bluuebunny/crossref_metadata_2025 huggingface dataset:
Catalytic asymmetric addition of Grignard reagents to alkenyl-substituted aromatic
N
-heterocycles
Authors: Jumde Ravindra P. , Lanza Francesco , Veenstra Marieke J. , Harutyunyan Syuzanna R.
Published: April 2016

Copper adds alkyls asymmetrically

Nitrogen-bearing rings are very common features in the molecular structures of modern drugs. Reactions that can modify these N heterocycles selectively are thus especially useful to optimize pharmaceutical properties. Jumde
et al.
developed a method to append alkyl groups in a single mirror-image orientation to substituted C=C double bonds dangling from N heterocycles. The copper-catalyzed reaction, which relies on Grignard reagents to introduce the alkyl groups, manifests high selectivity across a broad range of substrates, with no interference from the nitrogen.

Science
, this issue p.
433
Waste not, want not, emit less
Authors: Aschemann-Witzel Jessica
Published: April 2016

Reducing food waste in the supply chain and at home can help to reduce carbon emissions
A monumental challenge
Cleopatra's Needles The Lost Obelisks of Egypt
Bob Brier
Bloomsbury Academic, 2016. 238 pp.
Authors: Robinson Andrew
Published: April 2016

Erecting an Egyptian obelisk is no small feat
Electrons catch a terahertz wave
Authors: Ropers Claus
Published: April 2016

Far-infrared fields control ultrashort electron pulses

[Also see Research Article by

Kealhofer
et al.

]
Due process in the Twitter age
Authors: McNutt Marcia
Published: April 2016

Until recently, the only insight that the research community had into published papers that might be suspect, and thus candidates for retraction, was through the formal investigations by institutions or funders and the official announcements made by journals. Today, online communities such as PubPeer and Retraction Watch could be the first to raise an alert that a paper may require additional scrutiny. The involvement of such online entities has made the standardization of processes to address allegations more complex and has led to less patience from the scientific community and the public with what are often long timelines in institutional misconduct investigations. Editors are caught in the middle: They want to correct the literature as quickly as possible to avoid misleading readers with flawed information, but they also want to ensure that authors have received due process, even as rumors of scientific misconduct may be amplified through social media. Last month, a Journals Summit convened by the U.S. National Academy of Sciences tackled this topic in the larger context of research integrity, and several solutions emerged.
Clinical Impact of Laboratory Implementation of Verigene BC-GN Microarray-Based Assay for Detection of Gram-Negative Bacteria in Positive Blood Cultures
Authors: Walker Tamar , Dumadag Sandrea , Lee Christine Jiyoun , Lee Seung Heon , Bender Jeffrey M. , Cupo Abbott Jennifer , She Rosemary C.
Published: April 2016

Gram-negative bacteremia is highly fatal, and hospitalizations due to sepsis have been increasing worldwide. Molecular tests that supplement Gram stain results from positive blood cultures provide specific organism information to potentially guide therapy, but more clinical data on their real-world impact are still needed. We retrospectively reviewed cases of Gram-negative bacteremia in hospitalized patients over a 6-month period before (
n =
98) and over a 6-month period after (
n =
97) the implementation of a microarray-based early identification and resistance marker detection system (Verigene BC-GN; Nanosphere) while antimicrobial stewardship practices remained constant. Patient demographics, time to organism identification, time to effective antimicrobial therapy, and other key clinical parameters were compared. The two groups did not differ statistically with regard to comorbid conditions, sources of bacteremia, or numbers of intensive care unit (ICU) admissions, active use of immunosuppressive therapy, neutropenia, or bacteremia due to multidrug-resistant organisms. The BC-GN panel yielded an identification in 87% of Gram-negative cultures and was accurate in 95/97 (98%) of the cases compared to results using conventional culture. Organism identifications were achieved more quickly post-microarray implementation (mean, 10.9 h versus 37.9 h;
P
< 0.001). Length of ICU stay, 30-day mortality, and mortality associated with multidrug-resistant organisms were significantly lower in the postintervention group (
P
< 0.05). More rapid implementation of effective therapy was statistically significant for postintervention cases of extended-spectrum beta-lactamase-producing organisms (
P
= 0.049) but not overall (
P
= 0.12). The Verigene BC-GN assay is a valuable addition for the early identification of Gram-negative organisms that cause bloodstream infections and can significantly impact patient care, particularly when resistance markers are detected.
Evaluation of Various Culture Media for Detection of Rapidly Growing Mycobacteria from Patients with Cystic Fibrosis
Authors: Preece Clair L. , Wichelhaus Thomas A. , Perry Audrey , Jones Amanda L. , Cummings Stephen P. , Perry John D. , Hogardt Michael
Published: April 2016

Isolation of nontuberculous mycobacteria (NTM) from the sputum of patients with cystic fibrosis (CF) is challenging due to overgrowth by rapidly growing species that colonize the lungs of patients with CF. Extended incubation onBurkholderia cepaciaselective agar (BCSA) has been recommended as an expedient culture method for the isolation of rapidly growing NTM in this setting. The aim of this study was to assess five selective media designed for the isolation ofBurkholderia cepaciacomplex, along with two media designed for the isolation of mycobacteria (rapidly growing mycobacteria [RGM] medium and Middlebrook 7H11 agar), for their abilities to isolate NTM. All seven media were challenged with 147 isolates of rapidly growing mycobacteria and 185 isolates belonging to other species. RGM medium was then compared with the most selective brand of BCSA for the isolation of NTM from 224 sputum samples from patients with CF. Different agars designed for the isolation ofB. cepaciacomplex varied considerably in their inhibition of other bacteria and fungi. RGM medium supported the growth of all isolates of mycobacteria and was more selective than any other medium. NTM were recovered from 17 of 224 sputum samples using RGM medium, compared with only 7 samples using the most selective brand of BCSA (P= 0.023). RGM medium offers a superior option, compared to other selective agars, for the isolation of rapidly growing mycobacteria from the sputum of patients with CF. Furthermore, the convenience of using RGM medium enables routine screening for rapidly growing NTM in all submitted sputum samples from patients with CF.
RNA-Free and Ribonucleoprotein-Associated Influenza Virus Polymerases Directly Bind the Serine-5-Phosphorylated Carboxyl-Terminal Domain of Host RNA Polymerase II
Authors: Martínez-Alonso Mónica , Hengrung Narin , Fodor Ervin
Published: April 2016

