bluuebunny/crossref_metadata_2025_split huggingface dataset:
A novel mutant Na+/HCO3− cotransporter NBCe1 in a case of compound‐heterozygous inheritance of proximal renal tubular acidosis
Authors: Myers Evan J. , Yuan Lu , Felmlee Melanie A. , Lin Yuan‐Yuan , Jiang Yan , Pei Yu , Wang Ou , Li Mei , Xing Xiao‐Ping , Marshall Aniko , Xia Wei‐Bo , Parker Mark D. 
Published: June 2016

Key points
The inheritance of two defective alleles of SLC4A4, the gene that encodes the widely‐expressed electrogenic sodium bicarbonate cotransporter NBCe1, results in the bicarbonate‐wasting disease proximal renal tubular acidosis (pRTA).
In the present study, we report the first case of compound‐heterozygous inheritance of pRTA (p.Arg510His/p.Gln913Arg) in an individual with low blood pH, blindness and neurological signs that resemble transient ischaemic attacks.
We employ fluorescence microscopy on non‐polarized (human embryonic kidney) and polarized (Madin–Darby canine kidney) renal cell lines and electrophysiology on Xenopus oocytes to characterize the mutant transporters (R510H and Q913R).
Both mutant transporters exhibit enhanced intracellular retention in renal cells, an observation that probably explains the HCO3− transport deficit in the individual.
Both mutants retain a close‐to‐normal per molecule Na+/HCO3− cotransport activity in Xenopus oocytes, suggesting that they are suitable candidates for folding‐correction therapy. However, Q913R expression is uniquely associated with a depolarizing, HCO3− independent, Cl−‐conductance in oocytes that could have pathological consequences if expressed in the cells of patients.
AbstractProximal renal tubular acidosis (pRTA) is a rare, recessively‐inherited disease characterized by abnormally acidic blood, blindness, as well as below average height and weight. pRTA is typically associated with homozygous mutation of the solute carrier 4 family gene SLC4A4. SLC4A4 encodes the electrogenic sodium bicarbonate cotransporter NBCe1, a membrane protein that acts to maintain intracellular and plasma pH. We present the first description of a case of compound‐heterozygous inheritance of pRTA. The individual has inherited two mutations in NBCe1: p.Arg510His (R510H) and p.Gln913Arg (Q913R), one from each parent. In addition to the usual features of pRTA, the patient exhibits unusual signs, such as muscle spasms and fever. We have recreated these mutant transporters for expression in model systems. We find that both of the mutant proteins exhibit substantial intracellular retention when expressed in mammalian renal cell lines. When expressed in Xenopus oocytes, we find that the R510H and Q913R‐mutant NBCe1 molecules exhibit apparently normal Na+/HCO3− cotransport activity but that Q913R is associated with an unusual HCO3− independent anion‐leak. We conclude that a reduced accumulation of NBCe1 protein in the basolateral membrane of proximal‐tubule epithelia is the most probable cause of pRTA in this case. We further note that the Q913R‐associated anion‐leak could itself be pathogenic if expressed in the plasma membrane of mammalian cells, compromising the benefit of strategies aiming to enhance mutant NBCe1 accumulation in the plasma membrane.
Upper limits of the photon fluence rate on CT detectors: Case study on a commercial scanner
Authors: Persson Mats , Bujila Robert , Nowik Patrik , Andersson Henrik , Kull Love , Andersson Jonas , Bornefalk Hans , Danielsson Mats 
Published: June 2016

