bluuebunny/crossref_metadata_2025_split huggingface dataset:
Use of mCherry Red Fluorescent Protein for Studies of Protein Localization and Gene Expression in Clostridium difficile
Authors: Ransom Eric M. , Ellermeier Craig D. , Weiss David S. 
Published: December 2014

ABSTRACT

Fluorescent proteins are powerful reporters in biology, but most require O
2
for chromophore maturation, making them inherently difficult to use in anaerobic bacteria.
Clostridium difficile
, a strict anaerobe with a genomic GC content of only 29%, is the leading cause of hospital-acquired diarrhea in developed countries, and new methods for studying this pathogen are sorely needed. We recently demonstrated that a cyan fluorescent protein called CFP
opt
that has been codon optimized for production in low-GC bacteria can be used to study protein localization in
C. difficile
provided the cells are fixed prior to exposure to air. We describe here a codon-optimized variant of mCherry (mCherryOpt) that exhibits faster acquisition of fluorescence and a better signal-to-noise ratio than CFP
opt
. We utilized
mCherryOpt
to construct plasmids for studying protein localization (pRAN473) and gene expression (pDSW1728) in
C. difficile
. Plasmid pRAN473 is an
mCherryOpt
fusion vector with a tetracycline-inducible promoter. To document its biological utility, we demonstrated septal localization of two cell division proteins, MldA and ZapA. Plasmid pDSW1728 is designed for cloning a promoter of interest upstream of
mCherryOpt
. As proof of principle, we studied the expression of the
pdaV
operon, which is required for lysozyme resistance. In confirmation and extension of previous reports, we found that expression of the
pdaV
operon requires the alternative sigma factor σ
v
and that induction by lysozyme is dose dependent and uniform across the population of lysozyme-treated cells.
Integrative Conjugative Elements Are Widespread in Field Isolates of Mycoplasma Species Pathogenic for Ruminants
Authors: Tardy Florence , Mick Virginie , Dordet-Frisoni Emilie , Marenda Marc Serge , Sirand-Pugnet Pascal , Blanchard Alain , Citti Christine 
Published: December 2014

ABSTRACT

Comparative genomics have revealed massive horizontal gene transfer (HGT) between
Mycoplasma
species sharing common ruminant hosts. Further results pointed toward an integrative conjugative element (ICE) as an important contributor of HGT in the small-ruminant-pathogen
Mycoplasma agalactiae
. To estimate the prevalence of ICEs in ruminant mycoplasmas, we surveyed their occurrence in a collection of 166 field strains representing 4 (sub)species that are recognized as major pathogens. Based on available sequenced genomes, we first defined the conserved, minimal ICE backbone as composed of 4 coding sequences (CDSs) that are evenly distributed and predicted to be essential for ICE chromosomal integration-excision and horizontal transfer. Screening of the strain collection revealed that these 4 CDSs are well represented in ruminant
Mycoplasma
species, suggesting widespread occurrence of ICEs. Yet their prevalence varies within and among species, with no correlation found with the individual strain history. Extrachromosomal ICE forms were also often detected, suggesting that ICEs are able to circularize in all species, a first and essential step in ICE horizontal transfer. Examination of the junction of the circular forms and comparative sequence analysis of conserved CDSs clearly pointed toward two types of ICE, the
hominis
and
spiroplasma
types, most likely differing in their mechanism of excision-integration. Overall, our data indicate the occurrence and maintenance of functional ICEs in a large number of field isolates of ruminant mycoplasmas. These may contribute to genome plasticity and gene exchanges and, presumably, to the emergence of diverse genotypes within pathogenic mycoplasmas of veterinary importance.
SUMOylation Is Developmentally Regulated and Required for Cell Pairing during Conjugation in Tetrahymena thermophila
Authors: Nasir Amjad M. , Yang Qianyi , Chalker Douglas L. , Forney James D. 
Published: December 2014

