bluuebunny/crossref_metadata_2025 huggingface dataset:
Requirement of Siderophore Biosynthesis for Plant Colonization by Salmonella enterica
Authors: Hao Ling-yun , Willis David Kyle , Andrews-Polymenis Helene , McClelland Michael , Barak Jeri D.
Published: April 2012

Contaminated fresh produce has become the number one vector of nontyphoidal salmonellosis to humans. However,
Salmonella enterica
genes essential for the life cycle of the organism outside the mammalian host are for the most part unknown. Screening deletion mutants led to the discovery that an
aroA
mutant had a significant root colonization defect due to a failure to replicate. AroA is part of the chorismic acid biosynthesis pathway, a central metabolic node involved in aromatic amino acid and siderophore production. Addition of tryptophan or phenylalanine to alfalfa root exudates did not restore
aroA
mutant replication. However, addition of ferrous sulfate restored replication of the
aroA
mutant, as well as alfalfa colonization. Tryptophan and phenylalanine auxotrophs had minor plant colonization defects, suggesting that suboptimal concentrations of these amino acids in root exudates were not major limiting factors for
Salmonella
replication. An
entB
mutant defective in siderophore biosynthesis had colonization and growth defects similar to those of the
aroA
mutant, and the defective phenotype was complemented by the addition of ferrous sulfate. Biosynthetic genes of each
Salmonella
siderophore, enterobactin and salmochelin, were upregulated in alfalfa root exudates, yet only enterobactin was sufficient for plant survival and persistence. Similar results in lettuce leaves indicate that siderophore biosynthesis is a widespread or perhaps universal plant colonization fitness factor for
Salmonella
, unlike phytobacterial pathogens, such as
Pseudomonas
and
Xanthomonas
.
Investigating the Function of Ddr48p in Candida albicans
Authors: Cleary I. A. , MacGregor N. B. , Saville S. P. , Thomas D. P.
Published: April 2012

Candidiasis now represents the fourth most frequent nosocomial infection both in the United States and worldwide.Candida albicansis an increasingly common threat to human health as a consequence of AIDS, steroid therapy, organ and tissue transplantation, cancer therapy, broad-spectrum antibiotics, and other immune defects. The pathogenic potential ofC. albicansis intimately related to certain key processes, including biofilm formation and filamentation. Ddr48p is a damage response protein that is significantly upregulated during both biofilm formation and filamentation, but its actual function is unknown. Previous studies have indicated that this protein may be essential inC. albicansbut notSaccharomyces cerevisiae. Here we examined the function of Ddr48p and investigated the role of this protein in biofilm formation and filamentation. We demonstrated that this protein is not essential inC. albicansand appears to be dispensable for filamentation. However,DDR48is required for the flocculation response stimulated by 3-aminotriazole-induced amino acid starvation. Furthermore, we examined the response of this deletion strain to a wide variety of environmental stressors and antifungal compounds. We observed several mild sensitivity or resistance phenotypes and also found that Ddr48p contributes to the DNA damage response ofC. albicans. The results of this study reveal that the role of this highly expressed protein goes beyond a general stress response and impinges on a key facet of pathogenesis, namely, the ability to sense and respond to changes in the host environment.
Mechanism for Regulation of the Putrescine Utilization Pathway by the Transcription Factor PuuR in Escherichia coli K-12
Authors: Nemoto Naoki , Kurihara Shin , Kitahara Yuzuru , Asada Kei , Kato Kenji , Suzuki Hideyuki
Published: April 2012

In
Escherichia coli
, putrescine is metabolized to succinate for use as a carbon and nitrogen source by the putrescine utilization pathway (Puu pathway). One gene in the
puu
gene cluster encodes a transcription factor, PuuR, which has a helix-turn-helix DNA-binding motif. DNA microarray analysis of an
E. coli puuR
mutant, in which three amino acid residues in the helix-turn-helix DNA binding motif of PuuR were mutated to alanine to eliminate DNA binding of PuuR, suggested that PuuR is a negative regulator of
puu
genes. Results of gel shift and DNase I footprint analyses suggested that PuuR binds to the promoter regions of
puuA
and
puuD
. The binding of wild-type PuuR to a DNA probe containing PuuR recognition sites was diminished with increasing putrescine concentrations
in vitro
. These results suggest that PuuR regulates the intracellular putrescine concentration by the transcriptional regulation of genes in the Puu pathway, including
puuR
itself. The
puu
gene cluster is found in
E. coli
and closely related enterobacteria, but this gene cluster is uncommon in other bacterial groups.
E. coli
and related enterobacteria may have gained the Puu pathway as an adaptation for survival in the mammalian intestine, an environment in which polyamines exist at relatively high concentrations.
Cyclic Diguanylate Inversely Regulates Motility and Aggregation in Clostridium difficile
Authors: Purcell Erin B. , McKee Robert W. , McBride Shonna M. , Waters Christopher M. , Tamayo Rita
Published: April 2012

Clostridium difficile
-associated disease is increasing in incidence and is costly to treat. Our understanding of how this organism senses its entry into the host and adapts for growth in the large bowel is limited. The small-molecule second messenger cyclic diguanylate (c-di-GMP) has been extensively studied in Gram-negative bacteria and has been shown to modulate motility, biofilm formation, and other processes in response to environmental signals, yet little is known about the functions of this signaling molecule in Gram-positive bacteria or in
C. difficile
specifically. In the current study, we investigated the function of the second messenger c-di-GMP in
C. difficile
. To determine the role of c-di-GMP in
C. difficile
, we ectopically expressed genes encoding a diguanylate cyclase enzyme, which synthesizes c-di-GMP, or a phosphodiesterase enzyme, which degrades c-di-GMP. This strategy allowed us to artificially elevate or deplete intracellular c-di-GMP, respectively, and determine that c-di-GMP represses motility in
C. difficile
, consistent with previous studies in Gram-negative bacteria, in which c-di-GMP has a negative effect on myriad modes of bacterial motility. Elevated c-di-GMP levels also induced clumping of
C. difficile
cells, which may signify that
C. difficile
is capable of forming biofilms in the host. In addition, we directly quantified, for the first time, c-di-GMP production in a Gram-positive bacterium. This work demonstrates the effect of c-di-GMP on the motility of a Gram-positive bacterium and on aggregation of
C. difficile
, which may be relevant to the function of this signaling molecule during infection.
Complete DNA Sequence Analysis of Enterohemorrhagic Escherichia coli Plasmid pO157_2 in β-Glucuronidase-Positive E. coli O157:H7 Reveals a Novel Evolutionary Path
Authors: Rump L. V. , Meng J. , Strain E. A. , Cao G. , Allard M. W. , Gonzalez-Escalona N.
Published: April 2012

