bluuebunny/crossref_metadata_2025 huggingface dataset:
Can the Third Part of Duodenum Behind SMA Be Detected With Confidence on CT as a Proposed Mechanism for Imaging Suspected Malrotation in Children? A Preliminary Proof of Concept Study
Authors: Werndle Melissa C. , Venkatakrishna Shyam Sunder B. , Holjar-Erlic Izidora , Khumalo Zonah , Menegotto Flavia , Zouvani Andrea , Andronikou Savvas
Published: March 2022

Aim

The aims of this study were to determine frequency and reliability of computed tomography (CT) detection of anatomic landmarks for imaging suspected midgut malrotation in infants and children, and to calculate an estimated effective dose of an upper abdominal CT scan in our patient population.

Materials and Methods

Fifty consecutive pediatric patients who underwent a CT scan that included their upper abdomen between August 2016 and February 2018 were included. Four pediatric radiology consultants independently reviewed CT scans for detection of the third part of the duodenum and defined their confidence level of this through identification of continuity with the pyloric antrum, D1, D2, and D4 components of the duodenum, as well as the duodenojejunal flexure.

Interobserver variability was assessed using Fleiss κ for agreement. A dose estimate, per scan, was calculated using the scanner dose-length product and published conversion factors by Deak.

Results

Thirty patients were boys. The average age was 7.5 ± 5.4 years (6 days to 16 years). The D3 segment was definitely identified in 70% of scans, with 68% to 73%, moderate agreement between the readers and a Fleiss κ of 0.47 to 0.52. The DJ flexure was definitely identified in only 30.5% cases, with 35%, poor agreement between readers (Fleiss κ of 0.03). The average estimated dose for a targeted CT scan of the abdomen was 0.9 mSv (0.04–2.4 mSv).

Conclusions

The third part of the duodenum, which is integral in excluding malrotation on cross-sectional studies, was “definitely” identified in 70% of CT scans of children in our study, with 68% to 73% agreement between the readers and a Fleiss κ of 0.47 to 0.52.

These preliminary proof of concept results demonstrating a combination of a comparable CT dose in relation to upper gastrointestinal contrast studies and an acceptable number of cases delineating the third part of the duodenum with moderate agreement are a first step in suggesting low-dose CT for an imaging diagnosis of malrotation. Malrotation can be excluded in cases where D3 is well demonstrated in the normal position, which negates the need to automatically refer children with bilious emesis to specialist centers for upper gastrointestinal contrast studies.
Intravenous Contrast Material for Cardiac Computed Tomography
Authors: La Grutta Ludovico , Maffei Erica , Clemente Alberto , Privitera Giambattista , Rengo Marco , Seitun Sara , Malagò Roberto , Toia Patrizia , Acquafresca Manlio , Secchi Francesco , Gravina Matteo , Mantini Cesare , Scardapane Arnaldo , De Rosa Roberto , Carusi Luca Maria , Cossa Stefano , Falzea Fabio , Chiappino Dante , Annoni Andrea , Francone Marco , Di Cesare Ernesto , Midiri Massimo , Cademartiri Filippo
Published: March 2022

Purpose:

The Italian Registry of Contrast Material use in Cardiac Computed Tomography (iRCM-CCT) is a multicenter, multivendor, observational study on the use of contrast media (CM) in patients undergoing cardiac computed tomography (CCT). The aim of iRCM-CCT is to assess image quality and safety profile of intravenous CM compounds.

Materials and Methods:

iRCM-CCT enrolled 1842 consecutive patients undergoing CCT (≥50 per site) at 20 cluster sites with the indication of suspected coronary artery disease. Demographic characteristics, CCT, and CM protocols, clinical indications, safety markers, radiation dose reports, qualitative (ie, poor vascular enhancement) and quantitative (ie, HU attenuation values) image parameters were recorded. A centralized coordinating center collected and assessed all image parameters.

