 2006 to 2008.

Setting

Records of dog and cat cases were reviewed from the American Society for the Prevention of Cruelty to Animals, Animal Poison Control Center database.

Animals

Thirty‐nine cats and 9 dogs with history of exposure to natural flea preventatives.

Measurements and Main Results

The following information was retrieved from each incident: number of animals, species involved, frequency, types, onset time, duration of clinical signs, exposure appropriateness, final outcome, and treatment information. Ninety‐two percent of animals (n = 44) showed presence of one or more adverse effects. The frequency of adverse effects in dogs (n = 8; 89%) and cats (n = 36; 92%) was similar. Onset time of adverse effects in 39 of 44 animals occurred within 24 hours. The duration of signs in 24 animals ranged from 30 minutes to 149 hours. The products were used as per label in 77% animals (n = 37). Of 28 animals with known outcome, 50% (n = 14) recovered with bathing alone while others received intravenous fluids, muscle relaxants, and anticonvulsive medications. Death (1 cat; n = 1/28; 4%) or euthanasia (1 cat and 1 dog; n = 2/28; 7%) was reported in 3 animals.

Conclusion

Dogs and cats can experience significant adverse effects when exposed to plant‐derived flea preventatives even when used according to label directions. The number of reports of exposure in cats was higher than dogs, but the frequency of reported adverse effects was similar between the 2 species. Agitation and hypersalivation were common in cats, whereas lethargy and vomiting were common in dogs.
The effect of using an impedance threshold device on hemodynamic parameters during cardiopulmonary resuscitation in dogs
Authors: Buckley Gareth J. , Shih Andre , Garcia‐Pereira Fernando L. , Bandt Carsten
Published: July 2012

Objective

To investigate the hemodynamic effects following the use of an impedance threshold device (ITD) in a canine model of cardiopulmonary arrest.

Design

Experimental, randomized crossover study.

Setting

Cardiovascular research laboratory at a university veterinary center.

Animals

Eight purpose bred beagle dogs.

Interventions

Dogs were anesthetized and instrumented for the measurement of right atrial pressure , systolic blood pressure, mean blood pressure, and diastolic arterial blood pressure, end‐tidalCO2, and carotid blood flow (CBF).CBFwas determined via ultrasonic flow probe placed around the carotid artery. Animals were taking part in an unassociated terminal study and following subsequent euthanasia with pentobarbital, standardized cardiopulmonary resuscitation (CPR) was performed with an impedance threshold device attached (ITD‐CPRgroup) and without (S‐CPRgroup). Order of treatment was randomized.

Measurements and Main Results

ITDgroup had increasedCBF, coronary perfusion pressure, and a decrease in right atrial diastolic pressure. No differences in end‐tidalCO2, diastolic arterial blood pressure, mean blood pressure, or systolic blood pressure were seen. Return of spontaneous circulation was not observed in any of the animals

Conclusions

Use of theITDresulted in favorable changes in hemodynamic parameters in dogs undergoingCPR. TheITDmay be a useful adjunct duringCPRin dogs and warrants clinical evaluation.
Wound Healing Society (WHS) venous ulcer treatment guidelines: What's new in five years?
Authors: Tang Jennifer C. , Marston William A. , Kirsner Robert S.
Published: July 2012

Since the establishment of the guidelines for the treatment of venous ulcers by the Wound Healing Society in 2006, there has been an abundance of new literature, both in accord and discord with the guidelines. The goal of this update is to highlight new findings since the publication of these guidelines to assist practitioner and patient in appropriate health care decisions, as well as to drive future research endeavors.
Personal Identification of Cold Case Remains Through Combined Contribution from Anthropological, mtDNA, and Bomb‐Pulse Dating Analyses
Authors: Speller Camilla F. , Spalding Kirsty L. , Buchholz Bruce A. , Hildebrand Dean , Moore Jason , Mathewes Rolf , Skinner Mark F. , Yang Dongya Y.
Published: July 2012

In 1968, a child's cranium was recovered from the banks of a northern Canadian river and held in a trust until the “cold case” was reopened in 2005. The cranium underwent reanalysis at the Centre for Forensic Research, Simon Fraser University, using recently developed anthropological analysis, “bomb‐pulse” radiocarbon analysis, and forensic DNA techniques. Craniometrics, skeletal ossification, and dental formation indicated an age‐at‐death of 4.4 ± 1 year. Radiocarbon analysis of enamel from two teeth indicated a year of birth between 1958 and 1962. Forensic DNA analysis indicated the child was a male, and the obtained mitochondrial profile matched a living maternal relative to the presumed missing child. These multidisciplinary analyses resulted in a legal identification 41 years after the discovery of the remains, highlighting the enormous potential of combining radiocarbon analysis with anthropological and mtDNA analyses in producing confident personal identifications for forensic cold cases dating to within the last 60 years.
What digital games and literacy have in common: a heuristic for understanding pupils' gaming literacy
Authors: Apperley Thomas , Walsh Christopher
Published: July 2012