Strains of enterohemorragic
Escherichia coli
(EHEC) O157:H7 that are non-sorbitol fermenting (NSF) and β-glucuronidase negative (GUD
−
) carry a large virulence plasmid, pO157 (>90,000 bp), whereas closely related sorbitol-fermenting (SF)
E. coli
O157:H
−
strains carry plasmid pSFO157 (>120,000 bp). GUD
+
NSF O157:H7 strains are presumed to be precursors of GUD
−
NSF O157:H7 strains that also carry pO157. In this study, we report the complete sequence of a novel virulence plasmid, pO157-2 (89,762 bp), isolated from GUD
+
NSF O157:H7 strain G5101. PCR analysis confirmed the presence of pO157-2 in six other strains of GUD
+
NSF O157:H7. pO157-2 carries genes associated with virulence (e.g., hemolysin genes) and conjugation (
tra
and
trb
genes) but lacks
katP
and
espP
present in pO157. Comparative analysis of the three EHEC plasmids shows that pO157-2 is highly related to pO157 and pSFO157 but not ancestral to pO157. These results indicated that GUD
+
NSF O157:H7 strains might not be direct precursors to GUD
−
NSF O157:H7 as previously proposed but rather have evolved independently from a common ancestor.
Effects of Lipopolysaccharide Biosynthesis Mutations on K1 Polysaccharide Association with the Escherichia coli Cell Surface
Authors: Jiménez Natalia , Senchenkova Sofya N. , Knirel Yuriy A. , Pieretti Giuseppina , Corsaro Maria M. , Aquilini Eleonora , Regué Miguel , Merino Susana , Tomás Juan M.
Published: April 2012

The presence of cell-bound K1 capsule and K1 polysaccharide in culture supernatants was determined in a series of in-frame nonpolar core biosynthetic mutants from
Escherichia coli
KT1094 (K1, R1 core lipopolysaccharide [LPS] type) for which the major core oligosaccharide structures were determined. Cell-bound K1 capsule was absent from mutants devoid of phosphoryl modifications on
l
-
glycero
-
d
-
manno
-heptose residues (HepI and HepII) of the inner-core LPS and reduced in mutants devoid of phosphoryl modification on HepII or devoid of HepIII. In contrast, in all of the mutants, K1 polysaccharide was found in culture supernatants. These results were confirmed by using a mutant with a deletion spanning from the
hldD
to
waaQ
genes of the
waa
gene cluster to which individual genes were reintroduced. A nuclear magnetic resonance (NMR) analysis of core LPS from HepIII-deficient mutants showed an alteration in the pattern of phosphoryl modifications. A cell extract containing both K1 capsule polysaccharide and LPS obtained from an O-antigen-deficient mutant could be resolved into K1 polysaccharide and core LPS by column chromatography only when EDTA and deoxycholate (DOC) buffer were used. These results suggest that the K1 polysaccharide remains cell associated by ionically interacting with the phosphate-negative charges of the core LPS.
The Response Regulator RcsB Activates Expression of Mat Fimbriae in Meningitic Escherichia coli
Authors: Lehti Timo A. , Heikkinen Johanna , Korhonen Timo K. , Westerlund-Wikström Benita
Published: April 2012

The common colonization factor of
Escherichia coli
, the Mat (also termed ECP) fimbria, functions to advance biofilm formation on inert surfaces as well as bacterial adherence to epithelial cells and subsequent colonization. We used global mini-Tn
5
transposon mutagenesis to identify novel regulators of biofilm formation by the meningitic
E. coli
isolate IHE 3034. Of the 4,418 transformants, we found 17 that were impaired in biofilm formation. Most of these mutants were affected in lipopolysaccharide synthesis and were reduced in growth but not in Mat fimbria expression. In contrast, two mutants grew well but did not express Mat fimbria. The insertions in these two mutants were located at different sites of the
rcsB
gene, which encodes a DNA-binding response regulator of the Rcs response regulon. The mutations abrogated temperature-dependent biofilm formation by IHE 3034, and the phenotype correlated with loss of
mat
expression. The defect in biofilm formation in the
rcsB
mutant was reversed upon complementation with
rcsB
as well as by overexpression of structural
mat
genes but not by overexpression of the fimbria-specific activator gene
matA
. Monitoring of the
mat
operon promoter activity with chromosomal reporter fusions showed that the RcsB protein and an RcsAB box in the
mat
regulatory region, but not RcsC, RcsD, AckA, and Pta, are essential for initiation of
mat
transcription. Gel retardation assays showed that RcsB specifically binds to the
mat
promoter DNA, which enables its function in promoting biofilm formation by
E. coli
.
Diagnosis, treatment and prevention of ankle sprains: an evidence-based clinical guideline
Authors: Kerkhoffs Gino M , van den Bekerom Michel , Elders Leon A M , van Beek Peter A , Hullegie Wim A M , Bloemers Guus M F M , de Heus Elly M , Loogman Masja C M , Rosenbrand Kitty C J G M , Kuipers Ton , Hoogstraten J W A P , Dekker Rienk , ten Duis Henk-Jan , van Dijk C Niek , van Tulder Maurits W , van der Wees Philip J , de Bie Rob A
Published: April 2012