Influenza viruses subvert the transcriptional machinery of their hosts to synthesize their own viral mRNA. Ongoing transcription by cellular RNA polymerase II (Pol II) is required for viral mRNA synthesis. By a process known as cap snatching, the virus steals short 5′ capped RNA fragments from host capped RNAs and uses them to prime viral transcription. An interaction between the influenza A virus RNA polymerase and the C-terminal domain (CTD) of the large subunit of Pol II has been established, but the molecular details of this interaction remain unknown. We show here that the influenza virus ribonucleoprotein (vRNP) complex binds to the CTD of transcriptionally engaged Pol II. Furthermore, we provide evidence that the viral polymerase binds directly to the serine-5-phosphorylated form of the Pol II CTD, both in the presence and in the absence of viral RNA, and show that this interaction is conserved in evolutionarily distant influenza viruses. We propose a model in which direct binding of the viral RNA polymerase in the context of vRNPs to Pol II early in infection facilitates cap snatching, while we suggest that binding of free viral polymerase to Pol II late in infection may trigger Pol II degradation.

IMPORTANCE
Influenza viruses cause yearly epidemics and occasional pandemics that pose a threat to human health, as well as represent a large economic burden to health care systems globally. Existing vaccines are not always effective, as they may not exactly match the circulating viruses. Furthermore, there are a limited number of antivirals available, and development of resistance to these is a concern. New measures to combat influenza are needed, but before they can be developed, it is necessary to better understand the molecular interactions between influenza viruses and their host cells. By providing further insights into the molecular details of how influenza viruses hijack the host transcriptional machinery, we aim to uncover novel targets for the development of antivirals.
HIV-1 Protease, Reverse Transcriptase, and Integrase Variation
Authors: Rhee Soo-Yon , Sankaran Kris , Varghese Vici , Winters Mark A. , Hurt Christopher B. , Eron Joseph J. , Parkin Neil , Holmes Susan P. , Holodniy Mark , Shafer Robert W.
Published: April 2016

HIV-1 protease (PR), reverse transcriptase (RT), and integrase (IN) variability presents a challenge to laboratories performing genotypic resistance testing. This challenge will grow with increased sequencing of samples enriched for proviral DNA such as dried blood spots and increased use of next-generation sequencing (NGS) to detect low-abundance HIV-1 variants. We analyzed PR and RT sequences from >100,000 individuals and IN sequences from >10,000 individuals to characterize variation at each amino acid position, identify mutations indicating APOBEC-mediated G-to-A editing, and identify mutations resulting from selective drug pressure. Forty-seven percent of PR, 37% of RT, and 34% of IN positions had one or more amino acid variants with a prevalence of ≥1%. Seventy percent of PR, 60% of RT, and 60% of IN positions had one or more variants with a prevalence of ≥0.1%. Overall 201 PR, 636 RT, and 346 IN variants had a prevalence of ≥0.1%. The median intersubtype prevalence ratios were 2.9-, 2.1-, and 1.9-fold for these PR, RT, and IN variants, respectively. Only 5.0% of PR, 3.7% of RT, and 2.0% of IN variants had a median intersubtype prevalence ratio of ≥10-fold. Variants at lower prevalences were more likely to differ biochemically and to be part of an electrophoretic mixture compared to high-prevalence variants. There were 209 mutations indicative of APOBEC-mediated G-to-A editing and 326 mutations nonpolymorphic treatment selected. Identification of viruses with a high number of APOBEC-associated mutations will facilitate the quality control of dried blood spot sequencing. Identifying sequences with a high proportion of rare mutations will facilitate the quality control of NGS.

IMPORTANCEMost antiretroviral drugs target three HIV-1 proteins: PR, RT, and IN. These proteins are highly variable: many different amino acids can be present at the same position in viruses from different individuals. Some of the amino acid variants cause drug resistance and occur mainly in individuals receiving antiretroviral drugs. Some variants result from a human cellular defense mechanism called APOBEC-mediated hypermutation. Many variants result from naturally occurring mutation. Some variants may represent technical artifacts. We studied PR and RT sequences from >100,000 individuals and IN sequences from >10,000 individuals to quantify variation at each amino acid position in these three HIV-1 proteins. We performed analyses to determine which amino acid variants resulted from antiretroviral drug selection pressure, APOBEC-mediated editing, and naturally occurring variation. Our results provide information essential to clinical, research, and public health laboratories performing genotypic resistance testing by sequencing HIV-1 PR, RT, and IN.
Role of WDHD1 in Human Papillomavirus-Mediated Oncogenesis Identified by Transcriptional Profiling of E7-Expressing Cells
Authors: Zhou Yunying , Zhang Qishu , Gao Ge , Zhang Xiaoli , Liu Yafei , Yuan Shoudao , Wang Xiaowei , Chen Jason J.
Published: April 2016