ABSTRACT

The covalent attachment of
s
mall
u
biquitin-like
mo
difier (SUMO) to target proteins regulates numerous nuclear events in eukaryotes, including transcription, mitosis and meiosis, and DNA repair. Despite extensive interest in nuclear pathways within the field of ciliate molecular biology, there have been no investigations of the SUMO pathway in
Tetrahymena
. The developmental program of sexual reproduction of this organism includes cell pairing, micronuclear meiosis, and the formation of a new somatic macronucleus. We identified the
Tetrahymena thermophila
SMT3
(SUMO) and
UBA2
(SUMO-activating enzyme) genes and demonstrated that the corresponding green fluorescent protein (GFP) tagged gene products are found predominantly in the somatic macronucleus during vegetative growth. Use of an anti-Smt3p antibody to perform immunoblot assays with whole-cell lysates during conjugation revealed a large increase in SUMOylation that peaked during formation of the new macronucleus. Immunofluorescence using the same antibody showed that the increase was localized primarily within the new macronucleus. To initiate functional analysis of the SUMO pathway, we created germ line knockout cell lines for both the
SMT3
and
UBA2
genes and found both are essential for cell viability. Conditional Smt3p and Uba2p cell lines were constructed by incorporation of the cadmium-inducible metallothionein promoter. Withdrawal of cadmium resulted in reduced cell growth and increased sensitivity to DNA-damaging agents. Interestingly, Smt3p and Uba2p conditional cell lines were unable to pair during sexual reproduction in the absence of cadmium, consistent with a function early in conjugation. Our studies are consistent with multiple roles for SUMOylation in
Tetrahymena
, including a dynamic regulation associated with the sexual life cycle.
Effects of lixisenatide on elevated liver transaminases: systematic review with individual patient data meta-analysis of randomised controlled trials on patients with type 2 diabetes
Authors: Gluud Lise L , Knop Filip K , Vilsbøll Tina 
Published: December 2014

ObjectiveTo evaluate the effects of the glucagon-like peptide-1 receptor agonist lixisenatide on elevated liver blood tests in patients with type 2 diabetes.DesignSystematic review.Data sourcesElectronic and manual searches were combined.Study selectionRandomised controlled trials (RCTs) on lixisenatide versus placebo or active comparators for type 2 diabetes were included.ParticipantsIndividual patient data were retrieved to calculate outcomes for patients with elevated liver blood tests.Main outcome measuresNormalisation of alanine aminotransferase (ALT) and aspartate aminotransferase (AST).Data synthesisThe results of included trials were combined in meta-analyses. Sequential, subgroup and regression analyses were performed to evaluate heterogeneity and bias.ResultsWe included 12 RCTs on lixisenatide versus placebo and 3 RCTs with the active comparators liraglutide, exenatide or sitagliptin. The mean treatment duration was 29 weeks. Lixisenatide increased the proportion of patients with normalisation of ALT (risk difference: 0.07; 95% CI 0.01 to 0.14; number needed to treat: 14 patients, p=0.042). The effect was not confirmed in sequential analysis. No effects of lixisenatide were identified on AST, alkaline phosphatase or bilirubin. No evidence of bias was identified. Mixed effect multilevel meta-regression analyses suggest that the benefit of lixisenatide on ALT was limited to patients who were overweight or obese.ConclusionsThis review suggests that lixisenatide increases the proportion of obese or overweight patients with type 2 diabetes who achieve normalisation of ALT. Additional research is needed to determine if the findings translate to clinical outcome measures.Trial registration numberPROSPERO; CRD42013005779.
Diagnostic accuracy study of three alcohol breathalysers marketed for sale to the public
Authors: Ashdown Helen F , Fleming Susannah , Spencer Elizabeth A , Thompson Matthew J , Stevens Richard J 
Published: December 2014

ObjectivesTo assess the diagnostic accuracy of three personal breathalyser devices available for sale to the public marketed to test safety to drive after drinking alcohol.DesignProspective comparative diagnostic accuracy study comparing two single-use breathalysers and one digital multiuse breathalyser (index tests) to a police breathalyser (reference test).SettingEstablishments licensed to serve alcohol in a UK city.ParticipantsOf 222 participants recruited, 208 were included in the main analysis. Participants were eligible if they were 18 years old or over, had consumed alcohol and were not intending to drive within the following 6 h.Outcome measuresSensitivity and specificity of the breathalysers for the detection of being at or over the UK legal driving limit (35 µg/100 mL breath alcohol concentration).Results18% of participants (38/208) were at or over the UK driving limit according to the police breathalyser. The digital multiuse breathalyser had a sensitivity of 89.5% (95% CI 75.9% to 95.8%) and a specificity of 64.1% (95% CI 56.6% to 71.0%). The single-use breathalysers had a sensitivity of 94.7% (95% CI 75.4% to 99.1%) and 26.3% (95% CI 11.8% to 48.8%), and a specificity of 50.6% (95% CI 40.4% to 60.7%) and 97.5% (95% CI 91.4% to 99.3%), respectively. Self-reported alcohol consumption threshold of 5 UK units or fewer had a higher sensitivity than all personal breathalysers.ConclusionsOne alcohol breathalyser had sensitivity of 26%, corresponding to false reassurance for approximately one person in four who is over the limit by the reference standard, at least on the evening of drinking alcohol. The other devices tested had 90% sensitivity or higher. All estimates were subject to uncertainty. There is no clearly defined minimum sensitivity for this safety-critical application. We conclude that current regulatory frameworks do not ensure high sensitivity for these devices marketed to consumers for a decision with potentially catastrophic consequences.
The impact of exacerbation frequency on mortality following acute exacerbations of COPD: a registry-based cohort study
Authors: Schmidt Sigrun Alba Johannesdottir , Johansen Martin Berg , Olsen Morten , Xu Xiao , Parker Joseph M , Molfino Nestor A , Lash Timothy L , Sørensen Henrik Toft , Christiansen Christian Fynbo 
Published: December 2014