Purpose:The highest photon fluence rate that a computed tomography (CT) detector must be able to measure is an important parameter. The authors calculate the maximum transmitted fluence rate in a commercial CT scanner as a function of patient size for standard head, chest, and abdomen protocols.Methods:The authors scanned an anthropomorphic phantom (Kyoto Kagaku PBU‐60) with the reference CT protocols provided by AAPM on a GE LightSpeed VCT scanner and noted the tube current applied with the tube current modulation (TCM) system. By rescaling this tube current using published measurements on the tube current modulation of a GE scanner [N. Keat, “CT scanner automatic exposure control systems,” MHRA Evaluation Report 05016, ImPACT, London, UK, 2005], the authors could estimate the tube current that these protocols would have resulted in for other patient sizes. An ECG gated chest protocol was also simulated. Using measured dose rate profiles along the bowtie filters, the authors simulated imaging of anonymized patient images with a range of sizes on a GE VCT scanner and calculated the maximum transmitted fluence rate. In addition, the 99th and the 95th percentiles of the transmitted fluence rate distribution behind the patient are calculated and the effect of omitting projection lines passing just below the skin line is investigated.Results:The highest transmitted fluence rates on the detector for the AAPM reference protocols with centered patients are found for head images and for intermediate‐sized chest images, both with a maximum of 3.4 ⋅ 108 mm−2 s−1, at 949 mm distance from the source. Miscentering the head by 50 mm downward increases the maximum transmitted fluence rate to 5.7 ⋅ 108 mm−2 s−1. The ECG gated chest protocol gives fluence rates up to 2.3 ⋅ 108 − 3.6 ⋅ 108 mm−2 s−1 depending on miscentering.Conclusions:The fluence rate on a CT detector reaches 3 ⋅ 108 − 6 ⋅ 108 mm−2 s−1 in standard imaging protocols, with the highest rates occurring for ECG gated chest and miscentered head scans. These results will be useful to developers of CT detectors, in particular photon counting detectors.
Sequence Analysis of Fluoroquinolone Resistance-Associated Genes
gyrA
and
gyrB
in Clinical Mycobacterium tuberculosis Isolates from Patients Suspected of Having Multidrug-Resistant Tuberculosis in New Delhi, India
Authors: Singhal Ritu , Reynolds Paul R. , Marola Jamie L. , Epperson L. Elaine , Arora Jyoti , Sarin Rohit , Myneedu Vithal Prasad , Strong Michael , Salfinger Max 
Published: June 2016

ABSTRACT

Fluoroquinolones (FQs) are broad-spectrum antibiotics recommended for the treatment of multidrug-resistant tuberculosis (MDR-TB) patients. FQ resistance, caused by mutations in the
gyrA
and
gyrB
genes of
Mycobacterium tuberculosis
, is increasingly reported worldwide; however, information on mutations occurring in strains from the Indian subcontinent is scarce. Hence, in this study, we aimed to characterize mutations in the
gyrA
and
gyrB
genes of acid-fast bacillus (AFB) smear-positive sediments or of
M. tuberculosis
isolates from AFB smear-negative samples from patients in India suspected of having MDR-TB. A total of 152 samples from patients suspected of having MDR-TB were included in the study. One hundred forty-six strains detected in these samples were characterized by sequencing of the
gyrA
and
gyrB
genes. The extracted DNA was subjected to successive amplifications using a nested PCR protocol, followed by sequencing. A total of 27 mutations were observed in the
gyrA
genes of 25 strains, while no mutations were observed in the
gyrB
genes. The most common mutations occurred at amino acid position 94 (13/27 [48.1%]); of these, the D94G mutation was the most prevalent. The
gyrA
mutations were significantly associated with patients with rifampin (RIF)-resistant TB. Heterozygosity was seen in 4/27 (14.8%) mutations, suggesting the occurrence of mixed populations with different antimicrobial susceptibilities. A high rate of FQ-resistant mutations (17.1%) was obtained among the isolates of TB patients suspected of having MDR-TB. These observations emphasize the need for accurate and rapid molecular tests for the detection of FQ-resistant mutations at the time of MDR-TB diagnosis.
Verification of an Automated, Digital Dispensing Platform for At-Will Broth Microdilution-Based Antimicrobial Susceptibility Testing
Authors: Smith Kenneth P. , Kirby James E. 
Published: June 2016

ABSTRACT

With rapid emergence of multidrug-resistant bacteria, there is often a need to perform susceptibility testing for less commonly used or newer antimicrobial agents. Such testing can often be performed only by using labor-intensive, manual dilution methods and lies outside the capacity of most clinical labs, necessitating reference laboratory testing and thereby delaying the availability of susceptibility data. To address the compelling clinical need for microbiology laboratories to perform such testing in-house, we explored a novel, automated, at-will broth microdilution-based susceptibility testing platform. Specifically, we used the modified inkjet printer technology in the HP D300 digital dispensing system to dispense, directly from stock solutions into a 384-well plate, the 2-fold serial dilution series required for broth microdilution testing. This technology was combined with automated absorbance readings and data analysis to determine MICs. Performance was verified by testing members of the
Enterobacteriaceae
for susceptibility to ampicillin, cefazolin, ciprofloxacin, colistin, gentamicin, meropenem, and tetracycline in comparison to the results obtained with a broth microdilution reference standard. In precision studies, essential and categorical agreement levels were 96.8% and 98.3%, respectively. Furthermore, significantly fewer D300-based measurements were outside ±1 dilution from the modal MIC, suggesting enhanced reproducibility. In accuracy studies performed using a panel of 80 curated clinical isolates, rates of essential and categorical agreement and very major, major, and minor errors were 94%, 96.6%, 0%, 0%, and 3.4%, respectively. Based on these promising initial results, it is anticipated that the D300-based methodology will enable hospital-based clinical microbiology laboratories to perform at-will broth microdilution testing of antimicrobials and to address a critical testing gap.
CD4-Induced Antibodies Promote Association of the HIV-1 Envelope Glycoprotein with CD4-Binding Site Antibodies
Authors: Gardner Matthew R. , Fellinger Christoph H. , Prasad Neha R. , Zhou Amber S. , Kondur Hema R. , Joshi Vinita R. , Quinlan Brian D. , Farzan Michael 
Published: June 2016

