bluuebunny/crossref_metadata_2025 huggingface dataset:
Temporal components of cholinergic terminal to dopaminergic terminal transmission in dorsal striatum slices of mice
Authors: Wang Li , Zhang Xiaoyu , Xu Huadong , Zhou Li , Jiao Ruiying , Liu Wei , Zhu Feipeng , Kang Xinjiang , Liu Bin , Teng Sasa , Wu Qihui , Li Mingli , Dou Haiqiang , Zuo Panli , Wang Changhe , Wang Shirong , Zhou Zhuan
Published: June 2014

Key points

The timing of synaptic transmission is critical to synaptic plasticity in the striatum.

However, the timing of striatal dopamine (DA) release induced by cholinergic interneurons (ChIs) in the striatum is unclear.

In this study, we focused on the temporal components of DA release and replenishment triggered by different pathways.

We show that stimulation of ChIs induces DA release with a total delay of 20.6 ms, including 2.8 ms for action potential firing of ChIs, 7.0 ms for cholinergic transmission between acetylcholine terminals and DA terminals, and 10.8 ms for downstream DA release. The delay of DA release via this ChI pathway is 1.9 times that via the nigrostriatal pathways.

We describe the time course of recovery of DA release via the two pathways and that of vesicle replenishment in DA terminals.

Our work provides an example of unravelling the temporal building blocks during fundamental synaptic terminal–terminal transmission.

Striatal dopamine (DA) is critically involved in major brain functions such as motor control and deficits such as Parkinson's disease. DA is released following stimulation by two pathways: the nigrostriatal pathway and the cholinergic interneuron (ChI) pathway. The timing of synaptic transmission is critical in striatal circuits, because millisecond latency changes can reverse synaptic plasticity from long‐term potentiation to long‐term depression in a DA‐dependent manner. Here, we determined the temporal components of ChI‐driven DA release in striatal slices from optogenetic ChAT‐ChR2‐EYFP mice. After a light stimulus at room temperature, ChIs fired an action potential with a delay of 2.8 ms. The subsequent DA release mediated by nicotinic acetylcholine (ACh) receptors had a total latency of 17.8 ms, comprising 7.0 ms for cholinergic transmission and 10.8 ms for the downstream terminal DA release. Similar latencies of DA release were also found in striatal slices from wild‐type mice. The latency of ChI‐driven DA release was regulated by inhibiting the presynaptic vesicular ACh release. Moreover, we describe the time course of recovery of DA release via the two pathways and that of vesicle replenishment in DA terminals. Our work provides an example of unravelling the temporal building blocks during fundamental synaptic terminal–terminal transmission in motor regulation.
Airway turbulence and changes in upper airway hydraulic diameter can be estimated from the intensity of high frequency inspiratory sounds in sleeping adults
Authors: Rembold Christopher M. , Suratt Paul M.
Published: June 2014

Key points

Adults and children with obstructive sleep disordered breathing make loud high frequency inspiratory sounds (HFIS, defined as inspiratory sounds >2 kHz) while sleeping.

In an in vitro model, the intensity of high frequency sounds was found to be an estimate of airflow turbulence as reflected by the Reynold's number (Re).

Hydraulic diameter (D) of the in vitro model could be calculated using Re determined by sound intensity, the pressure gradient, the Swamee–Jain formula and the Darcy formula.

In four adult humans, D was calculated from measured HFIS intensity and the pressure gradient across the upper airway (estimated with oesophageal pressure, Pes).

At apnoea termination when the airway opens, we observed (1) an increase in HFIS intensity suggesting an increase in turbulence (higher Re), and (2) a larger calculated D.

This method allows dynamic estimation of changes in relative upper airway D in sleeping humans with narrowed upper airways.

Obstructive sleep disordered breathing can cause death and significant morbidity in adults and children. We previously found that children with smaller upper airways (measured by magnetic resonance imaging while awake) generated loud high frequency inspiratory sounds (HFIS, defined as inspiratory sounds > 2 kHz) while they slept. The purpose of this study was (1) to determine what characteristics of airflow predicted HFIS intensity, and (b) to determine if we could calculate changes in hydraulic diameter (D) in both an in vitro model and in the upper airways of sleeping humans. In an in vitro model, high frequency sound intensity was an estimate of airflow turbulence as reflected by the Reynold's number (Re). D of the in vitro model was calculated using Re, the pressure gradient, Swamee–Jain formula and Darcy formula. D was proportional to but smaller than the actual diameters (r2 = 0.94). In humans, we measured HFIS intensity and the pressure gradient across the upper airway (estimated with oesophageal pressure, Pes) during polysomnography in four adult volunteers and applied the same formulae to calculate D. At apnoea termination when the airway opens, we observed (1) an increase in HFIS intensity suggesting an increase in turbulence (higher Re), and (2) a larger calculated D. This method allows dynamic estimation of changes in relative upper airway hydraulic diameter (D) in sleeping humans with narrowed upper airways.
Central command dysfunction in rats with heart failure is mediated by brain oxidative stress and normalized by exercise training
Authors: Koba Satoshi , Hisatome Ichiro , Watanabe Tatsuo
Published: June 2014

Key points

In heart failure, sympathoexcitation elicited by central command, a parallel activation of the motor and autonomic neural circuits in the brain, is exaggerated.

Mechanisms underlying central command dysfunction in heart failure were unexplored, and effects of exercise training on central command dysfunction in heart failure were not determined.

Data presented here suggest that oxidative stress in the medulla in heart failure mediates central command dysfunction, and that exercise training in heart failure is capable of normalizing central command dysfunction through its antioxidant effects in the medulla.

