Background
Acute heart failure (AHF) is one of the most common diagnoses assigned to emergency department (ED) patients who are hospitalized. Despite its high prevalence in the emergency setting, the ...diagnosis of AHF in ED patients with undifferentiated dyspnea can be challenging.
Objectives
The primary objective of this study was to perform a systematic review and meta‐analysis of the operating characteristics of diagnostic elements available to the emergency physician for diagnosing AHF. Secondary objectives were to develop a test–treatment threshold model and to calculate interval likelihood ratios (LRs) for natriuretic peptides (NPs) by pooling patient‐level results.
Methods
PubMed, EMBASE, and selected bibliographies were searched from January 1965 to March 2015 using MeSH terms to address the ability of the following index tests to predict AHF as a cause of dyspnea in adult patients in the ED: history and physical examination, electrocardiogram, chest radiograph (CXR), B‐type natriuretic peptide (BNP), N‐terminal proB‐type natriuretic peptide (NT‐proBNP), lung ultrasound (US), bedside echocardiography, and bioimpedance. A diagnosis of AHF based on clinical data combined with objective test results served as the criterion standard diagnosis. Data were analyzed using Meta‐DiSc software. Authors of all NP studies were contacted to obtain patient‐level data. The Quality Assessment Tool for Diagnostic Accuracy Studies‐2 (QUADAS‐2) for systematic reviews was utilized to evaluate the quality and applicability of the studies included.
Results
Based on the included studies, the prevalence of AHF ranged from 29% to 79%. Index tests with pooled positive LRs ≥ 4 were the auscultation of S3 on physical examination (4.0, 95% confidence interval CI = 2.7 to 5.9), pulmonary edema on both CXR (4.8, 95% CI = 3.6 to 6.4) and lung US (7.4, 95% CI = 4.2 to 12.8), and reduced ejection fraction observed on bedside echocardiogram (4.1, 95% CI = 2.4 to 7.2). Tests with low negative LRs were BNP < 100 pg/mL (0.11, 95% CI = 0.07 to 0.16), NT‐proBNP < 300 pg/mL (0.09, 95% CI = 0.03 to 0.34), and B‐line pattern on lung US LR (0.16, 95% CI = 0.05 to 0.51). Interval LRs of BNP concentrations at the low end of “positive” results as defined by a cutoff of 100 pg/mL were substantially lower (100 to 200 pg/mL; 0.29, 95% CI = 0.23 to 0.38) than those associated with higher BNP concentrations (1000 to 1500 pg/mL; 7.12, 95% CI = 4.53 to 11.18). The interval LR of NT‐proBNP concentrations even at very high values (30,000 to 200,000 pg/mL) was 3.30 (95% CI = 2.05 to 5.31).
Conclusions
Bedside lung US and echocardiography appear to the most useful tests for affirming the presence of AHF while NPs are valuable in excluding the diagnosis.
CRISPR-Cas systems offer versatile technologies for genome engineering, yet their implementation has been outpaced by ongoing discoveries of new Cas nucleases and anti-CRISPR proteins. Here, we ...present the use of E. coli cell-free transcription-translation (TXTL) systems to vastly improve the speed and scalability of CRISPR characterization and validation. TXTL can express active CRISPR machinery from added plasmids and linear DNA, and TXTL can output quantitative dynamics of DNA cleavage and gene repression—all without protein purification or live cells. We used TXTL to measure the dynamics of DNA cleavage and gene repression for single- and multi-effector CRISPR nucleases, predict gene repression strength in E. coli, determine the specificities of 24 diverse anti-CRISPR proteins, and develop a fast and scalable screen for protospacer-adjacent motifs that was successfully applied to five uncharacterized Cpf1 nucleases. These examples underscore how TXTL can facilitate the characterization and application of CRISPR technologies across their many uses.
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•Active expression of multiple CRISPR nucleases and gRNAs in an E. coli TXTL system•Repression activity of dCas9 strongly correlated between TXTL and E. coli•A TXTL-based PAM assay allowed characterization of five Cpf1 nucleases•Determined specificities of 24 anti-CRISPR proteins against five Cas9 nucleases
Marshall et al. demonstrate that an E. coli cell-free transcription-translation (TXTL) system can be used to improve the speed and scalability of characterizing CRISPR nucleases and their accessory factors. The method will facilitate the discovery of uncharacterized CRISPR nucleases and anti-CRISPR proteins and aid the validation of designed gRNAs.