This article argues that digital games and school‐based literacy practices have much more in common than is reported in the research literature. We describe the role digital game paratexts – ancillary print and multimodal texts about digital games – can play in connecting pupils’ gaming literacy practices to ‘traditional’ school‐based literacies still needed for academic success. By including the reading, writing and design of digital game paratexts in the literacy curriculum, teachers can actively and legitimately include digital games in their literacy instruction. To help teachers understand pupils’ gaming literacy practices in relation to other forms of literacy practices, we present a heuristic for understanding gaming (HUG) literacy. We argue our heuristic can be used for effective teacher professional development because it assists teachers in identifying the elements of gameplay that would be appropriate for the demands of the literacy curriculum. The heuristic traces gaming literacy across the quadrants of actions, designs, situations and systems to provide teachers and practitioners with a knowledge of gameplay and a metalanguage for talking about digital games. We argue this knowledge will assist them in capitalising on pupils’ existing gaming literacy by connecting their out‐of‐school gaming literacy practices to the literacy and English curriculum.
HuPho: the human phosphatase portal
Authors: Liberti Susanna , Sacco Francesca , Calderone Alberto , Perfetto Livia , Iannuccelli Marta , Panni Simona , Santonico Elena , Palma Anita , Nardozza Aurelio P. , Castagnoli Luisa , Cesareni Gianni
Published: July 2012

Phosphatases and kinases contribute to the regulation of protein phosphorylation homeostasis in the cell. Phosphorylation is a key post‐translational modification underlying the regulation of many cellular processes. Thus, a comprehensive picture of phosphatase function and the identification of their target substrates would aid a systematic approach to a mechanistic description of cell signalling. Here we present a website designed to facilitate the retrieval of information about human protein phosphatases. To this end we developed a search engine to recover and integrate information annotated in several publicly available web resources. In addition we present a text‐mining‐assisted annotation effort aimed at extracting phosphatase related data reported in the scientific literature. The HuPho (human phosphatases) website can be accessed at http://hupho.uniroma2.it.
A distinct de novo expression of Nav1.5 sodium channels in human atrial fibroblasts differentiated into myofibroblasts
Authors: Chatelier Aurélien , Mercier Aurélie , Tremblier Boris , Thériault Olivier , Moubarak Majed , Benamer Najate , Corbi Pierre , Bois Patrick , Chahine Mohamed , Faivre Jean François
Published: July 2012

Key point

Fibroblasts play a major role in heart physiology. In pathological conditions, they can lead to cardiac fibrosis when they differentiate into myofibroblasts.

This differentiated status is associated with changes in expression profile leading to neo‐expression of proteins such as ionic channels.

The present study investigates electrophysiological changes associated with fibroblast differentiation focusing on voltage‐gated sodium channels in human atrial fibroblasts and myofibroblasts.

We show that human atrial fibroblast differentiation in myofibroblasts is associated with de novo expression of voltage gated sodium current. Multiple arguments support that this current is predominantly supported by the Nav1.5 α‐subunit which may generate a persistent sodium entry into myofibroblasts.

Our data revealed that Nav1.5 α‐subunit expression is not restricted to cardiac myocytes within the atrium. Since fibrosis is one of the fundamental mechanisms implicated in atrial fibrillation, it is of great interest to investigate how this channel could influence myofibroblasts function.

Abstract  Fibroblasts play a major role in heart physiology. They are at the origin of the extracellular matrix renewal and production of various paracrine and autocrine factors. In pathological conditions, fibroblasts proliferate, migrate and differentiate into myofibroblasts leading to cardiac fibrosis. This differentiated status is associated with changes in expression profile leading to neo‐expression of proteins such as ionic channels. The present study investigates further electrophysiological changes associated with fibroblast differentiation focusing on the activity of voltage‐gated sodium channels in human atrial fibroblasts and myofibroblasts. Using the patch clamp technique we show that human atrial myofibroblasts display a fast inward voltage gated sodium current with a density of 13.28 ± 2.88 pA pF−1 whereas no current was detectable in non‐differentiated fibroblasts. Quantitative RT‐PCR reveals a large amount of transcripts encoding the Nav1.5 α‐subunit with a fourfold increased expression level in myofibroblasts when compared to fibroblasts. Accordingly, half of the current was blocked by 1 μm of tetrodotoxin and immunocytochemistry experiments reveal the presence of Nav1.5 proteins. Overall, this current exhibits similar biophysical characteristics to sodium currents found in cardiac myocytes except for the window current that is enlarged for potentials between −100 and −20 mV. Since fibrosis is one of the fundamental mechanisms implicated in atrial fibrillation, it is of great interest to investigate how this current could influence myofibroblast properties. Moreover, since several Nav1.5 mutations are related to cardiac pathologies, this study offers a new avenue on the fibroblasts involvement of these mutations.
Heterotypic connexin50/connexin50 mutant gap junction channels reveal interactions between two hemichannels during transjunctional voltage‐dependent gating
Authors: Xin Li , Sun Yiguo , Bai Donglin
Published: July 2012

Key points

Gap junctions (GJs) composed of two hemichannels are intercellular channels that provide a direct pathway for the exchange of ions and signalling molecules between adjoining cells.

GJ conductance is regulated by transjunctional voltage (Vj, the voltage difference between the interiors of adjoining cells), known as Vj‐dependent gating.

The GJs made up of two identical hemichannels show mirror symmetrical Vj‐dependent gating to positive and negative Vj polarities; GJs formed by two non‐identical hemichannels show novel Vj‐gating properties that cannot be simply ascribed to the predicted functional properties of the component hemichannels.

We provide experimental evidence at both macroscopic and single channel levels that two hemichannels interact with each other in a complex manner for the Vj‐dependent gating phenotype of a GJ.