The present study contributes to our understanding of brain mechanisms underlying abnormal autonomic adjustments to exercise in heart failure.

Sympathoexcitation elicited by central command, a parallel activation of the motor and autonomic neural circuits in the brain, has been shown to become exaggerated in chronic heart failure (CHF). The present study tested the hypotheses that oxidative stress in the medulla in CHF plays a role in exaggerating central command‐elicited sympathoexcitation, and that exercise training in CHF suppresses central command‐elicited sympathoexcitation through its antioxidant effects in the medulla. In decerebrate rats, central command was activated by electrically stimulating the mesencephalic locomotor region (MLR) after neuromuscular blockade. The MLR stimulation at a current intensity greater than locomotion threshold in rats with CHF after myocardial infarction (MI) evoked larger (P < 0.05) increases in renal sympathetic nerve activity and arterial pressure than in sham‐operated healthy rats (Sham) and rats with CHF that had completed longterm (8–12 weeks) exercise training (MI + TR). In the Sham and MI + TR rats, bilateral microinjection of a superoxide dismutase (SOD) mimetic Tempol into the rostral ventrolateral medulla (RVLM) had no effects on MLR stimulation‐elicited responses. By contrast, in MI rats, Tempol treatment significantly reduced MLR stimulation‐elicited responses. In a subset of MI rats, treatment with Tiron, another SOD mimetic, within the RVLM also reduced responses. Superoxide generation in the RVLM, as evaluated by dihydroethidium staining, was enhanced in MI rats compared with that in Sham and MI + TR rats. Collectively, these results support the study hypotheses. We suggest that oxidative stress in the medulla in CHF mediates central command dysfunction, and that exercise training in CHF is capable of normalizing central command dysfunction through its antioxidant effects in the medulla.
Regulation of hypoxia‐inducible factor‐α isoforms and redox state by carotid body neural activity in rats
Authors: Peng Ying‐Jie , Yuan Guoxiang , Khan Shakil , Nanduri Jayasri , Makarenko Vladislav V. , Reddy Vaddi Damodara , Vasavda Chirag , Kumar Ganesh K. , Semenza Gregg L. , Prabhakar Nanduri R.
Published: June 2014

Key points

Rats exposed to chronic intermittent hypoxia (CIH) exhibited imbalanced expression of hypoxia‐inducible factor (HIF)‐α isoforms and oxidative stress in brainstem regions associated with the carotid body (CB) chemoreflex, and in the adrenal medulla, an end organ of the sympathetic nervous system.

Selective ablation of the CB abolished the effects of CIH on HIF‐α isoform expression and oxidative stress.

In the adrenal medulla, chemoreflex‐mediated sympathetic activation regulates HIF‐α isoform expression via muscarinic acetylcholine receptor‐mediated Ca2+ influx and the resultant activation of the mammalian target of rapamycin pathway and calpain proteases.

Thus, CB neural activity regulates HIF‐α isoform expressions and redox state in the central and peripheral nervous system associated with the chemoreflex pathway under the setting of CIH.

Previous studies reported that chronic intermittent hypoxia (CIH) results in an imbalanced expression of hypoxia‐inducible factor‐α (HIF‐α) isoforms and oxidative stress in rodents, which may be due either to the direct effect of CIH or indirectly via hitherto uncharacterized mechanism(s). As neural activity is a potent regulator of gene transcription, we hypothesized that carotid body (CB) neural activity contributes to CIH‐induced HIF‐α isoform expression and oxidative stress in the chemoreflex pathway. Experiments were performed on adult rats exposed to CIH for 10 days. Rats exposed to CIH exhibited: increased HIF‐1α and decreased HIF‐2α expression; increased NADPH oxidase 2 and decreased superoxide dismutase 2 expression; and oxidative stress in the nucleus tractus solitarius and rostral ventrolateral medulla as well as in the adrenal medulla (AM), a major end organ of the sympathetic nervous system. Selective ablation of the CB abolished these effects. In the AM, sympathetic activation by the CB chemoreflex mediates CIH‐induced HIF‐α isoform imbalance via muscarinic acetylcholine receptor‐mediated Ca2+ influx, and the resultant activation of mammalian target of rapamycin pathway and calpain proteases. Rats exposed to CIH presented with hypertension, elevated sympathetic activity and increased circulating catecholamines. Selective ablation of either the CB (afferent pathway) or sympathetic innervation to the AM (efferent pathway) abolished these effects. These observations uncover CB neural activity‐dependent regulation of HIF‐α isoforms and the redox state by CIH in the central and peripheral nervous systems associated with the chemoreflex.
Regulation of serum phosphate
Authors: Lederer Eleanor
Published: June 2014

The regulation of serum phosphate, an acknowledged risk factor for chronic kidney disease and cardiovascular mortality, is poorly understood. The discovery of fibroblast growth factor 23 (FGF23) as a key regulator of renal phosphate handling and activation of vitamin D has revolutionized our comprehension of phosphate homeostasis. Through as yet undetermined mechanisms, circulating and dietary phosphate appear to have a direct effect on FGF23 release by bone cells that, in turn, causes renal phosphate excretion and decreases intestinal phosphate absorption through a decrease in vitamin D production. Thus, the two major phosphaturic hormones, PTH and FGF23, have opposing effects on vitamin D production, placing vitamin D at the nexus of phosphate homeostasis. While our understanding of phosphate homeostasis has advanced, the factors determining regulation of serum phosphate level remain enigmatic. Diet, time of day, season, gender, age and genetics have all been identified as significant contributors to serum phosphate level. The effects of these factors on serum phosphate have major implications for what is understood as ‘normal’ and for studies of phosphate homeostasis and metabolism. Moreover, other hormonal mediators such as dopamine, insulin‐like growth factor, and angiotensin II also affect renal handling of phosphate. How the major hormone effects on phosphate handling are regulated and how the effect of these other factors are integrated to yield the measurable serum phosphate are only now beginning to be studied.
Knockdown of mineralocorticoid or angiotensin II type 1 receptor gene expression in the paraventricular nucleus prevents angiotensin II hypertension in rats
Authors: Chen Aidong , Huang Bing S. , Wang Hong‐Wei , Ahmad Monir , Leenen Frans H. H.
Published: June 2014