The E7 oncoprotein of the high-risk human papillomavirus (HPV) plays a major role in HPV-induced carcinogenesis. E7 abrogates the G
1
cell cycle checkpoint and induces genomic instability, but the mechanism is not fully understood. In this study, we performed RNA sequencing (RNA-seq) to characterize the transcriptional profile of keratinocytes expressing HPV 16 (HPV-16) E7. At the transcriptome level, 236 genes were differentially expressed between E7 and vector control cells. A subset of the differentially expressed genes, most of them novel to E7-expressing cells, was further confirmed by real-time PCR. Of interest, the activities of multiple transcription factors were altered in E7-expressing cells. Through bioinformatics analysis, pathways altered in E7-expressing cells were investigated. The upregulated genes were enriched in cell cycle and DNA replication, as well as in the DNA metabolic process, transcription, DNA damage, DNA repair, and nucleotide metabolism. Specifically, we focused our studies on the gene encoding WDHD1 (WD repeat and high mobility group [HMG]-box DNA-binding protein), one of the genes that was upregulated in E7-expressing cells. WDHD1 is a component of the replisome that regulates DNA replication. Recent studies suggest that WDHD1 may also function as a DNA replication initiation factor as well as a G
1
checkpoint regulator. We found that in E7-expressing cells, the steady-state level of WDHD1 protein was increased along with the half-life. Moreover, downregulation of WDHD1 reduced E7-induced G
1
checkpoint abrogation and rereplication, demonstrating a novel function for WDHD1. These studies shed light on mechanisms by which HPV induces genomic instability and have therapeutic implications.

IMPORTANCE
The high-risk HPV types induce cervical cancer and encode an E7 oncoprotein that plays a major role in HPV-induced carcinogenesis. However, the mechanism by which E7 induces carcinogenesis is not fully understood; specific anti-HPV agents are not available. In this study, we performed RNA-seq to characterize transcriptional profiling of keratinocytes expressing HPV-16 E7 and identified more than 200 genes that were differentially expressed between E7 and vector control cells. Through bioinformatics analysis, pathways altered in E7-expressing cells were identified. Significantly, the WDHD1 gene, one of the genes that is upregulated in E7-expressing cells, was found to play an important role in E7-induced G
1
checkpoint abrogation and rereplication. These studies shed light on mechanisms by which HPV induces genomic instability and have therapeutic implications.
Discovery of a Coregulatory Interaction between Kaposi's Sarcoma-Associated Herpesvirus ORF45 and the Viral Protein Kinase ORF36
Authors: Avey Denis , Tepper Sarah , Pifer Benjamin , Bahga Amritpal , Williams Hunter , Gillen Joseph , Li Wenwei , Ogden Sarah , Zhu Fanxiu
Published: April 2016

Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of three human malignancies. KSHV ORF36 encodes a serine/threonine viral protein kinase, which is conserved throughout all herpesviruses. Although several studies have identified the viral and cellular substrates of conserved herpesvirus protein kinases (CHPKs), the precise functions of KSHV ORF36 during lytic replication remain elusive. Here, we report that ORF36 interacts with another lytic protein, ORF45, in a manner dependent on ORF36 kinase activity. We mapped the regions of ORF36 and ORF45 involved in the binding. Their association appears to be mediated by electrostatic interactions, since deletion of either the highly basic N terminus of ORF36 or an acidic patch of ORF45 abolished the binding. In addition, the dephosphorylation of ORF45 protein dramatically reduced its association with ORF36. Importantly, ORF45 enhances both the stability and kinase activity of ORF36. Consistent with previous studies of CHPK homologs, we detected ORF36 protein in extracellular virions. To investigate the roles of ORF36 in the context of KSHV lytic replication, we used bacterial artificial chromosome mutagenesis to engineer both ORF36-null and kinase-dead mutants. We found that ORF36-null/mutant virions are moderately defective in viral particle production and are further deficient in primary infection. In summary, our results uncover a functionally important interaction between ORF36 and ORF45 and indicate a significant role of ORF36 in the production of infectious progeny virions.

IMPORTANCE
Kaposi's sarcoma-associated herpesvirus (KSHV) is a human tumor virus with a significant public health burden. KSHV ORF36 encodes a serine/threonine viral protein kinase, whose functions throughout the viral life cycle have not been elucidated. Here, we report that ORF36 interacts with another KSHV protein, ORF45. We mapped the regions of ORF36 and ORF45 involved in their association and further characterized the consequences of this interaction. We engineered ORF36 mutant viruses in order to investigate the functional roles of ORF36 in the context of KSHV lytic replication, and we confirmed that ORF36 is a component of KSHV virions. Moreover, we found that ORF36 mutants are defective in virion production and primary infection. In summary, we discovered and characterized a functionally important interaction between KSHV ORF36 and ORF45, and our results suggest a significant role of ORF36 in the production of infectious progeny virions, a process critical for KSHV pathogenesis.
Rotavirus NSP1 Associates with Components of the Cullin RING Ligase Family of E3 Ubiquitin Ligases
Authors: Lutz Lindy M. , Pace Chandler R. , Arnold Michelle M.
Published: April 2016

The rotavirus nonstructural protein NSP1 acts as an antagonist of the host antiviral response by inducing degradation of key proteins required to activate interferon (IFN) production. Protein degradation induced by NSP1 is dependent on the proteasome, and the presence of a RING domain near the N terminus has led to the hypothesis that NSP1 is an E3 ubiquitin ligase. To examine this hypothesis, pulldown assays were performed, followed by mass spectrometry to identify components of the host ubiquitination machinery that associate with NSP1. Multiple components of cullin RING ligases (CRLs), which are essential multisubunit ubiquitination complexes, were identified in association with NSP1. The mass spectrometry was validated in both transfected and infected cells to show that the NSP1 proteins from different strains of rotavirus associated with key components of CRL complexes, most notably the cullin scaffolding proteins Cul3 and Cul1.
In vitro
binding assays using purified proteins confirmed that NSP1 specifically interacted with Cul3 and that the N-terminal region of Cul3 was responsible for binding to NSP1. To test if NSP1 used CRL3 to induce degradation of the target protein IRF3 or β-TrCP, Cul3 levels were knocked down using a small interfering RNA (siRNA) approach. Unexpectedly, loss of Cul3 did not rescue IRF3 or β-TrCP from degradation in infected cells. The results indicate that, rather than actively using CRL complexes to induce degradation of target proteins required for IFN production, NSP1 may use cullin-containing complexes to prevent another cellular activity.