Ankle injuries are a huge medical and socioeconomic problem. Many people have a traumatic injury of the ankle, most of which are a result of sports. Total costs of treatment and work absenteeism due to ankle injuries are high. The prevention of recurrences can result in large savings on medical costs. A multidisciplinary clinical practice guideline was developed with the aim to prevent further health impairment of patients with acute lateral ankle ligament injuries by giving recommendations with respect to improved diagnostic and therapeutic opportunities. The recommendations are based on evidence from published scientific research, which was extensively discussed by the guideline committee. This clinical guideline is helpful for healthcare providers who are involved in the management of patients with ankle injuries.
Fitness levels and physical activity among class A drug users entering prison
Authors: Fischer Jan , Butt Christine , Dawes Helen , Foster Charlie , Neale Joanne , Plugge Emma , Wheeler Carly , Wright Nat
Published: April 2012

Background

Physical activity could benefit drug users' physiological and mental health. Previous research has suggested that physical activity levels change when drug users enter prison.

Methods

Twenty-five class A drug users who were new to prison answered physical activity and drug use cross-sectional questionnaires, took a submaximal fitness test and wore a pedometer for 1 week.

Results

Participants' mean aerobic capacity was estimated as 49 mls O2/kg/min (±12 SD). Their mean self-reported walking distance outside of prison was 4.67 miles on an average day (±4.14 SD). Pedometer data suggest they walked a mean of 1.8 miles/day in prison.

Conclusion

Many class A drug users entering prison had high levels of fitness and physical activity before admission, often gained from walking. Walking activity reduced when they entered prison, posing a challenge to maintaining healthy activity levels.
Thiazolidinediones for plaque psoriasis: a systematic review and meta-analysis
Authors: Malhotra Anita , Shafiq Nusrat , Rajagopalan Sujit , Dogra Sunil , Malhotra Samir
Published: April 2012

Background

There is some evidence for the role of inflammation and insulin resistance in the pathophysiology of plaque psoriasis. Thiazolidinediones are antidiabetic drugs that act by improving insulin sensitivity and also possess anti-inflammatory effects. Evidence from in vitro, animal and human studies has accumulated suggesting that these drugs may be of use in psoriasis.

Objective

This systematic review and meta-analysis was performed to assess the efficacy of thiazolidinediones on psoriasis severity.

Study selection

Randomised, open-label or single blind or double blind, published as well as unpublished, studies of thiazolidinedione administration compared with placebo, given to patients with plaque psoriasis for at least 8 weeks were considered for inclusion in this review. The primary outcomes were as follows: mean or mean percent change in Psoriasis Area and Severity Index (PASI) from baseline to end of treatment with pioglitazone, proportion of patients showing >75% improvement in PASI score, and proportion of patients showing >50% improvement in PASI score. Data analysis was done using Revman 5.

Findings

Twenty-seven relevant citations were identified. Two studies each for pioglitazone and rosiglitazone were included in the meta-analysis. There was a significantly greater mean decrease in PASI scores from baseline to end of treatment (−4.24 (95% CI −5.35 to −3.12)) in the pioglitazone group as compared to placebo. There was a non-significant improvement in PASI50/70 in the pooled analysis of rosiglitazone trials.

Conclusion

Pioglitazone appears to have efficacy for the treatment of psoriasis. The clinical significance of the effect and role in management of psoriasis deserve further study.
SchIBZ - Design Of Different Diesel Based Fuel Cell Systems for Seagoing Vessels and Their Evaluation
Authors: Leites Keno , Bauschulte Ansgar , Dragon Michael , Krummrich Stefan , Nehter Pedro
Published: April 2012

The project SchIBZ has the aim to develop and test a fuel cell based generator set for seagoing ships. Before the start of the system development a short study was carried out, to evaluate different solutions before choosing one. This paper contains a short review of the possibilities and selected results.
California's Hydrogen Infrastructure Funding Program
Authors: Muench Tobias R.
Published: April 2012

California's hydrogen funding program is part of the Alternative and Renewable Fuel and Vehicle Technology Program (ARFVTP) administered by the California Energy Commission (CEC). It's the single largest public initiative of its kind in the United States today. Together with various stakeholders, the program is designed to co-fund modern hydrogen stations for fueling hydrogen-powered vehicles. CEC's strategic hydrogen infrastructure planning focuses on early-adopter cluster communities. The program is strongly supported by a proactive group of seven auto-makers who committed specific numbers of vehicles (also transit buses) to be rolled out in order to sustain particular future fueling station locations. The program uses performance-based incentives to encourage lower cost, faster installed, higher capacity stations, and those that exceed California's 33.3% renewably-sourced hydrogen transportation fuel mandate. This allows for a more substantiated business case and helps leverage public funds for a successful outcome for the hydrogen fueling network in California.
The Influence of Channel Wettability on Two-Phase Flow and Polymer Electrolyte Membrane Fuel Cell Performance
Authors: Al Shakhshir Saher , Wang Yonxin , Alaefour Ibrahim E. , Li Xianguo
Published: April 2012

Water management is one of the critical issues affecting polymer electrolyte membrane fuel cell (PEMFC) performance, durability and cost. Modifying the surface wettability of flow field channels in PEMFC can improve the water management and fuel cell performance. In present work, an ex-situ investigation of the effects of channels with different surface wettability on the two phase-flow characteristics has been conducted. Horizontal graphite channel (slightly hydrophobic) and other four channels coated with polytetrafluoroethylene (PTFE) (hydrophobic), silica/Polydimethylsiloxane (PDMS) (super-hydrophobic), silica/PDMS at the bottom wall but with the side walls being raw graphite (combined surface wettability channel), and silica coated channel (superhydrophilic), are tested and visualized at room temperature and atmospheric pressure. Pressure drop measurements and two-phase flow visualization using high speed camera have been performed. Super-hydrophobic channel has shown desirable positive effect on the two-phase flow and on the PEMFC's performance compared with the other channels, especially at high current densities.
Analysis of the Potential for Cogeneration of a Proton Exchange Membrane Fuel Cell System
Authors: Almeida Silvio C. , Furtado Jose Geraldo D. , Serra Eduardo T. , Barroso Diego G.
Published: April 2012