A series of experiments was established to characterize biochars made from four feedstocks regionally available in the Pacific Northwest (wood pellets, softwood bark, switchgrass (Panicum virgatum ...L.) straw, and anaerobically digested fiber) to determine their effect on five soils. Soils were amended with 9.8, 19.5, and 39.0 Mg ha−1 of each of the four biochars and evaluated for changes in pH, water holding capacity, N mineralization, and soil C. The C content of biochars derived from the herbaceous feedstocks was 60 to 67 kg kg−1 whereas that of the woody feedstocks was >75 kg kg−1. In amended soils we found that biochars, regardless of origin, significantly raised the pH of all soil types 0.1 to 0.9 units, with the greatest impact on a sand soil. The biochars increased the soil C and water holding capacity at the higher rates of amendment depending on soil and biochar type. Nitrogen mineralization rates decreased in three of the five soils across all feedstocks. There were significant correlations (r ≥ 0.9) between biochar C measured and biochar C added regardless of feedstock or soil type. Our research demonstrates that in temperate soils, biochar feedstock may not be as important a variable as soil type for increasing C content and pH but can influence N mineralization and water holding capacity.
Non-vitamin K oral anticoagulants (NOACs) are now widely used as alternatives to warfarin for stroke prevention in atrial fibrillation and management of venous thromboembolism. In clinical practice, ...there is still widespread uncertainty on how to manage patients on NOACs who bleed or who are at risk for bleeding. Clinical trial data related to NOAC reversal for bleeding and perioperative management are sparse, and recommendations are largely derived from expert opinion. Knowledge of time of last ingestion of the NOAC and renal function is critical to managing these patients given that laboratory measurement is challenging because of the lack of commercially available assays in the United States. Idarucizumab is available as an antidote to rapidly reverse the effects of dabigatran. At present, there is no specific antidote available in the United States for the oral factor Xa inhibitors. Prothrombin concentrate may be considered in life-threatening bleeding. Healthcare institutions should adopt a NOAC reversal and perioperative management protocol developed with multidisciplinary input.
Background
Eosinophilic esophagitis (EoE) is a chronic, Th2‐type inflammatory disease. Chemoattractant receptor‐homologous molecule on Th2 cells (CRTH2) is a prostaglandin D2 (PGD2) receptor, ...expressed by Th2 cells and other inflammatory cells, including eosinophils and basophils, that mediates chemotaxis and activation. OC000459 is a selective CRTH2 antagonist and would be expected to suppress eosinophilic tissue inflammation. The purpose of this study was to evaluate the efficacy and safety of an OC000459 monotherapy in adult patients with active, corticosteroid‐dependent or corticosteroid‐refractory EoE.
Methods
In this randomized, double‐blind, placebo‐controlled trial, 26 adult patients (m/f = 22/4; mean age 41 years, range 22–69 years) with active EoE, dependent or resistant to corticosteroids, were treated either with 100 mg OC000459 (n = 14) or placebo (n = 12) twice daily. Pre‐ and post‐treatment disease activity was assessed clinically, endoscopically, histologically, and via biomarkers. The primary end point was the reduction in esophageal eosinophil infiltration.
Results
After an 8‐week OC000459 treatment, the esophageal eosinophil load decreased significantly, from 114.83 to 73.26 eosinophils per high‐power field (eos/hpf), P = 0.0256, whereas no reduction was observed with placebo (102.80–99.47 eos/hpf, P = 0.870). With OC000459, the physician's global assessment of disease activity improved from 7.13 to 5.18 (P = 0.035). OC000459 likewise reduced extracellular deposits of eosinophil peroxidase and tenascin C, the effects not seen with placebo. No serious adverse events were observed.
Conclusions
An 8‐week treatment with the CRTH2‐antagonist, OC000459, exerts modest, but significant, anti‐eosinophil and beneficial clinical effects in adult patients with active, corticosteroid‐dependent or corticosteroid‐refractory EoE and is well tolerated.