The results help reveal the molecular mechanisms of the gating properties of GJs formed by two non‐identical hemichannels.

Abstract  To investigate transjunctional voltage (Vj)‐dependent gating mechanisms of connexin50 (Cx50) gap junction (GJ) channels and to elucidate the relative contribution of each hemichannel of a heterotypic GJ channel to Vj‐dependent gating, we performed dual voltage‐clamp recordings on heterotypic GJ channels formed by Cx50 and a mutant, Cx50N9R or a chimera, Cx50‐Cx36N. Our results provide evidence that the two component hemichannels interact with each other during Vj‐dependent gating. Cx50/Cx50N9R heterotypic GJ channels exhibited asymmetrical Vj‐dependent gating which cannot be ascribed to the function of an individual hemichannel for a certain polarity of voltage; instead it can only be ascribed to the combined effects of both hemichannels. Single GJ channel open dwell‐time analyses showed that homotypic Cx50 channels adopted short‐lived and long‐lived open states. Heterotypic combinations of Cx50/Cx50N9R gave rise to shorter mean dwell‐times when Cx50‐expressing cells received relatively positive Vj, and longer mean dwell‐times when positive Vj was applied at the Cx50N9R side. In contrast, Cx50/Cx50‐Cx36N heterotypic channels showed asymmetrical Vj‐dependent gating, which appears to be caused by enhanced and reduced Vj‐gating sensitivity of Cx50‐Cx36N and Cx50 hemichannels, respectively. Unitary conductance of the main open state of both types of heterotypic GJ channel cannot be simply predicted by assuming a Vj redistribution across the two hemichannels arranged in series in heterotypic GJ channels. Our data also reveal reasons for the invisibility of fast Vj‐gating transitions from open to substate in homotypic Cx50N9R and Cx50‐Cx36N channels.
A universally conserved residue in the SUR1 subunit of the KATP channel is essential for translating nucleotide binding at SUR1 into channel opening
Authors: de Wet Heidi , Shimomura Kenju , Aittoniemi Jussi , Ahmad Nawaz , Lafond Mathilde , Sansom Mark S.P. , Ashcroft Frances M.
Published: July 2012

Key points

The sulphonylurea receptor (SUR1) subunit of the ATP‐sensitive potassium (KATP) channel is a member of the ATP‐binding cassette (ABC) protein family. Binding of MgADP to nucleotide‐binding domain 2 (NBD2) is critical for channel activation.

We identified a residue in the SUR1 subunit of the KATP channel that is essential for translating nucleotide binding to SUR1 into activation of the channel pore.

The ability of ATP to block KATP channel activity by binding to the Kir6.2 subunit of the channel was also altered by the mutation. This effect was dependent on the integrity of the NBDs of SUR1. This suggests SUR1 also modulates nucleotide inhibition of the channel at Kir6.2.

G1401 in SUR1 is one of 23 residues which are conserved throughout all ABC transporter proteins suggesting that it may have a universal role in coupling substrate binding to protein function.

Abstract  The sulphonylurea receptor (SUR1) subunit of the ATP‐sensitive potassium (KATP) channel is a member of the ATP‐binding cassette (ABC) protein family. Binding of MgADP to nucleotide‐binding domain 2 (NBD2) is critical for channel activation. We identified a residue in NBD2 (G1401) that is fully conserved among ABC proteins and whose functional importance is unknown. Homology modelling places G1401 on the outer surface of the protein, distant from the nucleotide‐binding site. The ATPase activity of purified SUR1‐NBD2‐G1410R (bound to maltose‐binding protein) was slightly inhibited when compared to the wild‐type protein, but its inhibition by MgADP was unchanged, indicating that MgADP binding is not altered. However, MgADP activation of channel activity was abolished. This implies that the G1401R mutation impairs the mechanism by which MgADP binding to NBD2 is translated into opening of the KATP channel pore. The location of G1401 would be consistent with interaction of this residue with the pore‐forming Kir6.2 subunit. Channel activity in the presence of MgATP reflects the balance between the stimulatory (at SUR1) and inhibitory (at Kir6.2) effects of nucleotides. Mutant channels were 2.5‐fold less sensitive to MgATP inhibition and not activated by MgATP. This suggests that ATP block of the channel is reduced by the SUR1 mutation. Interestingly, this effect was dependent on the functional integrity of the NBDs. These results therefore suggest that SUR1 modulates both nucleotide inhibition and activation of the KATP channel.
ATP‐sensitive potassium channels mediate the thermosensory response of orexin neurons
Authors: Parsons Matthew P. , Belanger‐Willoughby Natasha , Linehan Victoria , Hirasawa Michiru
Published: July 2012

Key points

High body temperature accompanies sleepiness, and loss of appetite and motivation.

Suppression of orexin neurons has been suggested to be involved in such behavioural responses.

In this study we determined that orexin neurons are inhibited by an increase in temperature itself.

The effect of warming is mediated by a novel temperature sensing mechanism involving ATP‐sensitive potassium (KATP) channels.

As KATP channels are also sensitive to energy balance, our results suggest that orexin neurons play an important role in regulating energy balance by inducing sickness behaviour when body temperature is elevated.