ObjectiveTo examine the association between exacerbation frequency and mortality following an acute exacerbation of chronic obstructive pulmonary disease (AECOPD).DesignCohort study using medical databases.SettingNorthern Denmark.ParticipantsOn 1 January 2005, we identified all patients with prevalent hospital-diagnosed chronic obstructive pulmonary disease (COPD) who had at least one AECOPD during 1 January 2005 to 31 December 2009. We followed patients from the first AECOPD during this period until death, emigration or 31 December 2009, whichever came first. We flagged all AECOPD events during follow-up and characterised each by the exacerbation frequency (0, 1, 2 or 3+) in the prior 12-month period.Main outcomes and measuresUsing Cox regression, we computed 0–30-day and 31–365-day age-adjusted, sex-adjusted, and comorbidity-adjusted mortality rate ratios (MRRs) with 95% CIs entering exacerbation frequency as a time-varying exposure.ResultsWe identified 16 647 eligible patients with prevalent COPD, of whom 6664 (40%) developed an AECOPD and were thus included in the study cohort. The 0–30-day MRRs were 0.97 (95% CI 0.80 to 1.18), 0.90 (95% CI 0.70 to 1.15) and 1.03 (95% CI 0.81 to 1.32) among patients with AECOPD with 1, 2 and 3+ AECOPDs versus no AECOPD within the past 12 months, respectively. The corresponding MRRs were 1.47 (95% CI 1.30 to 1.66), 1.89 (95% CI 1.59 to 2.25) and 1.59 (95% CI 1.23 to 2.05) for days 31–365.ConclusionAmong patients with AECOPD, one or more exacerbations in the previous year were not associated with 30-day mortality but were associated with an increased 31–365-day mortality.
Air ambulance tasking: mechanism of injury, telephone interrogation or ambulance crew assessment?
Authors: Wilmer Ian , Chalk Graham , Davies Gareth Edward , Weaver Anne Elizabeth , Lockey David John 
Published: December 2014

ObjectiveThe identification of serious injury is critical to the tasking of air ambulances. London’s Air Ambulance (LAA) is dispatched by a flight paramedic based on mechanism of injury (MOI), paramedical interrogation of caller (INT) or land ambulance crew request (REQ).This study aimed to demonstrate which of the dispatch methods was most effective (in accuracy and time) in identifying patients with serious injury.MethodsA retrospective review of 3 years of data (to December 2010) was undertaken. Appropriate dispatch was defined as the requirement for LAA to escort the patient to hospital or for resuscitation on-scene. Inaccurate dispatch was where LAA was cancelled or left the patient in the care of the land ambulance crew. The χ2test was used to calculate p values; with significance adjusted to account for multiple testing.ResultsThere were 2203 helicopter activations analysed: MOI 18.9% (n=417), INT 62.4% (n=1375) and REQ 18.7% (n=411). Appropriate dispatch rates were MOI 58.7% (245/417), INT 69.7% (959/1375) and REQ 72.2% (297/411). INT and REQ were both significantly more accurate than MOI (p<0.0001). There was no significant difference in accuracy between INT and REQ (p=0.36). Combining MOI and INT remotely identified 80.2% of patients, with an overtriage rate of 32.8%. Mean time to dispatch (in minutes) was MOI 4, INT 8 and REQ 21.ConclusionsTelephone interrogation of the caller by a flight paramedic is as accurate as ground ambulance crew requests, and both are significantly better than MOI in identifying serious injury. Overtriage remains an issue with all methods.
Strict Requirement for Vector-Induced Type I Interferon in Efficacious Antitumor Responses to Virally Encoded IL12
Authors: Melero Ignacio , Quetglas Jose I. , Reboredo Mercedes , Dubrot Juan , Rodriguez-Madoz Juan R. , Mancheño Uxua , Casales Erkuden , Riezu-Boj Jose I. , Ruiz-Guillen Marta , Ochoa Maria C. , Sanmamed Miguel F. , Thieblemont Nathalie , Smerdou Cristian , Hervas-Stubbs Sandra 
Published: December 2014