IMPORTANCE
The ubiquitin-proteasome pathway plays an important regulatory role in numerous cellular functions, and many viruses have evolved mechanisms to exploit or manipulate this pathway to enhance replication and spread. Rotavirus, a major cause of severe gastroenteritis in young children that causes approximately 420,000 deaths worldwide each year, utilizes the ubiquitin-proteasome system to subvert the host innate immune response by inducing the degradation of key components required for the production of interferon (IFN). Here, we show that NSP1 proteins from different rotavirus strains associate with the scaffolding proteins Cul1 and Cul3 of CRL ubiquitin ligase complexes. Nonetheless, knockdown of Cul1 and Cul3 suggests that NSP1 induces the degradation of some target proteins independently of its association with CRL complexes, stressing a need to further investigate the mechanistic details of how NSP1 subverts the host IFN response.
Impact of Ammonium on Syntrophic Organohalide-Respiring and Fermenting Microbial Communities
Authors: Delgado Anca G. , Fajardo-Williams Devyn , Kegerreis Kylie L. , Parameswaran Prathap , Krajmalnik-Brown Rosa
Published: April 2016

Contamination with ammonium and chlorinated solvents has been reported in numerous subsurface environments, and these chemicals bring significant challenges for
in situ
bioremediation.
Dehalococcoides mccartyi
is able to reduce the chlorinated solvent trichloroethene to the nontoxic end product ethene. Fermentative bacteria are of central importance for organohalide respiration and bioremediation to provide
D. mccartyi
with H
2
, their electron donor, acetate, their carbon source, and other micronutrients. In this study, we found that high concentrations of ammonium negatively correlated with rates of trichloroethene reductive dehalogenation and fermentation. However, detoxification of trichloroethene to nontoxic ethene occurred even at ammonium concentrations typical of those found in animal waste (up to 2 g liter
−1
NH
4
+
-N). To date, hundreds of subsurface environments have been bioremediated through the unique metabolic capability of
D. mccartyi
. These findings extend our knowledge of
D. mccartyi
and provide insight for bioremediation of sites contaminated with chlorinated solvents and ammonium.
Mouse Adenovirus Type 1 Early Region 1A Effects on the Blood-Brain Barrier
Authors: Tirumuru Nagaraja , Pretto Carla D. , Castro Jorge Luiza A. , Spindler Katherine R.
Published: April 2016

Encephalitis can be caused by viruses, and it is potentially life-threatening because of the vital nature of the brain and the lack of treatment options. MAV-1 produces viral encephalitis in its natural host, providing a model for investigating factors involved in development of encephalitis. MAV-1 infection disrupts the BBB and increases activity of matrix metalloproteinases in brains of infected mice. We investigated whether the major transcriptional regulator of adenoviruses, E1A protein, is responsible for any of the specific phenotypes that result from MAV-1 infection. For some of the functions assayed, an E1A mutant virus behaved like wild-type virus. However, expression of mRNA for one matrix metalloproteinase was higher in the virus lacking E1A protein production. This highlights the complex nature of encephalitis and suggests that E1A may have transcriptional effects on host genes important for the development of encephalitis.
Primary care randomised controlled trial of a tailored interactive website for the self-management of respiratory infections (Internet Doctor)
Authors: Little Paul , Stuart Beth , Andreou Panayiota , McDermott Lisa , Joseph Judith , Mullee Mark , Moore Mike , Broomfield Sue , Thomas Tammy , Yardley Lucy
Published: April 2016

Objective

To assess an internet-delivered intervention providing advice to manage respiratory tract infections (RTIs).

Design

Open pragmatic parallel group randomised controlled trial.

Setting

Primary care in UK.

Participants

Adults (aged ≥18) registered with general practitioners, recruited by postal invitation.

Intervention

Patients were randomised with computer-generated random numbers to access the intervention website (intervention) or not (control). The intervention tailored advice about the diagnosis, natural history, symptom management (particularly paracetamol/ibuprofen use) and when to seek further help.

Outcomes

Primary: National Health Service (NHS) contacts for those reporting RTIs from monthly online questionnaires for 20 weeks. Secondary: hospitalisations; symptom duration/severity.

Results

3044 participants were recruited. 852 in the intervention group and 920 in the control group reported 1 or more RTIs, among whom there was a modest increase in NHS direct contacts in the intervention group (intervention 37/1574 (2.4%) versus control 20/1661 (1.2%); multivariate risk ratio (RR) 2.25 (95% CI 1.00 to 5.07, p=0.048)). Conversely, reduced contact with doctors occurred (239/1574 (15.2%) vs 304/1664 (18.3%); RR 0.71, 0.52 to 0.98, p=0.037). Reduction in contacts occurred despite slightly longer illness duration (11.3 days vs 10.7 days, respectively; multivariate estimate 0.60 days longer (−0.15 to 1.36, p=0.118) and more days of illness rated moderately bad or worse illness (0.52 days; 0.06 to 0.97, p=0.026). The estimate of slower symptom resolution in the intervention group was attenuated when controlling for whether individuals had used web pages which advocated ibuprofen use (length of illness 0.22 days, −0.51 to 0.95, p=0.551; moderately bad or worse symptoms 0.36 days, −0.08 to 0.80, p=0.105). There was no evidence of increased hospitalisations (risk ratio 0.25; 0.05 to 1.12; p=0.069).