This work investigates the electrothermal performance of a fuel cell cogeneration system based on a Proton Exchange Membrane Fuel Cell (PEMFC) unit. A simple way to analyze the performance of a PEMFC at different operating conditions, developed in a spreadsheet, is presented. Electrical and thermal energy generated are calculated as function of the operational parameters of the system. Additionally, it was verified the influence of the fuel, operation & maintenance (O&M) and capital costs in the final cost of energy (COE). This work shows that the use of cogeneration at the PEMFC studied unit results in significant reduction in the final cost of energy.
Missions and Progressions of PEFC Evaluation Project in Japan
Authors: Shingu Hiroshi , Daimaru Akimasa
Published: April 2012

Many kinds of catalysts and membranes for PEFC (Polymer electrolyte fuel cell) have been being developed at various institutes, companies or universities. To find truly practical materials among such developed ones is very important to accelerate the commercialization of Fuel Cell Electric Vehicles (FCEV). In Japan, since 2010, such a mission has been executed under "PEFC Evaluation Project" which is one of NEDO projects. In PEFC Evaluation Project, a newly developed catalyst is processed to an MEA (Membrane Electrode Assembly), and the MEA is constructed to a single cell to evaluate the fuel cell performance.
Solid Oxide Fuel Cell Stack Module Development for Fuel Flexible Power Generation
Authors: Ghezel-Ayagh Hossein , Way Richard , Huang Peng , Walzak James , Jolly Stephen , Patel Dilip , Willman Carl , Davis Keith E. , Lukas Michael D. , Borglum Brian , Tang Eric , Pastula Michael , Petri Randy , Richards Mark
Published: April 2012

FuelCell Energy is developing solid oxide fuel cell (SOFC) systems and technology for very efficient, economically viable, coal-to-electricity power plants utilizing synthesis gas (syngas) from a coal gasifier for the Department of Energy Solid State Energy Conversion Alliance (SECA) program. SOFC technology development includes cell performance enhancement, stack building block scale-up, stack tower design and testing, and development of a 250 kW module accommodating multiple stack towers.
Evaluation of the Degradation of Fuel Cell Hybrids
Authors: Williams Mark C. , Winkler Wolfgang , Suzuki Ai , Miyamoto Akira
Published: April 2012

Fuel cell hybrids are combination of energy conversion sub-systems - fuel cells and heat engines. This paper is a study of fuel cell hybrids electrochemical performance when the fuel cell sub-system is undergoing degradation. In all cases one can utilize the waste heat to improve overall efficiency through hybridization. Even degradation rates of 0.25 percent per 1000 hours corresponding to 40,000 hour life produce significant amounts of waste heat. Power loss is especially high at EOL. Hybridization utilizes waste heat and can be used if degradation occurs and if long fuel cell life is expected. The common practice is to linearize degradation. Giving a linear representation to DR, however, gives a linear structure to the area specific resistance, ASR(t). Experimental evidence shows that ASR(t) is commonly an ohmic parabolic function. Degradation rate, DRavg(t), %/1000 hours varies throughout the life of the fuel cell for ohmic parabolic degradation behavior . This paper builds from and is an extension of several papers recently published in the Journal of The Electrochemical Society (ECS), ECS Transactions, and the Journal of Fuel Cell Science and Technology.
Validity of Substituting H2-H2O Gas Mixtures for Reformate Gases in Testing Practical-Sized Solid Oxide Fuel Cells
Authors: Tanaka Yohei , Momma Akihiko , Sato Katsutoshi , Kato Tohru
Published: April 2012

To carry out performance test of solid oxide fuel cells (SOFCs) simply and safely without dealing with toxic carbon monoxide, validity of substituting H2-H2O gas mixtures for reformate gases was investigated with practical-sized single cells. H2-H2O gas mixtures were prepared by replacing CO with H2 and CO2 with H2O in equilibrium reformate compositions at high temperatures (≥ 750ºC) where CH4 concentration is negligible. It was clarified that cell voltages for the H2-H2O substitutes agreed to ones for reformate gas of kerosene or methane by 0.6-0.2% at 0-90% fuel utilization and 650-750ºC. It was concluded that the way of H2-H2O substitution will be valid as long as for SOFCs show good internal reforming performance.
Degradation Mechanism with Impurities and Life Time Estimation for SOFCs
Authors: Horita Teruhisa , Cho Dohyung , Wang Fangfang , Nishi Mina , Shimonosono Taro , Kishimoto Haruo , Yamaji Katsuhiko , Brito Manuel E. , Yokokawa Harumi
Published: April 2012

The degradation of La0.6Sr0.4Co0.2Fe0.8O3 (LSCF) cathode performance were investigated under relatively high pressures of impurities with Cr and S. AC impedance and DC currents were measured as a function of operation time. The cathode polarization rapidly increased when exposure to CrO3 and SO2 due to active sites vanishing with impurities. The distributions of Cr and S in the porous LSCF were different due to the chemical reactivity and electrochemical effects. A significant amount of SrCrO4 was deposited at the gas inlet of cathode surface region whereas small spots of SrSO4 were distributed inside the porous LSCF cathode. From the trends of polarization increase, the decrease trends and life time of cathodes were estimated for the availability of cathodes.
Carbon Dioxide Reforming of Methane on Ni-ceria-based Oxide Cermet Anode for Solid Oxide Fuel Cells
Authors: Kawano Mitsunobu , Yoshida Hiroyuki , Ueno Daisuke , Hashigami Satoshi , Inagaki Toru
Published: April 2012