Abstract  High body temperatures are generally associated with somnolence, lethargy, hypophagia and anhedonia. Orexin neurons have been suggested to play a role in such sickness behaviours due to their known functions in appetite, behavioural and autonomic activation. Furthermore, the activity of orexin neurons is inhibited by lipopolysaccharide that induces fever. However, the cellular mechanism(s) underlying this suppression of orexin neurons was unknown. We used patch‐clamp recordings in acute rat brain slices to demonstrate that orexin neurons, including those projecting to the wake‐promoting locus coeruleus, are inhibited by increasing the ambient temperature by a 2–4°C increment between 26 and 40°C. This effect was not mediated by conventional thermosensing mechanisms but instead involved the activation of ATP‐sensitive potassium (KATP) channels. Since KATP channels can also sense energy substrate levels and cellular metabolism, our results suggest that orexin neurons can integrate the state of energy balance and body temperature, and adjust their output accordingly. Thus, the thermosensitivity of orexin neurons may be an important part of maintaining energy homeostasis during hyperthermia and fever.
Basolateral chloride loading by the anion exchanger type 2: role in fluid secretion by the human airway epithelial cell line Calu‐3
Authors: Huang Junwei , Shan Jiajie , Kim Dusik , Liao Jie , Evagelidis Alexandra , Alper Seth L. , Hanrahan John W.
Published: July 2012

Key points

The companion paper provided evidence for basolateral anion exchange during cAMP stimulation of chloride and fluid secretion by Calu‐3 monolayers; however, the molecular basis of this transport was not identified.

To test the role of AE2, an anion exchanger expressed at the basolateral membrane of Calu‐3 and many other epithelial cells, we used lentivirus‐mediated RNA interference to generate a stable Calu‐3 AE2 knock‐down cell line and characterized its fluid and anion transport properties.

AE2 knock‐down suppressed fluid secretion and increased the fraction of cAMP‐stimulated anion secretion that was sensitive to bumetanide inhibition.

Basolateral Cl−/HCO3− exchange was nearly abolished in AE2 knock‐down cells.

We conclude that AE2 is active during forskolin‐stimulated fluid secretion and mediates chloride uptake and bicarbonate recycling at the basolateral membrane.

Abstract  Anion exchanger type 2 (AE2 or SLC4A2) is an electroneutral Cl−/HCO3− exchanger expressed at the basolateral membrane of many epithelia. It is thought to participate in fluid secretion by airway epithelia. However, the role of AE2 in fluid secretion remains uncertain, due to the lack of specific pharmacological inhibitors, and because it is electrically silent and therefore does not contribute directly to short‐circuit current (Isc). We have studied the role of AE2 in Cl− and fluid secretion by the airway epithelial cell line Calu‐3. After confirming expression of its mRNA and protein, a knock‐down cell line called AE2‐KD was generated by lentivirus‐mediated RNA interference in which AE2 mRNA and protein levels were reduced ≥90%. Suppressing AE2 increased the expression of the cystic fibrosis transmembrane conductance regulator (CFTR) by ∼70% without affecting the levels of NKCC1 (Na+–K+–2Cl− cotransporter) or NBCe1 (Na+–nHCO3− cotransporter). cAMP agonists stimulated fluid secretion by parental Calu‐3 and scrambled shRNA cells >6.5‐fold. In AE2‐KD cells this response was reduced by ∼70%, and the secreted fluid exhibited elevated pH and [HCO3−] as compared with the control lines. Unstimulated equivalent short‐circuit current (Ieq) was elevated in AE2‐KD cells, but the incremental response to forskolin was unaffected. The modest bumetanide‐induced reductions in both Ieq and fluid secretion were more pronounced in AE2‐KD cells. Basolateral Cl−/HCO3− exchange measured by basolateral pH‐stat in cells with permeabilized apical membranes was abolished in AE2‐KD monolayers, and the intracellular alkalinization resulting from basolateral Cl− removal was reduced by ∼80% in AE2‐KD cells. These results identify AE2 as a major pathway for basolateral Cl− loading during cAMP‐stimulated secretion of Cl− and fluid by Calu‐3 cells, and help explain the large bumetanide‐insensitive component of fluid secretion reported previously in airway submucosal glands and some other epithelia.
New Correlations for Thermal Resistances of Vertical Single U-Tube Ground Heat Exchanger
Authors: Liao Quan , Zhou Chao , Cui Wenzhi , Jen T. C.
Published: July 2012

A new 2D numerical model of a single U-tube ground heat exchanger is proposed and a four-thermal-resistance model is adopted to evaluate the effective pipe-to-borehole, pipe-to-pipe, and borehole-to-borehole thermal resistances. The influence of temperature distributions on both borehole surface and outer diameter of two pipes to these thermal resistances has been thoroughly studied. The best-fit correlations of effective pipe-to-borehole, pipe-to-pipe, and borehole-to-borehole thermal resistances are proposed and compared with the available equations in the literature. It is found that the present correlations of thermal resistances for ground heat exchanger are more accurate than those of available formulas. Furthermore, based on these obtained thermal resistance correlations, an analytical model is proposed to evaluate the heat transfer performance of the ground heat changer.
Study of the fatigue wear behaviors of a tungsten carbide diamond-like carbon coating on 316L stainless steel
Authors: Chen Ying , Nie Xueyuan
Published: July 2012