Results:

The cohort included 891 men and 951 women (age: 63±14 y, body mass index: 26±4 kg/m2) studied with ≥64 detector rows computed tomography scanners and different iodinated intravenous CM protocols and compounds (iodixanol, iopamidol, iohexol, iobitridol, iopromide, and iomeprol). The following vascular attenuation was reported: 504±147 HU in the aorta, 451±146 HU in the right coronary artery, 474±146 HU in the left main, 451±146 HU in the left anterior descending artery, and 441±149 HU in the circumflex artery. In 4% of cases the image quality was not satisfactory due to poor enhancement. The following adverse reactions to CM were recorded: 6 (0.3%) extravasations and 17 (0.9%) reactions (11 mild, 4 moderate, 2 severe).

Conclusions:

In a multicenter registry on CM use during CCT the prevalence of CM-related adverse reactions was very low. The appropriate use of CM is a major determinant of image quality.
Healthy Food Choices, Physical Activity, and Screening Reduce the Risk of Colorectal Cancer
Authors: Morrow Linda , Greenwald Beverly
Published: March 2022

Cancer is the second leading cause of death in the United States with an estimated 1,806,590 new cases and 606,520 deaths in 2020. Cancer is a public health concern due to the numbers of cases, financial costs, and morbidity and mortality rates. An estimated 42% of all diagnosed cancers and 45% of cancer deaths in the United States in 2014 could be attributed to modifiable risk factors. Colorectal cancer screening is effective because small polyps take years to grow and turn to cancers allowing for early detection and removal of precancerous polyps. Primary prevention of colorectal cancer by risk factor reduction is also effective as these factors cause over half of colorectal cancer cases and deaths. Physical activity, weight control, healthy dietary choices, and abstinence from alcohol and tobacco are protective. The gastroenterology nurse can assess physical activity and food choices as vital signs and recommend a gradual addition of physical activity and a more plant-based diet with reduced processed food. All nurses need to advocate for policy improvements per our pledge to the Code of Ethics for Nurses. Policy changes improve the health of large numbers of people and make healthy behaviors a normalized way of life.
Stability of asymptotic behaviour within polarized T2-symmetric vacuum solutions with cosmological constant
Authors: Ames Ellery , Beyer Florian , Isenberg James , Oliynyk Todd A.
Published: March 2022

We prove the nonlinear stability of the asymptotic behaviour of perturbations of subfamilies of Kasner solutions in the contracting time direction within the class of polarizedT2-symmetric solutions of the vacuum Einstein equations with arbitrary cosmological constantΛ. This stability result generalizes the results proven in Ames Eet al.(2022 Stability of AVTD Behavior within the PolarizedT2-symmetric vacuum spacetimes.Ann. Henri Poincaré. (doi:10.1007/s00023-021-01142-0)), which focus on theΛ=0case, and as in that article, the proof relies on an areal time foliation and Fuchsian techniques. Even forΛ=0, the results established here apply to a wider class of perturbations of Kasner solutions within the family of polarizedT2-symmetric vacuum solutions than those considered in Ames Eet al.(2022 Stability of AVTD Behavior within the PolarizedT2-symmetric vacuum spacetimes.Ann. Henri Poincaré. (doi:10.1007/s00023-021-01142-0)) and Fournodavlos Get al.(2020Stable Big Bang formation for Einstein’s equations: the complete sub-critical regime. Preprint. (http://arxiv.org/abs/2012.05888)). Our results establish that the areal time coordinate takes all values in(0,T0]for someT0>0, for certain families of polarizedT2-symmetric solutions with cosmological constant.