Carbon dioxide reforming of methane on Ni-ceria-based oxide cermet anode of solid oxide fuel cells was carried out. Electrochemical characterizations revealed that direct internal carbon dioxide reforming operation was strongly influenced by the operation temperature. Direct internal carbon dioxide reforming operation resulted in sufficient level of conversion for power generation with methane at 750åC, while it gave insufficient level of conversion at 700 and 650åC. The results of gas analysis at 750°C also reveal that methane was effectively converted to hydrogen and carbon monoxide in the radial direction of the cell, and that hydrogen and carbon monoxide were effectively used as fuel species for electrochemical oxidation reactions.
Effect of a glucagon-like peptide 1 analog, ROSE-010, on GI motor functions in female patients with constipation-predominant irritable bowel syndrome
Authors: Camilleri Michael , Vazquez-Roque Maria , Iturrino Johanna , Boldingh Amy , Burton Duane , McKinzie Sanna , Wong Banny S. , Rao Archana S. , Kenny Enda , Månsson Maria , Zinsmeister Alan R.
Published: April 2012

The glucagon-like peptide 1 (GLP-1) analog ROSE-010 reduced pain during acute exacerbations of irritable bowel syndrome (IBS). Our objective was to assess effects of ROSE-010 on several gastrointestinal (GI) motor and bowel functions in constipation-predominant IBS (IBS-C). In a single-center, randomized, parallel-group, double-blind, placebo-controlled, dose-response study, we evaluated safety, pharmacodynamics, and pharmacokinetics in female patients with IBS-C. ROSE-010 (30, 100, or 300 μg sc) or matching placebo was administered once daily for 3 consecutive days and on 1 day 2–10 days later. We measured GI and colonic transit by validated scintigraphy and gastric volumes by single-photon emission computed tomography. The primary end points were half time of gastric emptying of solids, colonic transit geometric center at 24 h, and gastric accommodation volume. Analysis included intent-to-treat principle, analysis of covariance (with body mass index as covariate), and Dunnett-Hsu test for multiple comparisons. Exposure to ROSE-010 was approximately dose-proportional across the dose range tested. Demographic data in four treatment groups of female IBS-C patients (total 46) were not different. Gastric emptying was significantly retarded by 100 and 300 μg of ROSE-010. There were no significant effects of ROSE-010 on gastric volumes, small bowel or colonic transit at 24 h, or bowel functions. The 30- and 100-μg doses accelerated colonic transit at 48 h. Adverse effects were nausea ( P < 0.001 vs. placebo) and vomiting ( P = 0.008 vs. placebo). Laboratory safety results were not clinically significant. In IBS-C, ROSE-010 delayed gastric emptying of solids but did not retard colonic transit or alter gastric accommodation; the accelerated colonic transit at 48 h with 30 and 100 μg of ROSE-010 suggests potential for relief of constipation in IBS-C.
Butyrate suppresses colonic inflammation through HDAC1-dependent Fas upregulation and Fas-mediated apoptosis of T cells
Authors: Zimmerman Mary A. , Singh Nagendra , Martin Pamela M. , Thangaraju Muthusamy , Ganapathy Vadivel , Waller Jennifer L. , Shi Huidong , Robertson Keith D. , Munn David H. , Liu Kebin
Published: April 2012

Butyrate, an intestinal microbiota metabolite of dietary fiber, has been shown to exhibit protective effects toward inflammatory diseases such as ulcerative colitis (UC) and inflammation-mediated colorectal cancer. Recent studies have shown that chronic IFN-γ signaling plays an essential role in inflammation-mediated colorectal cancer development in vivo, whereas genome-wide association studies have linked human UC risk loci to IFNG, the gene that encodes IFN-γ. However, the molecular mechanisms underlying the butyrate-IFN-γ-colonic inflammation axis are not well defined. Here we showed that colonic mucosa from patients with UC exhibit increased signal transducer and activator of transcription 1 (STAT1) activation, and this STAT1 hyperactivation is correlated with increased T cell infiltration. Butyrate treatment-induced apoptosis of wild-type T cells but not Fas-deficient (Faslpr) or FasL-deficient (Fasgld) T cells, revealing a potential role of Fas-mediated apoptosis of T cells as a mechanism of butyrate function. Histone deacetylase 1 (HDAC1) was found to bind to the Fas promoter in T cells, and butyrate inhibits HDAC1 activity to induce Fas promoter hyperacetylation and Fas upregulation in T cells. Knocking down gpr109a or slc5a8, the genes that encode for receptor and transporter of butyrate, respectively, resulted in altered expression of genes related to multiple inflammatory signaling pathways, including inducible nitric oxide synthase (iNOS), in mouse colonic epithelial cells in vivo. Butyrate effectively inhibited IFN-γ-induced STAT1 activation, resulting in inhibition of iNOS upregulation in human colon epithelial and carcinoma cells in vitro. Our data thus suggest that butyrate delivers a double-hit: induction of T cell apoptosis to eliminate the source of inflammation and suppression of IFN-γ-mediated inflammation in colonic epithelial cells, to suppress colonic inflammation.
Pulmonary artery pressure and cardiac function in children and adolescents after rapid ascent to 3,450 m
Authors: Allemann Yves , Stuber Thomas , de Marchi Stefano F. , Rexhaj Emrush , Sartori Claudio , Scherrer Urs , Rimoldi Stefano F.
Published: April 2012