Conclusions

An internet-delivered intervention for the self-management of RTIs modifies help-seeking behaviour, and does not result in more hospital admissions due to delayed help seeking. Advising the use of ibuprofen may not be helpful.

Trial registration number

ISRCTN91518452.
Non-invasive versus invasive management in patients with prior coronary artery bypass surgery with a non-ST segment elevation acute coronary syndrome: study design of the pilot randomised controlled trial and registry (CABG-ACS)
Authors: Lee Matthew M Y , Petrie Mark C , Rocchiccioli Paul , Simpson Joanne , Jackson Colette , Brown Ammani , Corcoran David , Mangion Kenneth , McEntegart Margaret , Shaukat Aadil , Rae Alan , Hood Stuart , Peat Eileen , Findlay Iain , Murphy Clare , Cormack Alistair , Bukov Nikolay , Balachandran Kanarath , Papworth Richard , Ford Ian , Briggs Andrew , Berry Colin
Published: April 2016

Introduction

There is an evidence gap about how to best treat patients with prior coronary artery bypass grafts (CABGs) presenting with non-ST segment elevation acute coronary syndromes (NSTE-ACS) because historically, these patients were excluded from pivotal randomised trials. We aim to undertake a pilot trial of routine non-invasive management versus routine invasive management in patients with NSTE-ACS with prior CABG and optimal medical therapy during routine clinical care. Our trial is a proof-of-concept study for feasibility, safety, potential efficacy and health economic modelling. We hypothesise that a routine invasive approach in patients with NSTE-ACS with prior CABG is not superior to a non-invasive approach with optimal medical therapy.

Methods and analysis

60 patients will be enrolled in a randomised clinical trial in 4 hospitals. A screening log will be prospectively completed. Patients not randomised due to lack of eligibility criteria and/or patient or physician preference and who give consent will be included in a registry. We will gather information about screening, enrolment, eligibility, randomisation, patient characteristics and adverse events (including post-discharge). The primary efficacy outcome is the composite of all-cause mortality, rehospitalisation for refractory ischaemia/angina, myocardial infarction and hospitalisation for heart failure. The primary safety outcome is the composite of bleeding, stroke, procedure-related myocardial infarction and worsening renal function. Health status will be assessed using EuroQol 5 Dimensions (EQ-5D) assessed at baseline and 6 monthly intervals, for at least 18 months.

Trial registration number

NCT01895751 (ClinicalTrials.gov).
Mahogunin-mediated regulation of Gαi localisation during mitosis and its effect on spindle positioning
Authors: Srivastava Devika , Mukherjee Rukmini , Mookherjee Debdatto , Chakrabarti Oishee
Published: April 2016

Mahogunin RING Finger 1 (MGRN1) is a ubiquitin E3 ligase known to affect spindle tilt in mitotic cells by regulating α-tubulin ubiquitination and polymerization. In cell culture systems we have found that expressing truncated mutants of MGRN1 leads to various other mitotic anomalies, such as lateral and angular spindle displacements. This seems to be independent of the MGRN1 ligase activity. Our experiments suggest that MGRN1 regulates the balance between the lower molecular weight monomeric Gαi and larger trimeric G-protein complex, along with its abundance in the ternary complex that regulates spindle positioning. The cytosolic isoforms of MGRN1 lead to the enrichment of monomeric Gαi in the cytosol and its subsequent recruitment at the plasma membrane. Excess Gαi at the cell cortex results in an imbalance in the assembly of the ternary complex regulating spindle positioning during mitosis. These observations seem independent of the ligase activity of MGRN1, although we cannot exclude the involvement of an intermediate player that acts as a substrate for MGRN1, and in turn, regulates Gαi.
SPAIN AND THE CRISIS: HOUSING PRICES, CREDIT AND MACROPRUDENTIAL POLICIES
Authors: RUBIO MARGARITA , CARRASCO-GALLEGO JOSÉ A.
Published: April 2016

In this paper, we propose a two-country, two sector monetary union DSGE model with housing. One of the countries is calibrated to represent the Spanish economy while the other one is the rest of the European monetary union. First, we illustrate how looser credit conditions coming from the Euro area, together with increases in housing demand, lead to an increase in house prices and credit in Spain. Then, we analyze to what extent, macroprudential policies could have avoided the excess in credit that triggered the financial crisis in Spain. We find that a countercyclical loan-to-value (LTV) rule that mainly responds to house prices would have mitigated the credit boom in Spain. These results can also be applied to other countries facing similar problems in the housing sector and thinking about implementing macroprudential policies.
HOW CBN CONFRONTED THE MELTDOWN: THE GLOBAL FINANCIAL CRISIS AND THE CENTRAL BANK OF NIGERIA’S RESPONSE
Authors: OZSOZ EMRE , AKINKUNMI MUSTAPHA , AY ISMAIL CAGRI , BAMIDELE ADEMOLA
Published: April 2016

This paper provides an analysis of policy responses to the Global Financial Crisis by the Central Bank of Nigeria (CBN). Given its unique position as a major commodity exporter with a large population, Nigerian authorities utilized a mixture of policies including reductions in the monetary policy rate and capital reserve requirement, lending through the expanded discount window, money market interbank transactions guaranty and limitations on deposit money banks’ (DMBs) foreign exchange net open positions. CBN also rolled over margin loans that were extended to equity investors. As a result the country weathered the financial crisis with limited damage and recorded positive growth rates between 2008 and 2010.
Synthesis ofmeso-substituted porphyrins using sustainable chemical processes
Authors: Pinto Sara M. A. , Henriques César A. , Tomé Vanessa A. , Vinagreiro Carolina S. , Calvete Mário J. F. , Dąbrowski Janusz M. , Piñeiro Marta , Arnaut Luis G. , Pereira Mariette M.
Published: April 2016