This article is part of the theme issue ‘The future of mathematical cosmology, Volume 1’.
Some unusual wormholes in general relativity
Authors: Bronnikov Kirill A.
Published: March 2022

In this short review, we present some recently obtained traversable wormhole models in the framework of general relativity (GR) in four and six dimensions that somehow widen our common ideas on wormhole existence and properties. These are, first, rotating cylindrical wormholes, asymptotically flat in the radial direction and existing without exotic matter. The topological censorship theorems are not violated due to lack of asymptotic flatness in all spatial directions. Second, these are cosmological wormholes constructed on the basis of the Lemaître–Tolman–Bondi solution. They connect two copies of a closed Friedmann world filled with dust, or two otherwise distant parts of the same Friedmann world. Third, these are wormholes obtained in six-dimensional GR, whose one entrance is located in ‘our’ asymptotically flat world with very small extra dimensions while the other ‘end’ belongs to a universe with large extra dimensions and therefore different physical properties. The possible observable features of such wormholes are briefly discussed.

This article is part of the theme issue ‘The future of mathematical cosmology, Volume 1’.
Beyond energy balance in agrivoltaic food production: Emergent crop traits from wavelength-selective solar cells
Authors: Charles Melodi , Edwards Brianne , Ravishankar Eshwar , Calero John , Henry Reece , Rech Jeromy , Saravitz Carole , You Wei , Ade Harald , O’Connor Brendan , Sederoff Heike
Published: March 2022

The integration of semi-transparent organic solar cells (ST-OSCs) in greenhouses offers new agrivoltaic opportunities to meet the growing demands for sustainable food production. The tailored absorption/transmission spectra of ST-OSCs impacts the power generated as well as crop growth, development and responses to the biotic and abiotic environments. We grew lettuce and tomato, traditional greenhouse crops, under three ST-OSC filters that create different light spectra. Lettuce yield and early tomato development are not negatively affected by the modified light environment. Our genomic analysis reveals that lettuce production exhibits beneficial traits involving nutrient content and nitrogen utilization while select ST-OSCs impact regulation of flowering initiation in tomato. ST-OSCs integrated into greenhouses are not only a promising technology for energy-neutral, sustainable and climate-change protected crop production, but can deliver benefits beyond energy considerations.
sDarken: Next generation genetically encoded fluorescent sensors for serotonin
Authors: Kubitschke Martin , Müller Monika , Wallhorn Lutz , Pulin Mauro , Mittag Manuel , Pollok Stefan , Ziebarth Tim , Bremshey Svenja , Gerdey Jill , Claussen Kristin Carolin , Renken Kim , Groß Juliana , Meyer Niklas , Wiegert Simon , Reiner Andreas , Fuhrmann Martin , Masseck Olivia Andrea
Published: March 2022

We developed a new family of genetically encoded serotonin (5-HT) sensors (sDarken) on the basis of the native 5-HT1A receptor and circularly permuted GFP. sDarken 5-HT sensors are bright in the unbound state and diminish their fluorescence upon binding of 5-HT. Sensor variants with different affinities for serotonin were engineered to increase the versatility in imaging of serotonin dynamics. Experiments in vitro and in vivo showed the feasibility of imaging serotonin dynamics with high temporal and spatial resolution. As demonstrated here, the designed sensors showed excellent membrane expression, have high specificity, a superior signal-to-noise ratio, detect the endogenous release of serotonin and are suitable for two-photon in vivo imaging.
Evaluating spatial and network properties of NMDA-dependent neuronal connectivity in mixed cortical cultures
Authors: Rojvirat Catherine P. , Berlin Joshua R. , Nguyen Tuan D.
Published: March 2022