High-altitude destinations are visited by increasing numbers of children and adolescents. High-altitude hypoxia triggers pulmonary hypertension that in turn may have adverse effects on cardiac function and may induce life-threatening high-altitude pulmonary edema (HAPE), but there are limited data in this young population. We, therefore, assessed in 118 nonacclimatized healthy children and adolescents (mean ± SD; age: 11 ± 2 yr) the effects of rapid ascent to high altitude on pulmonary artery pressure and right and left ventricular function by echocardiography. Pulmonary artery pressure was estimated by measuring the systolic right ventricular to right atrial pressure gradient. The echocardiography was performed at low altitude and 40 h after rapid ascent to 3,450 m. Pulmonary artery pressure was more than twofold higher at high than at low altitude (35 ± 11 vs. 16 ± 3 mmHg; P < 0.0001), and there existed a wide variability of pulmonary artery pressure at high altitude with an estimated upper 95% limit of 52 mmHg. Moreover, pulmonary artery pressure and its altitude-induced increase were inversely related to age, resulting in an almost twofold larger increase in the 6- to 9- than in the 14- to 16-yr-old participants (24 ± 12 vs. 13 ± 8 mmHg; P = 0.004). Even in children with the most severe altitude-induced pulmonary hypertension, right ventricular systolic function did not decrease, but increased, and none of the children developed HAPE. HAPE appears to be a rare event in this young population after rapid ascent to this altitude at which major tourist destinations are located.
Activation of HIF-1 by metallothionein contributes to cardiac protection in the diabetic heart
Authors: Xue Wanli , Liu Yanlong , Zhao Jingchan , Cai Lu , Li Xiaokun , Feng Wenke
Published: April 2012

Metallothionein (MT) protects against heavy metal-induced cellular damage and may participate in other fundamental physiological and pathological processes, such as antioxidation, proliferation, and cell survival. Previously, we have shown that elevation of MT by transgene or by induction with zinc protects the heart against diabetic cardiomyopathy by mechanisms such as antidiabetes-induced oxidative stress and inactivation of glycogen synthase kinase-3, which mediates glucose metabolism. We also reported that MT overexpression rescued the diabetic-induced reduction of hypoxia-inducible factor (HIF)-1α, which plays an important role in glucose utilization and angiogenesis. Here, we showed that overexpression of MT increased hexokinase (HK)-II expression under control conditions and attenuated diabetes-decreased HK-II expression. Glycolytic flux assay demonstrated that MT increased glycolysis output in high glucose-containing media-cultured H9c2 cells. The diabetes-induced reduction in cardiac capillaries was also attenuated by MT overexpression. Furthermore, MT induction significantly increased HIF-1 expression under both control and diabetic conditions. Moreover, in the present study, we demonstrated that MT-enhanced HIF-1α activity is likely through a mechanism of protein nuclear translocation. These results suggest that MT induces HIF-1α expression, leading to increased HK-II in the diabetic heart.
The presence of LPS in OVA inhalations affects airway inflammation and AHR but not remodeling in a rodent model of asthma
Authors: Tsuchiya Kimitake , Siddiqui Sana , Risse Paul-André , Hirota Nobuaki , Martin James G.
Published: April 2012

Ovalbumin (OVA) is the most frequently used allergen in animal models of asthma. Lipopolysaccharide (LPS) contaminating commercial OVA may modulate the evoked airway inflammatory response to OVA. However, the effect of LPS in OVA on airway remodeling, especially airway smooth muscle (ASM) has not been evaluated. We hypothesized that LPS in commercial OVA may enhance allergen-induced airway inflammation and remodeling. Brown Norway rats were sensitized with OVA on day 0. PBS, OVA, or endotoxin-free OVA (Ef-OVA) was instilled intratracheally on days 14, 19, 24. Bronchoalveolar lavage (BAL) fluid, lung, and intrathoracic lymph node tissues were collected 48 h after the last challenge. Immunohistochemistry for α-smooth muscle actin, Periodic-Acid-Schiff staining, and real-time qPCR were performed. Airway hyperresponsiveness (AHR) was also measured. BAL fluid macrophages, eosinophils, neutrophils, and lymphocytes were increased in OVA-challenged animals, and macrophages and neutrophils were significantly lower in Ef-OVA-challenged animals. The ASM area in larger airways was significantly increased in both OVA and Ef-OVA compared with PBS-challenged animals. The mRNA expression of IFN-γ and IL-13 in lung tissues and IL-4 in lymph nodes was significantly increased by both OVA and Ef-OVA compared with PBS and were not significantly different between OVA and Ef-OVA. Monocyte chemoattractant protein (MCP)-1 in BAL fluid and AHR were significantly increased in OVA but not in Ef-OVA. LPS contamination in OVA contributes to the influx of macrophages and MCP-1 increase in the airways and to AHR after OVA challenges but does not affect OVA-induced Th1 and Th2 cytokine expression, goblet cell hyperplasia, and ASM remodeling.
A critical role for increased labile zinc in reducing sensitivity of cultured sheep pulmonary artery endothelial cells to LPS-induced apoptosis
Authors: Thambiayya Kalidasan , Wasserloos Karla , Kagan Valerian E. , Stoyanovsky Detcho , Pitt Bruce R.
Published: April 2012