Key points

Chronic subcutaneous infusion of Ang II causes a progressive increase in blood pressure (BP) associated with significant increases in angiotensin type 1 receptor (AT1R) and mineralocorticoid receptor (MR) expression in hypothalamic nuclei.

Intra‐paraventricular nucleus (PVN) infusion of AAV‐ AT1aR‐siRNA or of AAV‐MR‐siRNA markedly knockdown AT1a‐R or MR expression in the PVN but not in the subfornical organ; or supraoptic nucleus, and prevent most of the increase in BP.

These findings indicate that increased MR and AT1R activation in the PVN play a critical role in Ang II‐induced hypertension in rats on regular salt intake.

Circulating Ang II activates an aldosterone‐mineralocorticoid receptor (MR) – angiotensin II (Ang II) – angiotensin type 1 receptor (AT1R) pathway in the hypothalamus. To obtain insights into the actual neuronal projections involved, adeno‐associated virus carrying small interfering RNA against either AT1aR (AAV‐AT1aR‐siRNA) or MR (AAV‐MR‐siRNA) were infused into the paraventricular nucleus (PVN) in Wistar rats. Intra‐PVN infusion of AAV‐AT1aR‐siRNA or AAV‐MR‐siRNA decreased AT1R or MR expression in the PVN but not in the subfornical organ (SFO) or supraoptic nucleus (SON). Subcutaneous infusion of Ang II at 500 ng kg−1 min−1 for 2 weeks increased mean arterial pressure by 60–70 mmHg, and increased AT1R and MR expression in the SFO, SON and PVN. Intra‐PVN AT1aR‐siRNA prevented the Ang II‐induced increase in AT1R but not MR expression in the PVN, and MR‐siRNA prevented MR but not AT1R expression in the PVN. The increases in AT1R and MR expression in both the SFO and the SON were not changed by the two AAV‐siRNAs. Specific knockdown of AT1R or MR in the PVN by AAV‐siRNA each prevented most of the Ang II‐induced hypertension. Prevention of the subcutaneous Ang II‐induced increase in MR but not the increase in AT1R by knockdown of MR and vice versa suggests an independent regulation of MR and AT1R expression in the PVN. Both AT1R and MR activation in the PVN play a critical role in Ang II‐induced hypertension in rats.
Blood pressure is maintained during dehydration by hypothalamic paraventricular nucleus‐driven tonic sympathetic nerve activity
Authors: Holbein Walter W. , Bardgett Megan E. , Toney Glenn M.
Published: June 2014

Key points

At normal resting mean arterial pressure (MAP), sympathetic nerve activity (SNA) mostly consists of respiratory and cardiac rhythmic bursts of action potentials.

In animal models of sympathetic hyperactivity, elevated SNA and MAP become reliant on activity of neurones in the hypothalamic paraventricular nucleus (PVN).

Dehydrated (DH) rats (48 h water deprived) were used as a model of sympathetic hyperactivity. As expected, acute PVN inhibition reduced MAP and integrated splanchnic SNA (sSNA) in DH rats, but had no effect in euhydrated controls. Unexpectedly, the fall of sSNA in DH rats was due to a reduction of irregular, tonic activity as neither respiratory nor cardiac rhythmic bursting was significantly affected.

We conclude that MAP is largely maintained during dehydration by PVN‐driven tonic SNA and speculate that a normally quiescent tonic component of SNA might also be recruited in chronic diseases (hypertension, heart failure, obesity) where PVN activation drives sympathetic hyperactivity.

Resting sympathetic nerve activity (SNA) consists primarily of respiratory and cardiac rhythmic bursts of action potentials. During homeostatic challenges such as dehydration, the hypothalamic paraventricular nucleus (PVN) is activated and drives SNA in support of arterial pressure (AP). Given that PVN neurones project to brainstem cardio‐respiratory regions that generate bursting patterns of SNA, we sought to determine the contribution of PVN to support of rhythmic bursting of SNA during dehydration and to elucidate which bursts dominantly contribute to maintenance of AP. Euhydrated (EH) and dehydrated (DH) (48 h water deprived) rats were anaesthetized, bilaterally vagotomized and underwent acute PVN inhibition by bilateral injection of the GABA‐A receptor agonist muscimol (0.1 nmol in 50 nl). Consistent with previous studies, muscimol had no effect in EH rats (n = 6), but reduced mean AP (MAP; P < 0.001) and integrated splanchnic SNA (sSNA; P < 0.001) in DH rats (n = 6). Arterial pulse pressure was unaffected in both groups. Muscimol reduced burst frequency of phrenic nerve activity (P < 0.05) equally in both groups without affecting the burst amplitude–duration integral (i.e. area under the curve). PVN inhibition did not affect the amplitude of the inspiratory peak, expiratory trough or expiratory peak of sSNA in either group, but reduced cardiac rhythmic sSNA in DH rats only (P < 0.001). The latter was largely reversed by inflating an aortic cuff to restore MAP (n = 5), suggesting that the muscimol‐induced reduction of cardiac rhythmic sSNA in DH rats was an indirect effect of reducing MAP and thus arterial baroreceptor input. We conclude that MAP is largely maintained in anaesthetized DH rats by a PVN‐driven component of sSNA that is neither respiratory nor cardiac rhythmic.
Synaptic mechanisms underlying cholinergic control of thalamic reticular nucleus neurons
Authors: Beierlein Michael
Published: June 2014