ABSTRACT
The HIV-1 envelope glycoprotein (Env) is a trimer of gp120/gp41 heterodimers that mediates viral entry. Env binds cellular CD4, an association which stabilizes a conformation favorable to its subsequent association with a coreceptor, typically CCR5 or CXCR4. The CD4- and coreceptor-binding sites serve as epitopes for two classes of HIV-1-neutralizing antibodies: CD4-binding site (CD4bs) and CD4-induced (CD4i) antibodies, respectively. Here we observed that, at a fixed total concentration, mixtures of the CD4i antibodies (E51 or 412d) and the CD4bs antibody VRC01 neutralized the HIV-1 isolates 89.6, ADA, SG3, and SA32 more efficiently than either antibody alone. We found that E51, and to a lesser extent 412d and 17b, promoted association of four CD4bs antibodies to the Env trimer but not to monomeric gp120. We further demonstrated that the binding of the sulfotyrosine-binding pocket by CCR5mim2-Ig was sufficient for promoting CD4bs antibody binding to Env. Interestingly, the relationship is not reciprocal: CD4bs antibodies were not as efficient as CD4-Ig at promoting E51 or 412d binding to Env trimer. Consistent with these observations, CD4-Ig, but none of the CD4bs antibodies tested, substantially increased HIV-1 infection of a CD4-negative, CCR5-positive cell line. We conclude that the ability of CD4i antibodies to promote VRC01 association with Env trimers accounts for the increase potency of VRC01 and CD4i antibody mixtures. Our data further suggest that potent CD4bs antibodies avoid inducing Env conformations that bind CD4i antibodies or CCR5.

IMPORTANCE
Potent HIV-1-neutralizing antibodies can prevent viral transmission and suppress an ongoing infection. Here we show that CD4-induced (CD4i) antibodies, which recognize the conserved coreceptor-binding site of the HIV-1 envelope glycoprotein (Env), can increase the association of Env with potent broadly neutralizing antibodies that recognize the CD4-binding site (CD4bs antibodies). We further show that, unlike soluble forms of CD4, CD4bs antibodies poorly induce envelope glycoprotein conformations that efficiently bind CCR5. This study provides insight into the properties of potent CD4bs antibodies and suggests that, under some conditions, CD4i antibodies can improve their potency. These observations may be helpful to the development of vaccines designed to elicit specific antibody classes.
Opposing Roles of Double-Stranded RNA Effector Pathways and Viral Defense Proteins Revealed with CRISPR-Cas9 Knockout Cell Lines and Vaccinia Virus Mutants
Authors: Liu Ruikang , Moss Bernard 
Published: June 2016

ABSTRACT
Vaccinia virus (VACV) decapping enzymes and cellular exoribonuclease Xrn1 catalyze successive steps in mRNA degradation and prevent double-stranded RNA (dsRNA) accumulation, whereas the viral E3 protein can bind dsRNA. We showed that dsRNA and E3 colocalized within cytoplasmic viral factories in cells infected with a decapping enzyme mutant as well as with wild-type VACV and that they coprecipitated with antibody. An E3 deletion mutant induced protein kinase R (PKR) and eukaryotic translation initiation factor alpha (eIF2α) phosphorylation earlier and more strongly than a decapping enzyme mutant even though less dsRNA was made, leading to more profound effects on viral gene expression. Human HAP1 and A549 cells were genetically modified by clustered regularly interspaced short palindromic repeat-Cas9 (CRISPR-Cas9) to determine whether the same pathways restrict E3 and decapping mutants. The E3 mutant replicated in PKR knockout (KO) HAP1 cells in which RNase L is intrinsically inactive but only with a double knockout (DKO) of PKR and RNase L in A549 cells, indicating that both pathways decreased replication equivalently and that no additional dsRNA pathway was crucial. In contrast, replication of the decapping enzyme mutant increased significantly (though less than that of wild-type virus) in DKO A549 cells but not in DKO HAP1 cells where a smaller increase in viral protein synthesis occurred. Xrn1 KO A549 cells were viable but nonpermissive for VACV; however, wild-type and mutant viruses replicated in triple-KO cells in which RNase L and PKR were also inactivated. Since KO of PKR and RNase L was sufficient to enable VACV replication in the absence of E3 or Xrn1, the poor replication of the decapping mutant, particularly in HAP1 DKO, cells indicated additional translational defects.