We previously noted an important signaling role for decreased labile intracellular zinc ([ Zn ] i) in LPS-induced apoptosis in cultured sheep pulmonary artery endothelial cells (SPAEC) (Tang ZL, Wasserloos KJ, Liu X, Stitt MS, Reynolds IJ, Pitt BR, St Croix CM. Mol Cell Biochem 234–235: 211–217, 2002; Thambiayya K, Wasserloos KJ, Huang Z, Kagan VE, St Croix CM, Pitt BR. Am J Physiol Lung Cell Mol Physiol 300: L624–632, 2011). In the present study, we used small interfering RNA (siRNA) to important contributors of zinc homeostasis [ SLC39A14 or Zrt/Irt-like protein 14 (ZIP14), a zinc importer; metallothionein (MT), a zinc binding protein ] to define molecular pathways by which extracellular zinc or nitric oxide (NO) increase labile [ Zn ] i [ e.g., zinc-sensitive fluorophore (FluoZin-3) detectable and/or chelatable by N, N, N′, N′-tetrakis(2-pyridylmethyl)ethylenediamine ] and reduce the sensitivity of SPAEC to LPS. Addition of 10 μM zinc to serum-free medium of SPAEC increased [ Zn ] i and abolished LPS-induced apoptosis (e.g., increased annexin V binding). The increase in [ Zn ] i and the protective effect of extracellular zinc were sensitive to reduction in ZIP14 expression (by siRNA), but not affected by collectively knocking down major isoforms of sheep MT (sMT-Ia, -Ib, -Ic, and -II). Pretreatment of wild-type SPAEC with 250 μM of the NO donor S-nitroso- N-acetylpenicillamine (SNAP) increased labile zinc in a relatively similar fashion to addition of extracellular zinc and reduced sensitivity of SPAEC to LPS-induced apoptosis (e.g., caspase-3/7 activation) in a N, N, N′, N′-tetrakis(2-pyridylmethyl)ethylenediamine-sensitive fashion. The antiapoptotic effects of SNAP were insensitive to siRNA knockdown of ZIP14, but were abolished (along with SNAP-induced increase in [ Zn ] i) when SPAEC were pretreated with siRNA to sheep MT. Zinc was able to directly inhibit recombinant caspase-3 activity in an in vitro assay. Collectively, these data show that increases in labile [ Zn ] i are an important component of ZIP14- or NO-mediated resistance to LPS-induced apoptosis. Cytoprotection via ZIP14 appeared to be secondary to transcellular movement of extracellular zinc, whereas NO-mediated protection was secondary to S-nitrosation of MT and redistribution of [ Zn ] i.
MAPK signaling in the quadriceps of patients with chronic obstructive pulmonary disease
Authors: Lemire Bruno B. , Debigaré Richard , Dubé Annie , Thériault Marie-Eve , Côté Claude H. , Maltais François
Published: April 2012

Muscle atrophy in chronic obstructive pulmonary disease (COPD) is associated with reduced exercise tolerance, muscle strength, and survival. The molecular mechanisms leading to muscle atrophy in COPD remain elusive. The mitogen-activated protein kinases (MAPKs) such as p38 MAPK and ERK 1/2 can increase levels of MAFbx/Atrogin and MuRF1, which are specifically involved in muscle protein degradation and atrophy. Our aim was to investigate the level of activation of p38 MAPK, ERK 1/2, and JNK in the quadriceps of patients with COPD. A biopsy of the quadriceps was obtained in 18 patients with COPD as well as in 9 healthy controls. We evaluated the phosphorylated as well as total protein levels of p38 MAPK, ERK 1/2, and JNK as well as MAFbx/Atrogin and MuRF1 in these muscle samples. The corresponding mRNA expression was also assessed by RT-PCR. Ratios of phosphorylated to total level of p38 MAPK ( P = 0.02) and ERK 1/2 ( P = 0.01) were significantly elevated in patients with COPD compared with controls. Moreover, protein levels of MAFbx/Atrogin showed a tendency to be greater in patients with COPD ( P = 0.08). mRNA expression of p38 MAPK ( P = 0.03), ERK 1/2 ( P = 0.02), and MAFbx/Atrogin ( P = 0.04) were significantly elevated in patients with COPD. In addition, phosphorylated-to-total p38 MAPK ratio (Pearson's r = −0.45; P < 0.05) and phosphorylated-to-total ERK 1/2 ratio (Pearson's r = −0.47; P < 0.05) were negatively associated with the mid-thigh muscle cross-sectional area. These data support the hypothesis that the MAPKs might play a role in the development of muscle atrophy in COPD.
Stromal Estrogen Receptor-α Promotes Tumor Growth by Normalizing an Increased Angiogenesis
Authors: Péqueux Christel , Raymond-Letron Isabelle , Blacher Silvia , Boudou Frédéric , Adlanmerini Marine , Fouque Marie-José , Rochaix Philippe , Noël Agnès , Foidart Jean-Michel , Krust Andrée , Chambon Pierre , Brouchet Laurent , Arnal Jean-François , Lenfant Françoise
Published: April 2012

Estrogens directly promote the growth of breast cancers that express the estrogen receptor α (ERα). However, the contribution of stromal expression of ERα in the tumor microenvironment to the protumoral effects of estrogen has never been explored. In this study, we evaluated the molecular and cellular mechanisms by which 17β-estradiol (E2) impacts the microenvironment and modulates tumor development of ERα-negative tumors. Using different mouse models of ER-negative cancer cells grafted subcutaneously into syngeneic ovariectomized immunocompetent mice, we found that E2 potentiates tumor growth, increases intratumoral vessel density, and modifies tumor vasculature into a more regularly organized structure, thereby improving vessel stabilization to prevent tumor hypoxia and necrosis. These E2-induced effects were completely abrogated in ERα-deficient mice, showing a critical role of host ERα. Notably, E2 did not accelerate tumor growth when ERα was deficient in Tie2-positive cells, even in mice grafted with wild-type bone marrow. These results were extended by clinical evidence of ERα-positive stromal cell labeling in the microenvironment of human breast cancers. Together, our findings therefore show that E2 promotes the growth of ERα-negative cancer cells through the activation of stromal ERα (extra-hematopoietic Tie-2 positive cells), which normalizes tumor angiogenesis and allows an adaptation of blood supply to tumors, thereby preventing hypoxia and necrosis. These findings significantly deepen mechanistic insights into the impact of E2 on tumor development with potential consequences for cancer treatment. Cancer Res; 72(12); 3010–9. ©2012 AACR.
The Proto-oncogene PKCι Regulates the Alternative Splicing of Bcl-x Pre-mRNA
Authors: Shultz Jacqueline C. , Vu Ngoc , Shultz Michael D. , Mba Mba-Uzoma U. , Shapiro Brian A. , Chalfant Charles E.
Published: April 2012