The easy access and low price of pyrrole and aldehydes, conjugated with the simplicity in preparing meso-substituted porphyrins, makes this type of porphyrins very attractive for a broad range of applications. However, there is an increasing demand for the development of new synthetic processes involving more sustainable chemical principles substituting, whenever possible, dangerous organic solvents by alternative solvents, chromatographic purifications by precipitations and energy-intensive procedures by alternative energy sources such as microwaves and ultrasounds. In this review we will address some recent strategies to synthesize meso-substituted porphyrins using alternative energy sources, reaction media and catalysts, namely microwave irradiation, water as solvent, or solid microporous acid catalysts.
4-tert-butylphenoxy substituted phthalocyanine with RGD motif as highly selective one-photon and two-photon imaging probe for mitochondria and cancer cell
Authors: Luan Liqiang , Fang Wenjuan , Liu Wei , Tian Minggang , Ni Yuxing , Chen Xi , Yu Xiaoqiang , He Jing , Yang Yang , Li Xiangzhi
Published: April 2016

An unsymmetrical phthalocyanine based one- and two-photon fluorescence imaging probe that substituted with 4-tert-butylphenoxy and RDGyK moieties was developed and characterized by UV-vis and high-resolution MALDI-TOF/MS. The conjugate is non-aggregated in [Formula: see text],[Formula: see text]-dimethylformamide, with relatively weak fluorescence emission ([Formula: see text] 0.023) and high singlet oxygen quantum yield ([Formula: see text] 0.55). Conjugation of the cyclic peptide sequence c(RGDyK) can enhance the cellular uptake towards the DU145 and PC3 cells. While the fluorescence is greatly enhanced in mitochondria, the conjugate is non-cytotoxicity either in dark or upon exposure to red-light with dose up to 12 J.cm[Formula: see text]. The results suggest that this conjugate is a promising multifunctional imaging probe for mitochondria and cancer.
Modulation of the microbiota-gut-brain axis by probiotics in a murine model of inflammatory bowel disease
Authors: Emge Jacob R. , Huynh Kevin , Miller Elaine N. , Kaur Manvir , Reardon Colin , Barrett Kim E. , Gareau Mélanie G.
Published: April 2016

Anxiety, depression, and altered memory are associated with intestinal diseases, including inflammatory bowel disease (IBD). Understanding the link between these behavioral changes and IBD is important clinically since concomitant mood disorders often increase a patient's risk of requiring surgery and developing secondary functional gastrointestinal diseases. Anxiety-like behavior (light/dark box test) and recognition memory (novel object recognition task) were determined at the peak and during resolution of inflammation in the dextran sodium sulfate (DSS) mouse model of acute colitis. DSS (5 days) was administered via drinking water followed by 3 or 9 days of normal drinking water to assess behavior during active or resolving inflammation, respectively. Disease (weight, colon length, and histology) was assessed and the composition of the gut microbiota was characterized by using qPCR on fecal pellet DNA. In a subset of mice, pretreatment with probiotics was started 1 wk prior to commencing DSS. During active inflammation (8 days), mice demonstrated impaired recognition memory and exhibited anxiety-like behavior vs. controls. These behavioral defects were normalized by 14 days post-DSS. Shifts in the composition of the gut microbiota were evident during active inflammation, notably as decreases in lactobacilli and segmented filamentous bacteria, which were also reversed once the disease had resolved. Administration of probiotics could prevent the behavioral defects seen in acute DSS. Taken together, our findings indicate that changes in mood and behavior are present during acute inflammation in murine IBD and associated with dysbiosis and that these outcomes can be prevented by the administration of probiotics.
Deletion of thioredoxin-interacting protein improves cardiac inotropic reserve in the streptozotocin-induced diabetic heart
Authors: Myers Ronald B. , Fomovsky Gregory M. , Lee Samuel , Tan Max , Wang Bing F. , Patwari Parth , Yoshioka Jun
Published: April 2016

Although the precise pathogenesis of diabetic cardiac damage remains unclear, potential mechanisms include increased oxidative stress, autonomic nervous dysfunction, and altered cardiac metabolism. Thioredoxin-interacting protein (Txnip) was initially identified as an inhibitor of the antioxidant thioredoxin but is now recognized as a member of the arrestin superfamily of adaptor proteins that classically regulate G protein-coupled receptor signaling. Here we show that Txnip plays a key role in diabetic cardiomyopathy. High glucose levels induced Txnip expression in rat cardiomyocytes in vitro and in the myocardium of streptozotocin-induced diabetic mice in vivo. While hyperglycemia did not induce cardiac dysfunction at baseline, β-adrenergic challenge revealed a blunted myocardial inotropic response in diabetic animals (24-wk-old male and female C57BL/6;129Sv mice). Interestingly, diabetic mice with cardiomyocyte-specific deletion of Txnip retained a greater cardiac response to β-adrenergic stimulation than wild-type mice. This benefit in Txnip-knockout hearts was not related to the level of thioredoxin activity or oxidative stress. Unlike the β-arrestins, Txnip did not interact with β-adrenergic receptors to desensitize downstream signaling. However, our proteomic and functional analyses demonstrated that Txnip inhibits glucose transport through direct binding to glucose transporter 1 (GLUT1). An ex vivo analysis of perfused hearts further demonstrated that the enhanced functional reserve afforded by deletion of Txnip was associated with myocardial glucose utilization during β-adrenergic stimulation. These data provide novel evidence that hyperglycemia-induced Txnip is responsible for impaired cardiac inotropic reserve by direct regulation of insulin-independent glucose uptake through GLUT1 and plays a role in the development of diabetic cardiomyopathy.
Evidence of microvascular dysfunction in patients with cystic fibrosis
Authors: Rodriguez-Miguelez Paula , Thomas Jeffrey , Seigler Nichole , Crandall Reva , McKie Kathleen T. , Forseen Caralee , Harris Ryan A.
Published: April 2016