AbstractHost responses are increasingly considered important for the efficacious response to experimental cancer therapies that employ viral vectors, but little is known about the specific nature of host responses required. In this study, we investigated the role of host type I interferons (IFN-I) in the efficacy of virally delivered therapeutic genes. Specifically, we used a Semliki Forest virus encoding IL12 (SFV-IL12) based on its promise as an RNA viral vector for cancer treatment. Intratumoral injection of SFV-IL12 induced production of IFN-I as detected in serum. IFN-I production was abolished in mice deficient for the IFNβ transcriptional regulator IPS-1 and partially attenuated in mice deficient for the IFNβ signaling protein TRIF. Use of bone marrow chimeric hosts established that both hematopoietic and stromal cells were involved in IFN-I production. Macrophages, plasmacytoid, and conventional dendritic cells were each implicated based on cell depletion experiments. Further, mice deficient in the IFN-I receptor (IFNAR) abolished the therapeutic activity of SFV-IL12, as did a specific antibody-mediated blockade of IFNAR signaling. Reduced efficacy was not caused by an impairment in IL12 expression, because IFNAR-deficient mice expressed the viral IL12 transgene even more strongly than wild-type (WT) hosts. Chimeric host analysis for the IFNAR involvement established a strict requirement in hematopoietic cells. Notably, although tumor-specific CD8 T lymphocytes expanded robustly after intratumoral injection of WT mice with SFV-IL12, this did not occur in mice where IFNAR was inactivated genetically or pharmacologically. Overall, our results argued that the antitumor efficacy of a virally based transgene therapeutic relied strongly on a vector-induced IFN-I response, revealing an unexpected mechanism of action that is relevant to a broad array of current translational products in cancer research. Cancer Res; 75(3); 497–507. ©2014 AACR.
Physical Activity and Survival among Men Diagnosed with Prostate Cancer
Authors: Bonn Stephanie E. , Sjölander Arvid , Lagerros Ylva Trolle , Wiklund Fredrik , Stattin Pär , Holmberg Erik , Grönberg Henrik , Bälter Katarina 
Published: December 2014

AbstractBackground: Few studies have investigated the association between post-diagnosis physical activity and mortality among men diagnosed with prostate cancer. The aim of this study was to investigate the effect of physical activity after a prostate cancer diagnosis on both overall and prostate cancer–specific mortality in a large cohort.Methods: Data from 4,623 men diagnosed with localized prostate cancer 1997–2002 and followed-up until 2012 were analyzed. HRs with 95% confidence intervals (CI) were estimated using Cox proportional hazards models to examine the association between post-diagnosis recreational MET-h/d, time spent walking/bicycling, performing household work or exercising, and time to overall and prostate cancer–specific death. All models were adjusted for potential confounders.Results: During the follow-up, 561 deaths of any cause and 194 deaths from prostate cancer occurred. Statistically significantly lower overall mortality rates were found among men engaged in ≥5 recreational MET-h/d (HR, 0.63; 95% CI, 0.52–0.77), walking/bicycling ≥20 min/d (HR, 0.70; 95% CI, 0.57–0.86), performing household work ≥1 h/d (HR, 0.71; 95% CI, 0.59–0.86), or exercising ≥1 h/wk (HR, 0.74; 95% CI, 0.61–0.90), compared with less active men within each activity type. For prostate cancer–specific mortality, statistically significantly lower mortality rates were seen among men walking/bicycling ≥20 min/d (HR, 0.61; 95% CI, 0.43–0.87) or exercising ≥1 h/wk (HR, 0.68; 95% CI, 0.48–0.94).Conclusions: Higher levels of physical activity were associated with reduced rates of overall and prostate cancer–specific mortality.Impact: Our study further strengthens previous results indicating beneficial effects of physical activity on survival among men with prostate cancer. Cancer Epidemiol Biomarkers Prev; 24(1); 57–64. ©2014 AACR.
Inhibition of the GTPase Rac1 Mediates the Antimigratory Effects of Metformin in Prostate Cancer Cells
Authors: Dirat Béatrice , Ader Isabelle , Golzio Muriel , Massa Fabienne , Mettouchi Amel , Laurent Kathiane , Larbret Frédéric , Malavaud Bernard , Cormont Mireille , Lemichez Emmanuel , Cuvillier Olivier , Tanti Jean François , Bost Frédéric 
Published: December 2014