Two splice variants derived from the Bcl-x gene via alternative 5′ splice site selection (5′SS) are proapoptotic Bcl-x(s) and antiapoptotic Bcl-x(L). Previously, our laboratory showed that apoptotic signaling pathways regulated the alternative 5′SS selection via protein phosphatase-1 and de novo ceramide. In this study, we examined the elusive prosurvival signaling pathways that regulate the 5′SS selection of Bcl-x pre-mRNA in cancer cells. Taking a broad-based approach by using a number of small-molecule inhibitors of various mitogenic/survival pathways, we found that only treatment of non–small cell lung cancer (NSCLC) cell lines with the phosphoinositide 3-kinase (PI3K) inhibitor LY294002 (50 μmol/L) or the pan-protein kinase C (PKC) inhibitor Gö6983 (25 μmol/L) decreased the Bcl-x(L)/(s) mRNA ratio. Pan-PKC inhibitors that did not target the atypical PKCs, PKCι and PKCζ, had no effect on the Bcl-x(L)/(s) mRNA ratio. Additional studies showed that downregulation of the proto-oncogene, PKCι, in contrast to PKCζ, also resulted in a decrease in the Bcl-x(L)/(s) mRNA ratio. Furthermore, downregulation of PKCι correlated with a dramatic decrease in the expression of SAP155, an RNA trans-acting factor that regulates the 5′SS selection of Bcl-x pre-mRNA. Inhibition of the PI3K or atypical PKC pathway induced a dramatic loss of SAP155 complex formation at ceramide-responsive RNA cis-element 1. Finally, forced expression of Bcl-x(L) “rescued” the loss of cell survival induced by PKCι siRNA. In summary, the PI3K/PKCι regulates the alternative splicing of Bcl-x pre-mRNA with implications in the cell survival of NSCLC cells. Mol Cancer Res; 10(5); 660–9. ©2012 AACR.
Overexpression of Aurora-A Enhances Invasion and Matrix Metalloproteinase-2 Expression in Esophageal Squamous Cell Carcinoma Cells
Authors: Wang Xiaoxia , Lu Na , Niu Bo , Chen Xianjiu , Xie Jun , Cheng Niuliang
Published: April 2012

Esophageal squamous cell carcinoma (ESCC) is one of the most aggressive cancers, and metastasis is the principal cause of death in ESCC patients. It has been shown that amplification and overexpression of mitotic serine/threonine kinase Aurora-A occur in several types of human tumors, including ESCC. Moreover, increase in expression levels of Aurora-A has been predicted to correlate with the grades of tumor differentiation and invasive capability. However, the mechanisms by which Aurora-A mediates its invasive effects still remain elusive. In this article, we showed that Aurora-A overexpression significantly increased cell migration and invasion as well as secretion and expression of matrix metalloproteinase-2 (MMP-2). Conversely, siRNA-mediated knockdown of Aurora-A expression in human ESCC cells led to inhibition of cell invasiveness as well as secretion and expression of MMP-2. In addition, Aurora-A overexpression increased phosphorylation levels of p38 mitogen-activated protein kinase (MAPK) and Akt, and the knockdown of Aurora-A by siRNA decreased the activity of p38 MAPK and Akt. Moreover, the blocking of the activity of above kinases using chemical inhibitors suppressed the ability of Aurora-A to induce MMP-2 secretion and expression as well as cell invasion. These data show that overexpression of Aurora-A contributes to the malignancy development of ESCC by enhancing tumor cell invasion as well as MMP-2 activity and expression, which can occur through signaling pathways involving p38 MAPK and Akt protein kinases. Taken together, these studies provide a molecular basis for promoting the role of Aurora-A in malignancy development of ESCC. Mol Cancer Res; 10(5); 588–96. ©2012 AACR.
Genetic Evidence That Lipoprotein(a) Associates With Atherosclerotic Stenosis Rather Than Venous Thrombosis
Authors: Kamstrup Pia R. , Tybjærg­-Hansen Anne , Nordestgaard Børge G.
Published: April 2012

Objective—

The aim of the present study was to determine whether lipoprotein(a) [Lp(a)], considered a causal risk factor for cardiovascular disease, primarily promotes thrombosis or atherosclerosis.

Methods and Results—

Using a Mendelian randomization study design, we measured plasma Lp(a) and genetically elevated Lp(a) levels through the
LPA
kringle IV type 2 repeat genotype in 41231 individuals. We included 2 general population studies of both venous thrombosis and combined thrombosis and atherosclerosis in coronary arteries (=myocardial infarction), and 3 ­case–­control studies of atherosclerotic stenosis. Neither Lp(a) tertiles nor
LPA
kringle IV type 2 tertiles associated with the risk of venous thrombosis in general population studies (trend:
P
=0.12–0.76), but did each associate with risk of coronary, carotid, and femoral atherosclerotic stenosis in ­case–­control studies (trend:
P
<0.001 to 0.04). Lp(a) and
LPA
kringle IV type 2 tertiles also associated with the risk of myocardial infarction in general population studies (trend:
P
<0.001 to 0.003). For doubling of Lp(a) levels, instrumental variable estimates of hazard/odds ratios were 1.02 (95% CI 0.90–1.15) and 1.04 (0.93–1.16) for venous thrombosis in the 2 general population studies, 1.12 (1.01–1.25), 1.17 (1.05–1.32), and 1.16 (1.01–1.35), respectively, for coronary, carotid, and femoral atherosclerotic stenosis in ­case–control studies, and 1.21 (1.10–1.33) and 1.17 (1.05–1.29) for myocardial infarction in general population studies.

Conclusion—

This supports that Lp(a) primarily promotes atherosclerotic stenosis rather than venous thrombosis.