Neuronal networks of the thalamus are the target of extensive cholinergic projections from the basal forebrain and the brainstem. Activation of these afferents can regulate neuronal excitability, transmitter release, and firing patterns in thalamic networks, thereby altering the flow of sensory information during distinct behavioural states. However, cholinergic regulation in the thalamus has been primarily examined by using receptor agonist and antagonist, which has precluded a detailed understanding of the spatiotemporal dynamics that govern cholinergic signalling under physiological conditions. This review summarizes recent studies on cholinergic synaptic transmission in the thalamic reticular nucleus (TRN), a brain structure intimately involved in the control of sensory processing and the generation of rhythmic activity in the thalamocortical system. This work has shown that acetylcholine (ACh) released from individual axons can rapidly and reliably activate both pre‐ and postsynaptic cholinergic receptors, thereby controlling TRN neuronal activity with high spatiotemporal precision.
Power coupling and utilization efficiencies of silicon-depositing plasmas in mixtures of H2, SiH4, Si2H6, and Si3H8
Authors: Sobolewski Mark A. , Ridgeway Robert G. , Bitner Mark D. , Sinatore Dino , Hurley Patrick T.
Published: June 2014

Adding Si2H6 or Si3H8 additives to SiH4/H2 discharges increases the growth rates for thin films of microcrystalline and amorphous silicon, but the reasons for this increase are not well understood. To better distinguish the chemical and physical from electrical effects of these additives, a comprehensive electrical study was performed for mixtures of H2, SiH4, Si2H6, and Si3H8. The power coupling efficiency, power utilization efficiency, voltage, current, impedance, and phase were measured as a function of total pressure, electrode gap, gas mixture, rf power, and time. The measurements identified a regime of pressure and gap in which the electrical behavior is optimized. In this regime, the power coupling efficiency is quite high and insensitive to gas mixture, and the power utilization efficiency also does not vary dramatically with mixture. Therefore, in this regime, chemical or physical effects of additives on growth rates predominate over electrical effects. Impedance models of the plasma and sheaths provide explanations for the optimized regime and its correlation with impedance phase. In addition, electrical signals were identified that can be used to detect a transient in the gas-phase density of silicon-containing molecules during deposition as well as other transient phenomena. The signals show promise for use in process monitoring and control.
Astrocytes Control Appetite
Authors: Gough Nancy R.
Published: July 2014

Astrocytes respond to a hormone that signals satiety to control synaptic plasticity of neurons that regulate eating.
Ostreid Herpesvirus 1 Infection among Pacific Oyster (Crassostrea gigas) Spat: Relevance of Water Temperature to Virus Replication and Circulation Prior to the Onset of Mortality
Authors: Renault Tristan , Bouquet Anne Lise , Maurice Julien-Thomas , Lupo Coralie , Blachier Philippe
Published: June 2014

A number of bivalve species worldwide, including the Pacific oyster,
Crassostrea gigas
, have been affected by mass mortality events associated with herpesviruses, resulting in significant losses. A particular herpesvirus was purified from naturally infected larval Pacific oysters, and its genome was completely sequenced. This virus has been classified as
Ostreid herpesvirus 1
(OsHV-1) within the family
Malacoherpesviridae
. Since 2008, mass mortality outbreaks among
C. gigas
in Europe have been related to the detection of a variant of OsHV-1 called μVar. Additional data are necessary to better describe mortality events in relation to environmental-parameter fluctuations and OsHV-1 detection. For this purpose, a single batch of Pacific oyster spat was deployed in 4 different locations in the Marennes-Oleron area (France): an oyster pond (“claire”), a shellfish nursery, and two locations in the field. Mortality rates were recorded based on regular observation, and samples were collected to search for and quantify OsHV-1 DNA by real-time PCR. Although similar massive mortality rates were reported at the 4 sites, mortality was detected earlier in the pond and in the nursery than at both field sites. This difference may be related to earlier increases in water temperature. Mass mortality was observed among oysters a few days after increases in the number of PCR-positive oysters and viral-DNA amounts were recorded. An initial increment in the number of PCR-positive oysters was reported at both field sites during the survey in the absence of significant mortality. During this period, the water temperature was below 16°C.
Improved Production of a Heterologous Amylase in Saccharomyces cerevisiae by Inverse Metabolic Engineering
Authors: Liu Zihe , Liu Lifang , Österlund Tobias , Hou Jin , Huang Mingtao , Fagerberg Linn , Petranovic Dina , Uhlén Mathias , Nielsen Jens
Published: June 2014