A tungsten carbide (WC) diamond-like carbon (DLC) hard coating has higher lubricity and resilience than other types of nitride-, carbide- and oxide-based ceramic coatings. To evaluate the anti-wear properties of the DLC coating under extremely high impact-sliding loading conditions, a newly developed method, called the cycled inclined impact-sliding test, is introduced and utilized to study fatigue wear behaviors of the DLC coating under a combination of an impact force, Fi, and pressing force, Fp (Fi/Fp = 200 N/400 N), for up to 10 000 cycles. A 10 mm steel (AISI 52100) bearing ball is used as the impacting and sliding object. Due to the low coefficient of friction of the WC DLC coating against the steel counterface, which creates a reduced tangential force on the coating surface and less tensile stress within the coating, the WC DLC coating exhibits high endurance when impacted in dry air conditions. A scanning electron microscopy study showed that different types of fatigue wear cracks appeared and were distributed in the different areas (head and tail parts) of the impact and sliding tracks. Most of those cracks penetrated the coating and caused sawtooth-like deformation in the substrate. Material transfer from the counterface ball was not detected by energy-dispersive x-ray spectroscopy.
Consequences of Noncompliance for Therapy Efficacy and Emergence of Resistance in Murine Tuberculosis Caused by the Beijing Genotype of Mycobacterium tuberculosis
Authors: de Steenwinkel Jurriaan E. M. , ten Kate Marian T. , de Knegt Gerjo J. , Verbrugh Henri A. , Aarnoutse Rob E. , Boeree Martin J. , den Bakker Michael A. , van Soolingen Dick , Bakker-Woudenberg Irma A. J. M.
Published: July 2012

Despite great effort by health organizations worldwide in fighting tuberculosis (TB), morbidity and mortality are not declining as expected. One of the reasons is related to the evolutionary development of
Mycobacterium tuberculosis
, in particular the Beijing genotype strains. In a previous study, we showed the association between the Beijing genotype and an increased mutation frequency for rifampin resistance. In this study, we use a Beijing genotype strain and an East-African/Indian genotype strain to investigate with our mouse TB model whether the higher mutation frequency observed in a Beijing genotype strain is associated with treatment failure particularly during noncompliance therapy. Both genotype strains showed high virulence in comparison to that of
M. tuberculosis
strain H37Rv, resulting in a highly progressive infection with a rapid lethal outcome in untreated mice. Compliance treatment was effective without relapse of TB irrespective of the infecting strain, showing similar decreases in the mycobacterial load in infected organs and similar histopathological changes. Noncompliance treatment, simulated by a reduced duration and dosing frequency, resulted in a relapse of infection. Relapse rates were correlated with the level of noncompliance and were identical for Beijing infection and East African/Indian infection. However, only in Beijing-infected mice, isoniazid-resistant mutants were selected at the highest level of noncompliance. This is in line with the substantial selection of isoniazid-resistant mutants
in vitro
in a wide isoniazid concentration window observed for the Beijing strain and not for the EAI strain. These results suggest that genotype diversity of
M. tuberculosis
may be involved in emergence of resistance and indicates that genotype-tailor-made treatment should be investigated.
Altering the Proclivity towards Daptomycin Resistance in Methicillin-Resistant Staphylococcus aureus Using Combinations with Other Antibiotics
Authors: Berti Andrew D. , Wergin Justine E. , Girdaukas Gary G. , Hetzel Scott J. , Sakoulas George , Rose Warren E.
Published: July 2012

Daptomycin (DAP) is increasingly used as a part of combination therapy, particularly in complex methicillin-resistant
Staphylococcus aureus
(MRSA) infections. While multiple studies have reported the potential for synergy between DAP and adjunctive anti-infectives, few have examined the influence of adjunctive therapy on the emergence of DAP resistance. This study examined eight adjunctive antimicrobial combinations with DAP
in vitro
and the emergence of DAP resistance over time (up to 4 weeks) using clinical isolates of DAP-susceptible MRSA (MIC, 0.5 μg/ml) in which DAP resistance subsequently developed during patient therapy (MIC, 3 μg/ml). In addition to DAP susceptibility testing, selected strains were examined for phenotypic changes associated with DAP resistance, including changes to cell wall thickness (CWT) and cell membrane alterations. The addition of either oxacillin or clarithromycin in medium containing DAP significantly inhibited the development of DAP resistance through the entirety of the 4-week exposure (10- to 32-fold MIC reduction from that of DAP alone). Combinations with rifampin or fosfomycin were effective in delaying the emergence of DAP resistance through the end of week one only (week one MIC, 0.5 μg/ml; week four MIC, 24 μg/ml). Cell wall thickening was observed for all antibiotic combinations regardless of their effect on the DAP MIC (14 to 70% increase in CWT), while changes in cell membrane fluidity were variable and treatment dependent. DAP showed reduced activity against strains with DAP MICs of 1 to 12 μg/ml, but cell membrane integrity was still disrupted at concentrations achieved with doses greater than 10 mg/kg of body weight. The emergence of DAP resistance in MRSA is strongly influenced by the presence of subinhibitory concentrations of adjunctive antimicrobials. These data suggest that combining DAP with oxacillin or clarithromycin may delay the development of DAP resistance in cases requiring prolonged antibiotic therapy.
Effect of Ginkgo Biloba on the Pharmacokinetics of Raltegravir in Healthy Volunteers
Authors: Blonk Maren , Colbers Angela , Poirters Anne , Schouwenberg Bas , Burger David
Published: July 2012