Abstract
CRISPR technologies increasingly require spatiotemporal and dosage control of nuclease activity. One promising strategy involves linking nuclease activity to a cell's transcriptional state ...by engineering guide RNAs (gRNAs) to function only after complexing with a ‘trigger’ RNA. However, standard gRNA switch designs do not allow independent selection of trigger and guide sequences, limiting gRNA switch application. Here, we demonstrate the modular design of Cas12a gRNA switches that decouples selection of these sequences. The 5′ end of the Cas12a gRNA is fused to two distinct and non-overlapping domains: one base pairs with the gRNA repeat, blocking formation of a hairpin required for Cas12a recognition; the other hybridizes to the RNA trigger, stimulating refolding of the gRNA repeat and subsequent gRNA-dependent Cas12a activity. Using a cell-free transcription-translation system and Escherichia coli, we show that designed gRNA switches can respond to different triggers and target different DNA sequences. Modulating the length and composition of the sensory domain altered gRNA switch performance. Finally, gRNA switches could be designed to sense endogenous RNAs expressed only under specific growth conditions, rendering Cas12a targeting activity dependent on cellular metabolism and stress. Our design framework thus further enables tethering of CRISPR activities to cellular states.
Central nervous system (CNS) relapse of diffuse large B-cell lymphoma remains uncommon but catastrophic. The benefit of standalone intrathecal prophylaxis in reducing CNS recurrence is unclear and ...remains controversial. No systematic review analysing the evidence for stand-alone intrathecal prophylaxis has been performed in the era of anti-CD20 monoclonal antibody therapy. A comprehensive search (01/2002-01/2019) was systematically performed using Ovid MEDLINE
, Ovid EMBASE
and Cochrane. Studies were selected from a total of 804, screened based on predefined inclusion/exclusion criteria, and were critically appraised. Three post hoc analyses (RICOVER-60, RCHOP-14/21, GOYA), one prospective database and 10 retrospective series were included. 7,357 rituximab/obinutuzumab-exposed patients were analysed. The median percentage receiving intrathecal prophylaxis was 11.9%. Cumulative CNS relapse incidence ranged from 1.9% at 6.5 years to 8.4% at 5 years. Median time (of medians) to CNS relapse was 10 months. 73% developed isolated CNS relapses, 24% concurrent CNS/systemic relapse, and 3% post-systemic relapse. Reported CNS relapse sites were: parenchymal (58%), leptomeningeal (27%), and both (12%). Event rates were low resulting in limited power within each study to provide robust univariable/multivariable analysis. Intrathecal prophylaxis was not a univariable or multivariable factor associated with a reduction in CNS relapse in any study. We found no strong evidence for the benefit, or indeed genuine lack of benefit, of stand-alone intrathecal prophylaxis in preventing CNS relapse in diffuse large B-cell lymphoma-treated patients using anthracycline-based immunochemotherapy. Current published study designs limit the strength of such conclusions.
V-ATPases are proton pumps that function to acidify intracellular compartments in all eukaryotic cells, and to transport protons across the plasma membrane of certain specialized cells. V-ATPases ...function in many normal and disease processes, including membrane traffic, protein degradation, pathogen entry, and cancer cell invasion. An important mechanism of regulating V-ATPase activity
is regulated assembly, which is the reversible dissociation of the ATP-hydrolytic V
domain from the proton-conducting V
domain. Regulated assembly is highly conserved and occurs in response to various nutrient cues, suggesting that it plays an important role in cellular homeostasis. We have recently found that starvation of mammalian cells for either amino acids or glucose increases V-ATPase assembly on lysosomes, possibly to increase protein degradation (for amino acid homeostasis) or for the utilization of alternative energy sources (during glucose starvation). While regulation of assembly in response to amino acid starvation does not involve PI3K or mTORC1, glucose-regulated assembly involves both PI3K and AMPK. Another important form of V-ATPase regulation is the targeting of the enzyme to different cellular membranes, which is controlled by isoforms of subunit a. We have shown that V-ATPases are localized to the plasma membrane of highly invasive breast cancer cells, where they promote cell migration and invasion. Furthermore, overexpression of the a3 isoform is responsible for plasma membrane targeting of V-ATPases in breast tumor cells leading to their increased invasiveness.
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