A technique combining fluorescence imaging with Ca2+ indicators and single-cell laser scanning photostimulation of caged glutamate (LSPS) allows identification of functional connections between individual neurons in mixed cultures of rat neocortical cells as well as observation of synchronous spontaneous activity among neurons. LSPS performed on large numbers of neurons yielded maps of functional connections between neurons and allowed calculation of neuronal network parameters. LSPS also provided an indirect measure of excitability of neurons targeted for photostimulation. By repeating LSPS sessions with the same neurons, stability of connections and change in the number and strength of connections were also determined. Experiments were conducted in the presence of bicuculline to study the properties of excitatory neurotransmission. The AMPA receptor inhibitor, 6-Cyano-7-nitroquinoxaline-2,3-dione (CNQX), abolished synchronous neuronal activity but had no effect on connections mapped by LSPS. In contrast, the NMDA receptor inhibitor, 2-Amino-5-phosphono-pentanoic acid (APV), dramatically decreased the number of functional connections between neurons while also affecting synchronous spontaneous activity. Functional connections were also decreased by increasing extracellular Mg2+ concentration. These data demonstrated that LSPS mapping interrogates NMDA receptor-dependent connectivity between neurons in the network. A GluN2A-specific inhibitor, NVP-AAM077, decreased the number and strength of connections between neurons as well as neuron excitability. Conversely, the GluN2A-specific positive modulator, GNE-0723, increased these same properties. These data showed that LSPS can be used to directly study perturbations in the properties of NMDA receptor-dependent connectivity in neuronal networks. This approach should be applicable in a wide variety of in vitro and in vivo experimental preparations.
Gene selection by incorporating genetic networks into case-control association studies
Authors: Cao Xuewei , Liang Xiaoyu , Zhang Shuanglin , Sha Qiuying
Published: March 2022

Large-scale genome-wide association studies (GWAS) have been successfully applied to a wide range of genetic variants underlying complex diseases. The network-based penalized regression approach has been developed to overcome the challenges caused by the computational efficiency for analyzing high-dimensional genomic data by incorporating a biological genetic network. In this paper, we propose a gene selection approach by incorporating genetic networks into case-control association studies for DNA sequence data or DNA methylation data. Instead of using traditional dimension reduction techniques such as principal component analyses and supervised principal component analyses, we use a linear combination of genotypes at SNPs or methylation values at CpG sites in each gene to capture gene-level signals. We develop three approaches for the linear combination: optimally weighted sum (OWS), LD-adjusted polygenic risk score (LD-PRS), and beta-based weighted sum (BWS). OWS and LD-PRS are supervised approaches that depend on the effect of each SNP or CpG site on the case-control status, while BWS can be extracted without using the case-control status. After using one of the linear combinations of genotypes or methylation values in each gene to capture gene-level signals, we regularize them to perform gene selection based on the biological network. Simulation studies show that the proposed approaches have higher true positive rates than using traditional dimension reduction techniques. We also apply our approaches to DNA methylation data and UK Biobank DNA sequence data for analyzing rheumatoid arthritis. The results show that the proposed methods can select potentially rheumatoid arthritis related genes that are missed by existing methods.

Author Summary

There is strong evidence showing that when genes are functionally related to each other in a genetic network, statistical methods utilizing prior biological network knowledge can outperform other methods that ignore genetic network structures. Therefore, statistical methods that can incorporate genetic network information into association analysis in human genetic association studies have been widely used since 2008. Here, we take advantage of recently developed methods to capture the gene-level signals in network-based penalized regression of high-dimensional genetic data. We have shown that the selection performance of our proposed methods can outperform three traditional principal component-based dimension reduction techniques in several simulation scenarios in terms of true positive rates. Meanwhile, by applying our methods in both DNA methylation data and DNA sequence data, the genes identified by our proposed methods can be significantly enriched into the rheumatoid arthritis pathway, such as genesHLA-DMA,HLA-DPB1, andHLA-DQA2in the HLA region.
Physiological trait networks enhance understanding of crop growth and water use in contrasting environments
Authors: Gleason Sean M. , Barnard Dave M. , Green Timothy R. , Mackay D. Scott , Wang Diane R. , Ainsworth Elizabeth A. , Altenhofen Jon , Brodribb Timothy J. , Cochard Hervé , Comas Louise H. , Cooper Mark , Creek Danielle , DeJonge Kendall C. , Delzon Sylvain , Fritschi Felix B. , Hammer Graeme , Hunter Cameron , Lombardozzi Danica , Messina Carlos D. , Ocheltree Troy , Stevens Bo Maxwell , Stewart Jared J. , Vadez Vincent , Wenz Joshua , Wright Ian J. , Yemoto Kevin , Zhang Huihui
Published: March 2022