The increasing demand for industrial enzymes and biopharmaceutical proteins relies on robust production hosts with high protein yield and productivity. Being one of the best-studied model organisms and capable of performing posttranslational modifications, the yeast
Saccharomyces cerevisiae
is widely used as a cell factory for recombinant protein production. However, many recombinant proteins are produced at only 1% (or less) of the theoretical capacity due to the complexity of the secretory pathway, which has not been fully exploited. In this study, we applied the concept of inverse metabolic engineering to identify novel targets for improving protein secretion. Screening that combined UV-random mutagenesis and selection for growth on starch was performed to find mutant strains producing heterologous amylase 5-fold above the level produced by the reference strain. Genomic mutations that could be associated with higher amylase secretion were identified through whole-genome sequencing. Several single-point mutations, including an S196I point mutation in the
VTA1
gene coding for a protein involved in vacuolar sorting, were evaluated by introducing these to the starting strain. By applying this modification alone, the amylase secretion could be improved by 35%. As a complement to the identification of genomic variants, transcriptome analysis was also performed in order to understand on a global level the transcriptional changes associated with the improved amylase production caused by UV mutagenesis.
Spread and Transmission of Bacterial Pathogens in Experimental Populations of the Nematode Caenorhabditis elegans
Authors: Diaz S. Anaid , Restif Olivier
Published: June 2014

Caenorhabditis elegansis frequently used as a model species for the study of bacterial virulence and innate immunity. In recent years, diverse mechanisms contributing to the nematode's immune response to bacterial infection have been discovered. Yet despite growing interest in the biochemical and molecular basis of nematode-bacterium associations, many questions remain about their ecology. Although recent studies have demonstrated that free-living nematodes could act as vectors of opportunistic pathogens in soil, the extent to which worms may contribute to the persistence and spread of these bacteria has not been quantified. We conducted a series of experiments to test whether colonization of and transmission betweenC. elegansnematodes could enable two opportunistic pathogens (Salmonella entericaandPseudomonas aeruginosa) to spread on agar plates occupied byEscherichia coli. We monitored the transmission ofS. entericaandP. aeruginosafrom single infected nematodes to their progeny and measured bacterial loads both within worms and on the plates. In particular, we analyzed three factors affecting the dynamics of bacteria: (i) initial source of the bacteria, (ii) bacterial species, and (iii) feeding behavior of the host. Results demonstrate that worms increased the spread of bacteria through shedding and transmission. Furthermore, we found that despiteP. aeruginosa's relatively high transmission rate among worms, its pathogenic effects reduced the overall number of worms colonized. This study opens new avenues to understand the role of nematodes in the epidemiology and evolution of pathogenic bacteria in the environment.
Suicin 90-1330 from a Nonvirulent Strain of Streptococcus suis: a Nisin-Related Lantibiotic Active on Gram-Positive Swine Pathogens
Authors: LeBel Geneviève , Vaillancourt Katy , Frenette Michel , Gottschalk Marcelo , Grenier Daniel
Published: June 2014

Streptococcus suis
serotype 2 is known to cause severe infections (meningitis, endocarditis, and septicemia) in pigs and is considered an emerging zoonotic agent. Antibiotics have long been used in the swine industry for disease treatment/prevention and growth promoters. This pattern of utilization resulted in the spread of antibiotic resistance in
S. suis
worldwide. Interestingly, pigs may harbor
S. suis
in their tonsils without developing diseases, while North American strains belonging to the sequence type 28 (ST28) are nonvirulent in animal models. Consequently, the aim of this study was to purify and characterize a bacteriocin produced by a nonvirulent strain of
S. suis
serotype 2, with a view to a potential therapeutic and preventive application.
S. suis
90-1330 belonging to ST28 and previously shown to be nonvirulent in an animal model exhibited antibacterial activity toward all
S. suis
pathogenic isolates tested. The bacteriocin produced by this strain was purified to homogeneity by cationic exchange and reversed-phase fast protein liquid chromatography. Given its properties (molecular mass of <4 kDa, heat, pH and protease stability, and the presence of modified amino acids), the bacteriocin, named suicin 90-1330, belongs to the lantibiotic class. Using a DNA-binding fluorophore, the bacteriocin was found to possess a membrane permeabilization activity. When tested on other swine pathogens, the suicin showed activity against
Staphylococcus hyicus
and
Staphylococcus aureus
, whereas it was inactive against all Gram-negative bacteria tested. Amino acid sequencing of the purified bacteriocin showed homology (90.9% identity) with nisin U produced by
Streptococcus uberis
. The putative gene cluster involved in suicin production was amplified by PCR and sequence analysis revealed the presence of 11 open reading frames, including the structural gene and those required for the modification of amino acids, export, regulation, and immunity. Further studies will evaluate the ability of suicin 90-1330 or the producing strain to prevent experimental
S. suis
infections in pigs.
Multilocus Sequence Analysis of the Marine Bacterial Genus Tenacibaculum Suggests Parallel Evolution of Fish Pathogenicity and Endemic Colonization of Aquaculture Systems
Authors: Habib Christophe , Houel Armel , Lunazzi Aurélie , Bernardet Jean-François , Olsen Anne Berit , Nilsen Hanne , Toranzo Alicia E. , Castro Nuria , Nicolas Pierre , Duchaud Eric
Published: June 2014

The genus
Tenacibaculum
, a member of the family
Flavobacteriaceae
, is an abundant component of marine bacterial ecosystems that also hosts several fish pathogens, some of which are of serious concern for marine aquaculture. Here, we applied multilocus sequence analysis (MLSA) to 114 representatives of most known species in the genus and of the worldwide diversity of the major fish pathogen
Tenacibaculum maritimum
. Recombination hampers precise phylogenetic reconstruction, but the data indicate intertwined environmental and pathogenic lineages, which suggests that pathogenicity evolved independently in several species. At lower phylogenetic levels recombination is also important, and the species
T. maritimum
constitutes a cohesive group of isolates. Importantly, the data reveal no trace of long-distance dissemination that could be linked to international fish movements. Instead, the high number of distinct genotypes suggests an endemic distribution of strains. The MLSA scheme and the data described in this study will help in monitoring
Tenacibaculum
infections in marine aquaculture; we show, for instance, that isolates from tenacibaculosis outbreaks in Norwegian salmon farms are related to
T. dicentrarchi
, a recently described species.
LitR Is a Repressor of syp Genes and Has a Temperature-Sensitive Regulatory Effect on Biofilm Formation and Colony Morphology in Vibrio (Aliivibrio) salmonicida
Authors: Hansen Hilde , Bjelland Ane Mohn , Ronessen Maria , Robertsen Espen , Willassen Nils Peder
Published: June 2014