IMPORTANCE
Viruses have evolved ways of preventing or counteracting the cascade of antiviral responses that double-stranded RNA (dsRNA) triggers in host cells. We showed that the dsRNA produced in excess in cells infected with a vaccinia virus (VACV) decapping enzyme mutant and by wild-type virus colocalized with the viral E3 protein in cytoplasmic viral factories. Novel human cell lines defective in either or both protein kinase R and RNase L dsRNA effector pathways and/or the cellular 5′ exonuclease Xrn1 were prepared by CRISPR-Cas9 gene editing. Inactivation of both pathways was necessary and sufficient to allow full replication of the E3 mutant and reverse the defect cause by inactivation of Xrn1, whereas the decapping enzyme mutant still exhibited defects in gene expression. The study provided new insights into functions of the VACV proteins, and the well-characterized panel of CRISPR-Cas9-modified human cell lines should have broad applicability for studying innate dsRNA pathways.
Results of a nationally implemented de novo cardiac screening programme in elite rugby players in England
Authors: Ghani Saqib , Papadakis Michael , Kemp Simon , Zaidi Abbas , Sheikh Nabeel , Gati Sabiha , Raju Hariharan , Smith Andy , Palmer Corin , Somauroo John , Sharma Sanjay 
Published: June 2016

Background/aimScreening of young competitive athletes remains a contentious issue. In 2010, a nationwide cardiac screening for all elite rugby players was introduced in England. This provided a unique opportunity to prospectively assess the feasibility and cost-effectiveness of a de novo, ECG-based cardiac screening programme.MethodsBetween 2010 and 2012, 1191 rugby players aged ≥14 years underwent cardiac screening with a health questionnaire, 12-lead ECG and a consultation with a cardiologist. The players with concerning findings on initial evaluation were offered on-site transthoracic echocardiogram (TTE). Athletes were referred for further investigations as deemed necessary. The overall cost of the screening programme was estimated.ResultsAfter initial evaluation, 9.7% of athletes underwent on-site TTE; 8.2% underwent on-site TTE due to ECG anomalies and 1.4% underwent on-site TTE due to concerns on the questionnaire. After TTE, only 2.9% of the total cohort was referred for further evaluation. Two players were diagnosed with potentially serious conditions; one with Wolff-Parkinson-White, who resumed competition after catheter ablation, and one with hypertrophic cardiomyopathy, who withdrew from competition. During a mean follow-up of 52.8±5.5 months, none of the players who were reassured experienced any adverse cardiac events. The total cost of the screening programme was £59 875, which averaged to a cost of £50 per player or £29 938 per condition identified. Application of refined ECG criteria would reduce the ECG false-positive rate to 4.9%.ConclusionsPreparticipation cardiac screening with 12-lead ECG is feasible. Refinement of the ECG criteria, the use of on-site TTE and expert setting can minimise the burden of unnecessary investigations and reduce costs.
Clinical reasoning of junior doctors in emergency medicine: a grounded theory study
Authors: Adams E , Goyder C , Heneghan C , Brand L , Ajjawi R 
Published: June 2016

IntroductionEmergency medicine (EM) has a high case turnover and acuity making it a demanding clinical reasoning domain especially for junior doctors who lack experience. We aimed to better understand their clinical reasoning using dual cognition as a guiding theory.MethodsEM junior doctors were recruited from six hospitals in the south of England to participate in semi-structured interviews (n=20) and focus groups (n=17) based on recall of two recent cases. Transcripts were analysed using a grounded theory approach to identify themes and to develop a model of junior doctors' clinical reasoning in EM.ResultsWithin cases, clinical reasoning occurred in three phases. In phase 1 (case framing), initial case cues and first impressions were predominantly intuitive, but checked by analytical thought and determined the urgency of clinical assessment. In phase 2 (evolving reasoning), non-analytical single cue and pattern recognitions were common which were subsequently validated by specific analytical strategies such as use of red flags. In phase 3 (ongoing uncertainty) analytical self-monitoring and reassurance strategies were used to precipitate a decision regarding discharge.ConclusionWe found a constant dialectic between intuitive and analytical cognition throughout the reasoning process. Our model of clinical reasoning by EM junior doctors illustrates the specific contextual manifestations of the dual cognition theory. Distinct diagnostic strategies are identified and together these give EM learners and educators a framework and vocabulary for discussion and learning about clinical reasoning.
Safety in numbers for cyclists beyond national-level and city-level data: a study on the non-linearity of risk within the city of Hong Kong
Authors: Yao Shenjun , Loo Becky P Y 
Published: June 2016