Cystic fibrosis (CF) is a genetic, multisystemic disorder with broad clinical manifestations apart from the well-characterized pulmonary dysfunction. Recent findings have described impairment in conduit vessel function in patients with CF; however, whether microvascular function is affected in this population has yet to be elucidated. Using laser-Doppler imaging, we evaluated microvascular function through postocclusive reactive hyperemia (PORH), local thermal hyperemia (LTH), and iontophoresis with acetylcholine (ACh). PORH [518 ± 174% (CF) and 801 ± 125% (control), P = 0.039], LTH [1,338 ± 436% (CF) and 1,574 ± 620% (control), P = 0.045], and iontophoresis with ACh [416 ± 140% (CF) and 617 ± 143% (control), P = 0.032] were significantly lower in patients with CF than control subjects. In addition, the ratio of PORH to LTH was significantly ( P = 0.043) lower in patients with CF (55.3 ± 5.1%) than control subjects (68.8 ± 3.1%). Significant positive correlations between LTH and forced expiratory volume in 1 s (%predicted) ( r = 0.441, P = 0.013) and between the PORH-to-LTH ratio and exercise capacity ( r = 0.350, P = 0.049) were observed. These data provide evidence of microvascular dysfunction in patients with CF compared with control subjects. In addition, our data demonstrate a complex relationship between microvascular function and classical markers of disease severity (i.e., pulmonary function and exercise capacity) in CF.
PGE2-EP3 signaling exacerbates intracerebral hemorrhage outcomes in 24-mo-old mice
Authors: Leclerc Jenna L. , Lampert Andrew S. , Diller Matthew A. , Doré Sylvain
Published: April 2016

With the population aging at an accelerated rate, the prevalence of stroke and financial burden of stroke-related health care costs are expected to continue to increase. Intracerebral hemorrhage (ICH) is a devastating stroke subtype more commonly affecting the elderly population, who display increased mortality and worse functional outcomes compared with younger patients. This study aimed to investigate the contribution of the prostaglandin E2(PGE2) E prostanoid (EP) receptor subtype 3 in modulating anatomical outcomes and functional recovery following ICH in 24-mo-old mice. EP3 is the most abundant EP receptor in the brain and we have previously shown that signaling through the PGE2-EP3 axis exacerbates ICH outcomes in young mice. Here, we show that EP3 receptor deletion results in 17.9 ± 6.1% less ICH-induced brain injury ( P < 0.05) and improves neurological functional recovery ( P < 0.01), as identified by lower neurological deficit scores, decreased resting time, and more gross and fine motor movements. Immunohistological staining was performed to investigate possible mechanisms of EP3-mediated neurotoxicity. Identified mechanisms include reduced blood accumulation and modulation of angiogenic and astroglial responses. Using this aged cohort of mice, we have confirmed and extended our previous results in young mice demonstrating the deleterious role of the PGE2-EP3 signaling axis in modulating brain injury and functional recovery after ICH, further supporting the notion of the EP3 receptor as a putative therapeutic avenue for the treatment of ICH.
Renin overexpression leads to increased titin-based stiffness contributing to diastolic dysfunction in hypertensive mRen2 rats
Authors: Kovács Árpád , Fülöp Gábor Á. , Kovács Andrea , Csípő Tamás , Bódi Beáta , Priksz Dániel , Juhász Béla , Beke Lívia , Hendrik Zoltán , Méhes Gábor , Granzier Henk L. , Édes István , Fagyas Miklós , Papp Zoltán , Barta Judit , Tóth Attila
Published: April 2016

Hypertension (HTN) is a major risk factor for heart failure. We investigated the influence of HTN on cardiac contraction and relaxation in transgenic renin overexpressing rats (carrying mouse Ren-2 renin gene, mRen2, n = 6). Blood pressure (BP) was measured. Cardiac contractility was characterized by echocardiography, cellular force measurements, and biochemical assays were applied to reveal molecular mechanisms. Sprague-Dawley (SD) rats ( n = 6) were used as controls. Transgenic rats had higher circulating renin activity and lower cardiac angiotensin-converting enzyme two levels. Systolic BP was elevated in mRen2 rats (235.11 ± 5.32 vs. 127.03 ± 7.56 mmHg in SD, P < 0.05), resulting in increased left ventricular (LV) weight/body weight ratio (4.05 ± 0.09 vs. 2.77 ± 0.08 mg/g in SD, P < 0.05). Transgenic renin expression had no effect on the systolic parameters, such as LV ejection fraction, cardiomyocyte Ca2+-activated force, and Ca2+ sensitivity of force production. In contrast, diastolic dysfunction was observed in mRen2 compared with SD rats: early and late LV diastolic filling ratio (E/A) was lower (1.14 ± 0.04 vs. 1.87 ± 0.08, P < 0.05), LV isovolumetric relaxation time was longer (43.85 ± 0.89 vs. 28.55 ± 1.33 ms, P < 0.05), cardiomyocyte passive tension was higher (1.74 ± 0.06 vs. 1.28 ± 0.18 kN/m2, P < 0.05), and lung weight/body weight ratio was increased (6.47 ± 0.24 vs. 5.78 ± 0.19 mg/g, P < 0.05), as was left atrial weight/body weight ratio (0.21 ± 0.03 vs. 0.14 ± 0.03 mg/g, P < 0.05). Hyperphosphorylation of titin at Ser-12742 within the PEVK domain and a twofold overexpression of protein kinase C-α in mRen2 rats were detected. Our data suggest a link between the activation of renin-angiotensin-aldosterone system and increased titin-based stiffness through phosphorylation of titin's PEVK element, contributing to diastolic dysfunction.