Medicinal herbs may cause clinically relevant drug interactions with antiretroviral agents. Ginkgo biloba extract is a popular herbal product among HIV-infected patients because of its positive effects on cognitive function. Raltegravir, an HIV integrase inhibitor, is increasingly being used as part of combined antiretroviral therapy. Clinical data on the potential inhibitory or inductive effect of ginkgo biloba on the pharmacokinetics of raltegravir were lacking, and concomitant use was not recommended. We studied the effect of ginkgo biloba extract on the pharmacokinetics of raltegravir in an open-label, randomized, two-period, crossover phase I trial in 18 healthy volunteers. Subjects were randomly assigned to a regimen of 120 mg of ginkgo biloba twice daily for 15 days plus a single dose of raltegravir (400 mg) on day 15, a washout period, and 400 mg of raltegravir on day 36 or the test and reference treatments in reverse order. Pharmacokinetic sampling of raltegravir was performed up to 12 h after intake on an empty stomach. All subjects (9 male) completed the trial, and no serious adverse events were reported. Geometric mean ratios (90% confidence intervals) of the area under the plasma concentration-time curve from dosing to infinity (AUC
0-∞
) and the maximum plasma concentration (
C
max
) of raltegravir with ginkgo biloba versus raltegravir alone were 1.21 (0.93 to 1.58) and 1.44 (1.03 to 2.02). Ginkgo biloba did not reduce raltegravir exposure. The potential increase in the
C
max
of raltegravir is probably of minor importance, given the large intersubject variability of raltegravir pharmacokinetics and its reported safety profile.
Comparison of Plasma, Epithelial Lining Fluid, and Alveolar Macrophage Concentrations of Solithromycin (CEM-101) in Healthy Adult Subjects
Authors: Rodvold Keith A. , Gotfried Mark H. , Still J. Gordon , Clark Kay , Fernandes Prabhavathi
Published: July 2012

The steady-state concentrations of solithromycin in plasma were compared with concomitant concentrations in epithelial lining fluid (ELF) and alveolar macrophages (AM) obtained from intrapulmonary samples during bronchoscopy and bronchoalveolar lavage (BAL) in 30 healthy adult subjects. Subjects received oral solithromycin at 400 mg once daily for five consecutive days. Bronchoscopy and BAL were carried out once in each subject at either 3, 6, 9, 12, or 24 h after the last administered dose of solithromycin. Drug concentrations in plasma, ELF, and AM were assayed by a high-performance liquid chromatography-tandem mass spectrometry method. Solithromycin was concentrated extensively in ELF (range of mean [± standard deviation] concentrations, 1.02 ± 0.83 to 7.58 ± 6.69 mg/liter) and AM (25.9 ± 20.3 to 101.7 ± 52.6 mg/liter) in comparison with simultaneous plasma concentrations (0.086 ± 0.070 to 0.730 ± 0.692 mg/liter). The values for the area under the concentration-time curve from 0 to 24 h (AUC0–24values) based on mean and median ELF concentrations were 80.3 and 63.2 mg · h/liter, respectively. The ratio of ELF to plasma concentrations based on the mean and median AUC0–24values were 10.3 and 10.0, respectively. The AUC0–24values based on mean and median concentrations in AM were 1,498 and 1,282 mg · h/L, respectively. The ratio of AM to plasma concentrations based on the mean and median AUC0–24values were 193 and 202, respectively. Once-daily oral dosing of solithromycin at 400 mg produced steady-state concentrations that were significantly (P< 0.05) higher in ELF (2.4 to 28.6 times) and AM (44 to 515 times) than simultaneous plasma concentrations throughout the 24-h period after 5 days of solithromycin administration.
The Combination of Colistin and Doripenem Is Synergistic against Klebsiella pneumoniae at Multiple Inocula and Suppresses Colistin Resistance in an
In Vitro
Pharmacokinetic/Pharmacodynamic Model
Authors: Deris Zakuan Z. , Yu Heidi H. , Davis Kathryn , Soon Rachel L. , Jacob Jovan , Ku Caron K. , Poudyal Anima , Bergen Phillip J. , Tsuji Brian T. , Bulitta Jurgen B. , Forrest Alan , Paterson David L. , Velkov Tony , Li Jian , Nation Roger L.
Published: July 2012