Vibrio
(
Aliivibrio
)
salmonicida
is the etiological agent of cold water vibriosis, a disease in farmed Atlantic salmon (
Salmo salar
) that is kept under control due to an effective vaccine. A seawater temperature below 12°C is normally required for disease development. Quorum sensing (QS) is a cell density-regulated communication system that bacteria use to coordinate activities involved in colonization and pathogenesis, and we have previously shown that inactivation of the QS master regulator LitR attenuates the
V. salmonicida
strain LFI1238 in a fish model. We show here that strain LFI1238 and a panel of naturally occurring
V. salmonicida
strains are poor biofilm producers. Inactivation of
litR
in the LFI1238 strain enhances medium- and temperature-dependent adhesion, rugose colony morphology, and biofilm formation. Chemical treatment and electron microscopy of the biofilm identified an extracellular matrix consisting mainly of a fibrous network, proteins, and polysaccharides. Further, by microarray analysis of planktonic and biofilm cells, we identified a number of genes regulated by LitR and, among these, were homologues of the
Vibrio fischeri
symbiosis polysaccharide (
syp
) genes. The
syp
genes were regulated by LitR in both planktonic and biofilm lifestyle analyses. Disruption of
syp
genes in the
V. salmonicida
Δ
litR
mutant alleviated adhesion, rugose colony morphology, and biofilm formation. Hence, LitR is a repressor of
syp
transcription that is necessary for expression of the phenotypes examined. The regulatory effect of LitR on colony morphology and biofilm formation is temperature sensitive and weak or absent at temperatures above the bacterium's upper threshold for pathogenicity.
A Pseudomonas putida Strain Genetically Engineered for 1,2,3-Trichloropropane Bioremediation
Authors: Samin Ghufrana , Pavlova Martina , Arif M. Irfan , Postema Christiaan P. , Damborsky Jiri , Janssen Dick B.
Published: June 2014

1,2,3-Trichloropropane (TCP) is a toxic compound that is recalcitrant to biodegradation in the environment. Attempts to isolate TCP-degrading organisms using enrichment cultivation have failed. A potential biodegradation pathway starts with hydrolytic dehalogenation to 2,3-dichloro-1-propanol (DCP), followed by oxidative metabolism. To obtain a practically applicable TCP-degrading organism, we introduced an engineered haloalkane dehalogenase with improved TCP degradation activity into the DCP-degrading bacterium
Pseudomonas putida
MC4. For this purpose, the dehalogenase gene (
dhaA31
) was cloned behind the constitutive
dhlA
promoter and was introduced into the genome of strain MC4 using a transposon delivery system. The transposon-located antibiotic resistance marker was subsequently removed using a resolvase step. Growth of the resulting engineered bacterium,
P. putida
MC4-5222, on TCP was indeed observed, and all organic chlorine was released as chloride. A packed-bed reactor with immobilized cells of strain MC4-5222 degraded >95% of influent TCP (0.33 mM) under continuous-flow conditions, with stoichiometric release of inorganic chloride. The results demonstrate the successful use of a laboratory-evolved dehalogenase and genetic engineering to produce an effective, plasmid-free, and stable whole-cell biocatalyst for the aerobic bioremediation of a recalcitrant chlorinated hydrocarbon.
Design and Validation of Four New Primers for Next-Generation Sequencing To Target the 18S rRNA Genes of Gastrointestinal Ciliate Protozoa
Authors: Ishaq Suzanne L. , Wright André-Denis G.
Published: June 2014

Four new primers and one published primer were used to PCR amplify hypervariable regions within the protozoal 18S rRNA gene to determine which primer pair provided the best identification and statistical analysis. PCR amplicons of 394 to 498 bases were generated from three primer sets, sequenced using Roche 454 pyrosequencing with Titanium, and analyzed using the BLAST database (NCBI) and MOTHUR version 1.29. The protozoal diversity of rumen contents from moose in Alaska was assessed. In the present study, primer set 1, P-SSU-316F and GIC758R (amplicon of 482 bases), gave the best representation of diversity using BLAST classification, and the set amplified
Entodinium simplex
and
Ostracodinium
spp., which were not amplified by the other two primer sets. Primer set 2, GIC1080F and GIC1578R (amplicon of 498 bases), had similar BLAST results and a slightly higher percentage of sequences that were identified with a higher sequence identity. Primer sets 1 and 2 are recommended for use in ruminants. However, primer set 1 may be inadequate to determine protozoal diversity in nonruminants. The amplicons created by primer set 1 were indistinguishable for certain species within the genera
Bandia
,
Blepharocorys
,
Polycosta
, and
Tetratoxum
and between
Hemiprorodon gymnoprosthium
and
Prorodonopsis
coli
, none of which are normally found in the rumen.
Role of Bacillus subtilis Error Prevention Oxidized Guanine System in Counteracting Hexavalent Chromium-Promoted Oxidative DNA Damage
Authors: Santos-Escobar Fernando , Gutiérrez-Corona J. Félix , Pedraza-Reyes Mario
Published: June 2014