Abstract
Cell migration is a critical step in the progression of prostate cancer to the metastatic state, the lethal form of the disease. The antidiabetic drug metformin has been shown to display antitumoral properties in prostate cancer cell and animal models; however, its role in the formation of metastases remains poorly documented. Here, we show that metformin reduces the formation of metastases to fewer solid organs in an orthotopic metastatic prostate cancer cell model established in nude mice. As predicted, metformin hampers cell motility in PC3 and DU145 prostate cancer cells and triggers a radical reorganization of the cell cytoskeleton. The small GTPase Rac1 is a master regulator of cytoskeleton organization and cell migration. We report that metformin leads to a major inhibition of Rac1 GTPase activity by interfering with some of its multiple upstream signaling pathways, namely P-Rex1 (a Guanine nucleotide exchange factor and activator of Rac1), cAMP, and CXCL12/CXCR4, resulting in decreased migration of prostate cancer cells. Importantly, overexpression of a constitutively active form of Rac1, or P-Rex, as well as the inhibition of the adenylate cyclase, was able to reverse the antimigratory effects of metformin. These results establish a novel mechanism of action for metformin and highlight its potential antimetastatic properties in prostate cancer. Mol Cancer Ther; 14(2); 586–96. ©2014 AACR.
TRKing Down an Old Oncogene in a New Era of Targeted Therapy
Authors: Vaishnavi Aria , Le Anh T. , Doebele Robert C. 
Published: December 2014

Abstract
The use of high-throughput next-generation sequencing techniques in multiple tumor types during the last few years has identified NTRK1, 2, and 3 gene rearrangements encoding novel oncogenic fusions in 19 different tumor types to date. These recent developments have led us to revisit an old oncogene, Trk (originally identified as OncD), which encodes the TPM3–NTRK1 gene fusion and was one of the first transforming chromosomal rearrangements identified 32 years ago. However, no drug has yet been approved by the FDA for cancers harboring this oncogene. This review will discuss the biology of the TRK family of receptors, their role in human cancer, the types of oncogenic alterations, and drugs that are currently in development for this family of oncogene targets.
Significance: Precision oncology approaches have accelerated recently due to advancements in our ability to detect oncogenic mutations in tumor samples. Oncogenic alterations, most commonly gene fusions, have now been detected for the genes encoding the TRKA, TRKB, and TRKC receptor tyrosine kinases across multiple tumor types. The scientific rationale for the targeting of the TRK oncogene family will be discussed here. Cancer Discov; 5(1); 25–34. ©2014 AACR.
B7-H4 Expression by Nonhematopoietic Cells in the Tumor Microenvironment Promotes Antitumor Immunity
Authors: Rahbar Ramtin , Lin Albert , Ghazarian Magar , Yau Helen-Loo , Paramathas Sangeetha , Lang Philipp A. , Schildknecht Anita , Elford Alisha R. , Garcia-Batres Carlos , Martin Bernard , Berman Hal K. , Leong Wey L. , McCready David R. , Reedijk Michael , Done Susan J. , Miller Naomi , Youngson Bruce , Suh Woong-Kyung , Mak Tak W. , Ohashi Pamela S. 
Published: December 2014

Abstract
The B7 family plays a critical role in both positive and negative regulation of immune responses by engaging a variety of receptors on lymphocytes. Importantly, blocking coinhibitory molecules using antibodies specific for CTLA-4 and PD-1 enhances tumor immunity in a subset of patients. Therefore, it is critical to understand the role of different B7 family members since they may be suitable therapeutic targets. B7-H4 is another member that inhibits T-cell function, and it is also upregulated on a variety of tumors and has been proposed to promote tumor growth. Here, we investigate the role of B7-H4 in tumor development and show that B7-H4 expression inhibits tumor growth in two mouse models. Furthermore, we show that B7-H4 expression is required for antitumor immune responses in a mouse model of mammary tumorigenesis. We found that the expression levels of B7-H4 correlate with MHC class I expression in both mouse and human samples. We show that IFNγ upregulates B7-H4 expression on mouse embryo fibroblasts and that the upregulation of B7-H4 on tumors is dependent on T cells. Notably, patients with breast cancer with increased B7-H4 expression show a prolonged time to recurrence. These studies demonstrate a positive role for B7-H4 in promoting antitumor immunity. Cancer Immunol Res; 3(2); 184–95. ©2014 AACR.