Multidrug-resistant (MDR)
Klebsiella pneumoniae
may require combination therapy. We systematically investigated bacterial killing with colistin and doripenem mono- and combination therapy against MDR
K. pneumoniae
and emergence of colistin resistance. A one-compartment
in vitro
pharmacokinetic/pharmacodynamic model was employed over a 72-h period with two inocula (∼10
6
and ∼10
8
CFU/ml); a colistin-heteroresistant reference strain (ATCC 13883) and three clinical isolates (colistin-susceptible FADDI-KP032 [doripenem resistant], colistin-heteroresistant FADDI-KP033, and colistin-resistant FADDI-KP035) were included. Four combinations utilizing clinically achievable concentrations were investigated. Microbiological responses were examined by determining log changes and population analysis profiles (for emergence of colistin resistance) over 72 h. Against colistin-susceptible and -heteroresistant isolates, combinations of colistin (constant concentration regimens of 0.5 or 2 mg/liter) plus doripenem (steady-state peak concentration [
C
max
] of 2.5 or 25 mg/liter over 8 h; half-life, 1.5 h) generally resulted in substantial improvements in bacterial killing at both inocula. Combinations were additive or synergistic against ATCC 13883, FADDI-KP032, and FADDI-KP033 in 9, 9, and 14 of 16 cases (4 combinations at 6, 24, 48, and 72 h) at the 10
6
-CFU/ml inoculum and 14, 11, and 12 of 16 cases at the 10
8
-CFU/ml inoculum, respectively. Combinations at the highest dosage regimens resulted in undetectable bacterial counts at 72 h in 5 of 8 cases (4 isolates at 2 inocula). Emergence of colistin-resistant subpopulations in colistin-susceptible and -heteroresistant isolates was virtually eliminated with combination therapy. Against the colistin-resistant isolate, colistin at 2 mg/liter plus doripenem (
C
max
, 25 mg/liter) at the low inoculum improved bacterial killing. This investigation provides important information for optimization of colistin-doripenem combinations.
Vaccination with SesC Decreases Staphylococcus epidermidis Biofilm Formation
Authors: Shahrooei Mohammad , Hira Vishal , Khodaparast Laleh , Khodaparast Ladan , Stijlemans Benoit , Kucharíková Soňa , Burghout Peter , Hermans Peter W. M. , Van Eldere Johan
Published: July 2012

The increased use of medical implants has resulted in a concomitant rise in device-related infections. The majority of these infections are caused by
Staphylococcus epidermidis
biofilms. Immunoprophylaxis and immunotherapy targeting
in vivo
-expressed, biofilm-associated, bacterial cell surface-exposed proteins are promising new approaches to prevent and treat biofilm-related infections, respectively. Using an
in silico
procedure, we identified 64 proteins that are predicted to be

S
.
e
pidermidis

s
urface exposed (Ses), of which 36 were annotated as (conserved) hypothetical. Of these 36 proteins, 5 proteins—3 LPXTG motif-containing proteins (SesL, SesB, and SesC) and 2 of the largest ABC transporters (SesK and SesM)—were selected for evaluation as vaccine candidates. This choice was based on protein size, number of antigenic determinants, or the established role in
S. epidermidis
biofilm formation of the protein family to which the candidate protein belongs. Anti-SesC antibodies exhibited the greatest inhibitory effect on
S. epidermidis
biofilm formation
in vitro
and on colonization and infection in a mouse jugular vein catheter infection model that includes biofilms and organ infections. Active vaccination with a recombinant truncated SesC inhibited
S. epidermidis
biofilm formation in a rat model of subcutaneous foreign body infection. Antibodies to SesC were shown to be opsonic by an
in vitro
opsonophagocytosis assay. We conclude that SesC is a promising target for antibody mediated strategies against
S. epidermidis
biofilm formation.
A Ferritin Mutant of Mycobacterium tuberculosis Is Highly Susceptible to Killing by Antibiotics and Is Unable To Establish a Chronic Infection in Mice
Authors: Pandey Ruchi , Rodriguez G. Marcela
Published: July 2012

Iron is an essential, elusive, and potentially toxic nutrient for most pathogens, including
Mycobacterium tuberculosis
. Due to the poor solubility of ferric iron under aerobic conditions, free iron is not found in the host.
M. tuberculosis
requires specialized iron acquisition systems to replicate and cause disease. It also depends on a strict control of iron metabolism and intracellular iron levels to prevent iron-mediated toxicity. Under conditions of iron sufficiency,
M. tuberculosis
represses iron acquisition and induces iron storage, suggesting an important role for iron storage proteins in iron homeostasis.
M. tuberculosis
synthesizes two iron storage proteins, a ferritin (BfrB) and a bacterioferritin (BfrA). The individual contributions of these proteins to the adaptive response of
M. tuberculosis
to changes in iron availability are not clear. By generating individual knockout strains of
bfrA
and
bfrB
, the contribution of each one of these proteins to the maintenance of iron homeostasis was determined. The effect of altered iron homeostasis, resulting from impaired iron storage, on the resistance of
M. tuberculosis
to
in vitro
and
in vivo
stresses was examined. The results show that ferritin is required to maintain iron homeostasis, whereas bacterioferritin seems to be dispensable for this function.
M. tuberculosis
lacking ferritin suffers from iron-mediated toxicity, is unable to persist in mice, and, most importantly, is highly susceptible to killing by antibiotics, showing that endogenous oxidative stress can enhance the antibiotic killing of this important pathogen. These results are relevant for the design of new therapeutic strategies against
M. tuberculosis
.
Helicobacter pylori Requires TlpD-Driven Chemotaxis To Proliferate in the Antrum
Authors: Rolig Annah S. , Shanks James , Carter J. Elliot , Ottemann Karen M.
Published: July 2012