ObjectiveThis paper examines the relationship between bicycle collisions and the amount of cycling at the local level. Most previous research has focused on national and city comparisons, little is known about differences within a city (the mesoscale).MethodsThis study mainly used three types of data sets relating to bicycle collisions, use of bicycles and local neighbourhood characteristics in Hong Kong. In particular, bicycle usage, measured as bicycle-kilometres travelled, was estimated from travel surveys following the activity-based approach. Negative binomial regression models were established to model the relationship between the amount of cycling and the occurrence of bicycle collisions at the spatial scale of the Tertiary Planning Unit, which is the smallest planning unit of the city.ResultsThe numbers of bicycle collisions went up with the increasing use of bicycles, but the increase in the number of collisions in a given community was less than a linear proportion of the bicycle flow. When other local neighbourhood variables are controlled, the amount of cycling is a statistically significant variable in accounting for the number of collisions.ConclusionsEven in a highly motorised city where bicycles are a minor transport mode, cyclists are less likely to be involved in road collisions in communities with higher cycling volume. Since cycling activities are likely to vary within a city, a more local-based approach in promoting cycling is needed. In particular, the higher safety risks in neighbourhoods of low bicycle usage, especially at an initial stage of promoting cycling, need to be addressed properly.
Laundry pod and non-pod detergent related emergency department visits occurring in children in the USA
Authors: Swain Thomas A , McGwin Gerald , Griffin Russell 
Published: June 2016

ObjectivePrevious studies have reported that children are at risk of severe injuries from exposure to laundry detergent pods. For the first time, this study sought to compare demographic and exposure characteristics and risk among children exposed to pod and non-pod laundry detergents presenting to emergency departments (EDs).MethodsData from the National Electronic Injury Surveillance System (NEISS) from 2012–2014 were used. All observations with injuries involving laundry detergent (NEISS code 0949) were included in this study. The χ2test was used for bivariate analysis and logistic regression was used to determine the OR and 95% CI of hospitalisation for pod related versus non-pod laundry detergent exposures.ResultsFrom 2012–2014, there were an estimated 26 062 non-pod and 9814 pod laundry detergent related exposures among those aged 18 years and younger. For pod detergent, children aged 0–5 years had the most injuries. Poisoning (71.3%) was the most common diagnosis for pod detergent while contact dermatitis (72.2%) was most common for non-pod detergent. Hospitalisation occurred in 12.5% of pod detergent cases and just 3.0% of non-pod cases. Compared with non-pod detergent, those exposed to pod detergent were 4 times as likely to be hospitalised (OR 4.02; 95% CI 1.96 to 8.24).ConclusionsA greater effort should be made to appropriately educate the public about the dangers of laundry detergents, specifically pods, so a safe home environment can be established. While new regulations such as childproof containers, opaque packaging, and less appealing and colourful pods could reduce the number of pod related ED visits for children, caregivers should store detergents, along with other chemicals, in a secure location where children cannot easily access them.
Ipilimumab Combined with Nivolumab: A Standard of Care for the Treatment of Advanced Melanoma?
Authors: Carlino Matteo S. , Long Georgina V. 
Published: June 2016

Abstract
Ipilimumab, an inhibitor of CTLA-4 on T cells, was the first drug to improve overall survival in patients with advanced melanoma. Subsequently, inhibitors of PD-1, including nivolumab and pembrolizumab, were shown to be superior to ipilimumab with a more favorable safety profile. The combination of ipilimumab and nivolumab is associated with a further improvement in response rate and progression-free survival; however, the combination is associated with an increased rate of immune-related toxicities. In 2015, the FDA approved the combination for the treatment of BRAF wild-type advanced melanoma. This review examines the preclinical rationale for the combination of ipilimumab and nivolumab as well as the efficacy and toxicity data from clinical trials in patients with advanced melanoma. Finally, alternative treatment options are discussed with a focus on patient selection. Clin Cancer Res; 22(16); 3992–8. ©2016 AACR.