Listen to this article's corresponding podcast at http://ajpheart.podbean.com/e/diastolic-dysfunction-in-the-hypertensive-mren2-rat/ .
Cutaneous blood flow during intradermal NO administration in young and older adults: roles for calcium-activated potassium channels and cyclooxygenase?
Authors: Fujii Naoto , Meade Robert D. , Minson Christopher T. , Brunt Vienna E. , Boulay Pierre , Sigal Ronald J. , Kenny Glen P.
Published: April 2016

Nitric oxide (NO) increases cutaneous blood flow; however, the underpinning mechanism(s) remains to be elucidated. We hypothesized that the cutaneous blood flow response during intradermal administration of sodium nitroprusside (SNP, a NO donor) is regulated by calcium-activated potassium (KCa) channels and cyclooxygenase (COX) in young adults. We also hypothesized that these contributions are diminished in older adults given that aging can downregulate KCa channels and reduce COX-derived vasodilator prostanoids. In 10 young (23 ± 5 yr) and 10 older (54 ± 4 yr) adults, cutaneous vascular conductance (CVC) was measured at four forearm skin sites infused with 1) Ringer (Control), 2) 50 mM tetraethylammonium (TEA), a nonspecific KCa channel blocker, 3) 10 mM ketorolac, a nonspecific COX inhibitor, or 4) 50 mM TEA + 10 mM ketorolac via intradermal microdialysis. All skin sites were coinfused with incremental doses of SNP (0.005, 0.05, 0.5, 5, and 50 mM each for 25 min). During SNP administration, CVC was similar at the ketorolac site (0.005–50 mM, all P > 0.05) relative to Control, but lower at the TEA and TEA + ketorolac sites (0.005–0.05 mM, all P < 0.05) in young adults. In older adults, ketorolac increased CVC relative to Control during 0.005–0.05 mM SNP administration (all P < 0.05), but this increase was not observed when TEA was coadministered (all P > 0.05). Furthermore, TEA alone did not modulate CVC during any concentration of SNP administration in older adults (all P > 0.05). We show that during low-dose NO administration (e.g., 0.005–0.05 mM), KCa channels contribute to cutaneous blood flow regulation in young adults; however, in older adults, COX inhibition increases cutaneous blood flow through a KCa channel-dependent mechanism.
The CUL3/KLHL3-WNK-SPAK/OSR1 pathway as a target for antihypertensive therapy
Authors: Ferdaus Mohammed Z. , McCormick James A.
Published: April 2016

Chronic high blood pressure (hypertension) is the most common disease in the Unites States. While several classes of drugs exist to treat it, many patients (up to 10 million Americans) respond poorly to therapy, even when multiple classes are used. Recent evidence suggests that a significant portion of patients will always remain hypertensive despite maximum therapy with the drugs currently available. Therefore, there is a pressing need to develop novel antihypertensive agents. One limitation has been the identification of new targets, a limitation that has been overcome by recent insights into the mechanisms underlying monogenic forms of hypertension. The disease familial hyperkalemic hypertension is caused by mutations in with-no-lysine (WNK) kinases 1 and 4 and in cullin-3 and kelch-like 3, components of an E3 ubiquitin ligase complex that promotes WNK kinase degradation. The study of the mechanisms by which this pathway regulates blood pressure has identified several candidates for the development of new antihypertensive agents. This pathway is particularly attractive since its inhibition may not only reduce renal sodium reabsorption along multiple segments but may also reduce vascular tone. Here, we will describe the mechanisms by which this pathway regulate blood pressure and discuss the potential of targeting it to develop new antihypertensive drugs.
CaMKII as a pathological mediator of ER stress, oxidative stress, and mitochondrial dysfunction in a murine model of nephronophthisis
Authors: Bracken Christina , Beauverger Philippe , Duclos Olivier , Russo Ryan J. , Rogers Kelly A. , Husson Hervé , Natoli Thomas A. , Ledbetter Steven R. , Janiak Philip , Ibraghimov-Beskrovnaya Oxana , Bukanov Nikolay O.
Published: April 2016

Polycystic kidney diseases (PKDs) are genetic diseases characterized by renal cyst formation with increased cell proliferation, apoptosis, and transition to a secretory phenotype at the expense of terminal differentiation. Despite recent progress in understanding PKD pathogenesis and the emergence of potential therapies, the key molecular mechanisms promoting cystogenesis are not well understood. Here, we demonstrate that mechanisms including endoplasmic reticulum stress, oxidative damage, and compromised mitochondrial function all contribute to nephronophthisis-associated PKD. Ca2+/calmodulin-dependent protein kinase II (CaMKII) is emerging as a critical mediator of these cellular processes. Therefore, we reasoned that pharmacological targeting of CaMKII may translate into effective inhibition of PKD in jck mice. Our data demonstrate that CaMKII is activated within cystic kidney epithelia in jck mice. Blockade of CaMKII with a selective inhibitor results in effective inhibition of PKD in jck mice. Mechanistic experiments in vitro and in vivo demonstrated that CaMKII inhibition relieves endoplasmic reticulum stress and oxidative damage and improves mitochondrial integrity and membrane potential. Taken together, our data support CaMKII inhibition as a new and effective therapeutic avenue for the treatment of cystic diseases.