Chromium pollution is potentially detrimental to bacterial soil communities, compromising carbon and nitrogen cycles that are essential for life on earth. It has been proposed that intracellular reduction of hexavalent chromium [Cr(VI)] to trivalent chromium [Cr(III)] may cause bacterial death by a mechanism that involves reactive oxygen species (ROS)-induced DNA damage; the molecular basis of the phenomenon was investigated in this work. Here, we report that
Bacillus subtilis
cells lacking a functional error prevention oxidized guanine (GO) system were significantly more sensitive to Cr(VI) treatment than cells of the wild-type (WT) strain, suggesting that oxidative damage to DNA is involved in the deleterious effects of the oxyanion. In agreement with this suggestion, Cr(VI) dramatically increased the ROS concentration and induced mutagenesis in a GO-deficient
B. subtilis
strain. Alkaline gel electrophoresis (AGE) analysis of chromosomal DNA of WT and ΔGO mutant strains subjected to Cr(VI) treatment revealed that the DNA of the ΔGO strain was more susceptible to DNA glycosylase Fpg attack, suggesting that chromium genotoxicity is associated with 7,8-dihydro-8-oxodeoxyguanosine (8-oxo-G) lesions. In support of this notion, specific monoclonal antibodies detected the accumulation of 8-oxo-G lesions in the chromosomes of
B. subtilis
cells subjected to Cr(VI) treatment. We conclude that Cr(VI) promotes mutagenesis and cell death in
B. subtilis
by a mechanism that involves radical oxygen attack of DNA, generating 8-oxo-G, and that such effects are counteracted by the prevention and repair GO system.
Cross Talk between NDR Kinase Pathways Coordinates Cytokinesis with Cell Separation in Schizosaccharomyces pombe
Authors: Gupta Sneha , Govindaraghavan Meera , McCollum Dannel
Published: June 2014

NDR (nuclear Dbf-2-related) kinases constitute key regulatory nodes in signaling networks that control multiple biological processes such as growth, proliferation, mitotic exit, morphogenesis, and apoptosis. Two NDR pathways called the septation initiation network (SIN) and the morphogenesis Orb6 network (MOR) exist in the fission yeast
Schizosaccharomyces pombe
. The SIN promotes cytokinesis, and the MOR drives cell separation at the end of cytokinesis and polarized growth during interphase. We showed previously that cross talk exists between these two pathways, with the SIN inhibiting the MOR during cytokinesis through phosphorylation of the MOR component Nak1 by the SIN Sid2 kinase. The reason for this inhibition remained uncertain. We show here that failure to inhibit MOR signaling during cytokinesis results in cell lysis at the site of septum formation. Time-lapse analysis revealed that MOR signaling during cytokinesis causes cells to prematurely initiate septum degradation/cell separation. The cell lysis phenotype is due to premature initiation of cell separation because it can be rescued by mutations in genes required for cell separation/septum degradation. We also shed further light on how the SIN inhibits the MOR. Sid2 phosphorylation of the MOR proteins Sog2 and Nak1 is required to prevent cell lysis during cytokinesis. Together, these results show that SIN inhibition of the MOR enforces proper temporal ordering of cytokinetic events.
Multicentre observational cohort study of NSAIDs as risk factors for postoperative adverse events in gastrointestinal surgery
Authors: Nepogodiev Dmitri , Chapman Stephen J , Glasbey James C D , Kelly Michael , Khatri Chetan , Fitzgerald J Edward , Bhangu Aneel
Published: June 2014

Introduction

Non-steroidal anti-inflammatory drugs (NSAIDs) are recommended as postoperative analgesia by the Enhanced Recovery After Surgery Society. Recent studies have raised concerns that NSAID administration following colorectal anastomosis may be associated with increased risk of anastomotic leak. This multicentre study aims to determine NSAIDs’ safety profile following gastrointestinal resection.

Methods and analysis

This prospective, multicentre cohort study will be performed over a 2-week period utilising a collaborative methodology. Consecutive adults undergoing open or laparoscopic, elective or emergency gastrointestinal resection will be included. The primary end point will be the 30-day morbidity, assessed using the Clavien-Dindo classification. This study will be disseminated through medical student networks, with an anticipated recruitment of at least 900 patients. The study will be powered to detect a 10% increase in complication rates with NSAID use.

Ethics and dissemination

Following the Research Ethics Committee Chairperson's review, a formal waiver was received. This study will be registered as a clinical audit or service evaluation at each participating hospital. Dissemination will take place through previously described novel research collaborative networks.
Effect of hyperglycaemia in pregnancy on adiposity in their infants in India: a protocol of a multicentre cohort study
Authors: Babu Giridhara R , Garadi Lavanya , Murthy G V S , Kinra Sanjay
Published: June 2014

Introduction

The carbohydrate ‘fuel’ metabolism in a pregnant woman may have a long-term impact on the development of her offspring (‘fuel-mediated teratogenesis’ hypothesis) including in utero exposure to maternal hyperglycaemia leading to fetal hyperinsulinaemia, and the consequent increase in fetal fat cells. Therefore, a feed-forward loop can exist of rising adiposity and hyperinsulinaemia throughout childhood, perhaps leading to obesity and diabetes in later life. There is a need for prospective examination of body fat distribution in children born to mothers with different glycaemic levels to understand the plausible association between glucose metabolism and future risk of diabetes in offspring. The hypothesis is that maternal glucose levels in pregnant women are related to skinfold thickness in their infants.