Different disease outcomes of
Helicobacter pylori
infection correlate with distinct inflammation patterns. These different inflammatory distributions may be initiated by differences in bacterial localization. One
H. pylori
property known to affect murine stomach localization is chemotaxis, the ability to move in response to chemical cues. In this report, we used nonchemotactic mutants (Che
−
) to analyze whether chemotaxis is required for initial colonization of particular stomach regions or for subsequent growth therein. We found that
H. pylori
behaves differently in the corpus, antrum, and corpus-antrum transition zone subregions of the stomach. This outcome suggests that these regions contain unique chemotactic signals. In the corpus,
H. pylori
utilizes chemotaxis for initial localization but not for subsequent growth. In contrast, in the antrum and the corpus-antrum transition zone, chemotaxis does not help initial colonization but does promote subsequent proliferation. To determine which chemoreceptor is responsible for the corpus-antrum phenotypes, we infected mice with strains lacking each chemoreceptor. Strains lacking TlpA, TlpB, or TlpC displayed only modest deviations from the wild-type phenotype, while strains lacking TlpD resembled the Che
−
mutant in their antral colonization defect and fared even worse than the Che
−
mutant in the corpus. Additional analysis showed that inflammation is worse in the antrum than in the corpus in both wild-type and Che
−
mutant infections. These results suggest that chemotaxis, specifically, that controlled by TlpD, is necessary for
H. pylori
to survive or grow in the environment of increased inflammation in the antrum.
Abscess Formation and Alpha-Hemolysin Induced Toxicity in a Mouse Model of Staphylococcus aureus Peritoneal Infection
Authors: Rauch Sabine , DeDent Andrea C. , Kim Hwan Keun , Bubeck Wardenburg Juliane , Missiakas Dominique M. , Schneewind Olaf
Published: July 2012

Staphylococcus aureus
is a frequent cause of skin infection and sepsis in humans. Preclinical vaccine studies with
S. aureus
have used a mouse model with intraperitoneal challenge and survival determination as a measure for efficacy. To appreciate the selection of protective antigens in this model, we sought to characterize the pathological attributes of
S. aureus
infection in the peritoneal cavity. Testing C57BL/6J and BALB/c mice, >10
9
CFU of
S. aureus
Newman were needed to produce a lethal outcome in 90% of animals infected via intraperitoneal injection. Both necropsy and histopathology revealed the presence of intraperitoneal abscesses in the vicinity of inoculation sites. Abscesses were comprised of fibrin as well as collagen deposits and immune cells with staphylococci replicating at the center of these lesions. Animals that succumbed to challenge harbored staphylococci in abscess lesions and in blood. The establishment of lethal infections, but not the development of intraperitoneal abscesses, was dependent on
S. aureus
expression of alpha-hemolysin (Hla). Active immunization with nontoxigenic Hla
H35L
or passive immunization with neutralizing monoclonal antibodies protected mice against early lethal events associated with intraperitoneal
S. aureus
infection but did not affect the establishment of abscess lesions. These results characterize a mouse model for the study of intraperitoneal abscess formation by
S. aureus
, a disease that occurs frequently in humans undergoing continuous ambulatory peritoneal dialysis for end-stage renal disease.
Exacerbation of AIH in a patient with an AIH/systemic sclerosis overlap syndrome and pulmonary arterial hypertension treated with the endothelin-1 receptor antagonist sitaxentan
Authors: Klein Reinhild , Hintz Eva , Staehler Gerd
Published: July 2012

Increase of liver enzymes during therapy with endothelin receptor antagonist (ERA) because of pulmonary arterial hypertension (PAH) has been observed quite frequently the cause of which is unknown. Here we describe a female patient who suffered from autoimmune hepatitis (AIH) type I [positive for antinuclear (ANA) and antiactin antibodies] who developed systemic sclerosis (SSc) with PAH. AIH was treated with corticosteroids and azathioprine, and PAH with the ERA sitaxentan. Reactivation of AIH was observed in the course of therapy with sitaxentan as shown by an increase of liver enzymes, immunoglobulin G globulins, the reappearance of antinuclear and antiactin antibodies and the induction of a further AIH marker antibody reacting with the soluble liver/liver pancreas antigen. Therapy with ERA for pulmonary hypertension may increase the risk for development or exacerbation of AIH.
A longitudinal qualitative exploration of healthcare and informal support needs among survivors of critical illness: the RELINQUISH protocol
Authors: Ramsay Pam , Huby Guro , Rattray Janice , Salisbury Lisa G , Walsh Timothy Simon , Kean Susanne
Published: July 2012

Introduction and background

Survival following critical illness is associated with a significant burden of physical, emotional and psychosocial morbidity. Recovery can be protracted and incomplete, with important and sustained effects upon everyday life, including family life, social participation and return to work. In stark contrast with other critically ill patient groups (eg, those following cardiothoracic surgery), there are comparatively few interventional studies of rehabilitation among the general intensive care unit patient population. This paper outlines the protocol for a sub study of the RECOVER study: a randomised controlled trial evaluating a complex intervention of enhanced ward-based rehabilitation for patients following discharge from intensive care.

Methods and analysis

The RELINQUISH study is a nested longitudinal, qualitative study of family support and perceived healthcare needs among RECOVER participants at key stages of the recovery process and at up to 1 year following hospital discharge. Its central premise is that recovery is a dynamic process wherein patients' needs evolve over time. RELINQUISH is novel in that we will incorporate two parallel strategies into our data analysis: (1) a pragmatic health services-oriented approach, using an a priori analytical construct, the ‘Timing it Right’ framework and (2) a constructivist grounded theory approach which allows the emergence of new themes and theoretical understandings from the data. We will subsequently use Qualitative Health Needs Assessment methodology to inform the development of timely and responsive healthcare interventions throughout the recovery process.

Ethics and dissemination

The protocol has been approved by the Lothian Research Ethics Committee (protocol number HSRU011). The study has been added to the UK Clinical Research Network Database (study ID. 9986). The authors will disseminate the findings in peer reviewed publications and to relevant critical care stakeholder groups.