Methods and analysis

Hyperglycaemia in pregnancy and adiposity in infants is a multicentre cohort study to evaluate the effects of glucose levels in pregnancy on the risk of adverse infant outcomes, especially in predicting the risk of chronic diseases in infants. The study aims to recruit 1045 participants over a period of 1 year, who will be followed up irrespective of their glycaemia status for a period of 15 months, beginning in the 24th week of gestation. The glucose levels in pregnant women would be obtained through oral glucose tolerance testing. The primary outcome of our study was skinfold thickness in infants at the age of 6 weeks, 3rd and 9th month (as a proxy for fat distribution/adiposity).

Ethics and dissemination

The institutional review board at The Indian Institute of Public Health (IIPH)-Hyderabad, Public Health Foundation of India has approved the protocol. All participants are required to provide written informed consent.
KIR channel activation contributes to onset and steady-state exercise hyperemia in humans
Authors: Crecelius Anne R. , Luckasen Gary J. , Larson Dennis G. , Dinenno Frank A.
Published: June 2014

We tested the hypothesis that activation of inwardly rectifying potassium (KIR) channels and Na+-K+-ATPase, two pathways that lead to hyperpolarization of vascular cells, contributes to both the onset and steady-state hyperemic response to exercise. We also determined whether after inhibiting these pathways nitric oxide (NO) and prostaglandins (PGs) are involved in the hyperemic response. Forearm blood flow (FBF; Doppler ultrasound) was determined during rhythmic handgrip exercise at 10% maximal voluntary contraction for 5 min in the following conditions: control [saline; trial 1 ( T1)]; with combined inhibition of KIR channels and Na+-K+-ATPase alone [via barium chloride (BaCl2) and ouabain, respectively; trial 2 ( T2)]; and with additional combined nitric oxide synthase ( NG-monomethyl-l-arginine) and cyclooxygenase inhibition [ketorolac; trial 3 ( T3)]. In T2, the total hyperemic responses were attenuated ∼50% from control ( P < 0.05) at exercise onset, and there was minimal further effect in T3 ( protocol 1; n = 11). In protocol 2 ( n = 8), steady-state FBF was significantly reduced during T2 vs. T1 (133 ± 15 vs. 167 ± 17 ml/min; Δ from control: −20 ± 3%; P < 0.05) and further reduced during T3 (120 ± 15 ml/min; −29 ± 3%; P < 0.05 vs. T2). In protocol 3 ( n = 8), BaCl2 alone reduced FBF during onset (∼50%) and steady-state exercise (∼30%) as observed in protocols 1 and 2, respectively, and addition of ouabain had no further impact. Our data implicate activation of KIR channels as a novel contributing pathway to exercise hyperemia in humans.
Sex differences in the mechanisms underlying long QT syndrome
Authors: Salama Guy , Bett Glenna C. L.
Published: June 2014

Sexual dimorphism is a well-established phenomenon, but its degree varies tremendously among species. Since the early days of Einthoven's development of the three-lead galvanometer ECG, we have known there are marked differences in QT intervals of men and women. It required over a century to appreciate the profound implications of sex-based electrophysiological differences in QT interval on the panoply of sex differences with respect to arrhythmia risk, drug sensitivity, and treatment modalities. Little is known about the fundamental mechanism responsible for sex differences in electrical substrate of the human heart, in large part due to the lack of tissue availability. Animal models are an important research tool, but species differences in the sexual dimorphism of the QT interval, the ionic currents underlying the cardiac repolarization, and effects of sex steroids make it difficult to interpolate animal to human sex differences. In addition, in some species, different strains of the same animal model yield conflicting data. Each model has its strengths, such as ease of genetic manipulation in mice or size in dogs. However, many animals do not reproduce the sexual dimorphism of QT seen in humans. To match sex linked prolongation of QT interval and arrhythmogenic phenotype, the current data suggest that the rabbit may be best suited to provide insight into sex differences in humans. In the future, emerging technologies such as induced pluripotent stem cell derived cardiac myocyte systems may offer the opportunity to study sex differences in a controlled hormonal situation in the context of a sex specific human model system.
Age-related loss of SirT1 expression results in dysregulated human vascular smooth muscle cell function
Authors: Thompson Anne Marie , Wagner Robert , Rzucidlo Eva M.
Published: June 2014

Loss of vascular smooth muscle cell (VSMC) function is a hallmark of vascular disease. VSMCs become increasingly dysregulated, apoptotic, and senescent as we age. Sirtuin 1 (SirT1) is a deactylase that regulates substrates associated with stress mitigation, metabolism, and aging. Our aim was to examine the role of SirT1 in vascular aging and the function this protein plays in the context of cellular response to stress and senescence. We compared endogenous SirT1 expression in young and old human donors. Human VSMC (HuVSMC) from donors ranging in age from 12 to 88 ( n = 14) were isolated and cultured. In cultured HuVSMC the levels of endogenous SirT1 were examined by Western blot analysis. We found that endogenous SirT1 protein expression inversely correlated with donor age. Additionally, we demonstrated that age-related loss of SirT1 correlated in functional deficits, diminished stress response, reduced capacity for migration, and proliferation and increased senescence. Manipulation of SirT1 levels in young cells confirmed the role of SirT1 in cellular migration and proliferation capability. Furthermore, we demonstrated that age-related loss of SirT1 was associated with the induction of VSMC senescence. With correlation to symptomatic disease, we demonstrated a significant difference in SirT1 levels from HuVSMC isolated from aged arteries that were occluded with atherosclerotic lesions ( n = 7), compared with patent sections of the same artery. Having demonstrated that endogenous SirT1 is lost with age, which correlates with a loss of capacity for vascular repair, our data explain one of the molecular changes that occurs in the aged vasculature.