Plant function arises from a complex network of structural and physiological traits. Explicit representation of these traits, as well as their connections with other biophysical processes, is required to advance our understanding of plant-soil-climate interactions. We used the Terrestrial Regional Ecosystem Exchange Simulator (TREES) to evaluate physiological trait networks in maize. Net primary productivity (NPP) and grain yield were simulated across five contrasting climate scenarios. Simulations achieving high NPP and grain yield in high precipitation environments featured trait networks conferring high water use strategies: deep roots, high stomatal conductance at low water potential (“risky” stomatal regulation), high xylem hydraulic conductivity, and high maximal leaf area index. In contrast, high NPP and grain yield was achieved in dry environments with low late-season precipitation via water conserving trait networks: deep roots, high embolism resistance, and low stomatal conductance at low leaf water potential (“conservative” stomatal regulation). We suggest that our approach, which allows for the simultaneous evaluation of physiological traits and their interactions (i.e., networks), has potential to improve crop growth predictions in different environments. In contrast, evaluating single traits in isolation of other coordinated traits does not appear to be an effective strategy for predicting plant performance.

Summary statement

Our process-based model uncovered two beneficial but contrasting trait networks for maize which can be understood by their integrated effect on water use/conservation. Modification of multiple, physiologically aligned, traits were required to bring about meaningful improvements in NPP and yield.
Reproductive barriers and genomic hotspots of adaptation during allopatric species divergence
Authors: Deshmukh Riddhi , Baral Saurav , Kuwalekar Muktai , Kizhakke Athulya Girish , Kunte Krushnamegh
Published: March 2022

Theory predicts that in allopatric populations, genomic divergence and reproductive barriers will be driven largely by random genetic drift, and thereby evolve slowly in large populations. However, local adaptation and divergence under selection may also play important roles, which remain poorly characterised. Here we address three key questions in young allopatric species: (a) How widespread are genomic signatures of adaptive divergence?, (b) What is the functional space along which young sister species show divergence at the genomic level?, and (c) How quickly might prezygotic and postzygotic reproductive barriers evolve? Analysis of 82 re-sequenced genomes of the OrientalPapilio polytesspecies group revealed surprisingly widespread hotspots of intense selection and selective sweeps at hundreds of genes unique to each species, and spanning all chromosomes, rather than divergence only in a few genomic islands. These genes perform diverse ecologically important adaptive functions such as wing development, colour patterning, courtship behaviour, mimicry, pheromone synthesis and olfaction, and host plant use and digestion of secondary metabolites, that could contribute to local adaptation and subsequent reproductive isolation. Divergence at such functional genes appeared to have reproductive consequences: behavioural and hybridisation experiments revealed strong assortative mate preference (prezygotic barriers) as well as postzygotic barriers to hybridisation in timespans as short as 1.27 my, indicating that speciation was already complete, rather than incipient. Our study thus demonstrates an underappreciated role of intense selection and potential local adaptation in creating genome-wide hotspots of rapid molecular evolution and divergence, during differentiation and speciation in young allopatric species.
Chemogenetic enhancement of cAMP signaling renders hippocampal synaptic plasticity resilient to the impact of acute sleep deprivation
Authors: Walsh Emily Nicole , Shetty Mahesh Shivarama , Diba Kamran , Abel Ted
Published: March 2022

Sleep facilitates memory storage and even brief periods of sleep loss lead to impairments in memory, particularly memories that are hippocampus dependent. In previous studies, we have shown that the deficit in memory seen after sleep loss is accompanied by deficits in synaptic plasticity. Our previous work has also found that sleep deprivation is associated with reduced levels of cyclic adenosine monophosphate (cAMP) in the hippocampus, and that the reduction of cAMP mediates the diminished memory performance. Based on these findings, we hypothesized that cAMP acts as a mediator for not only the cognitive deficits caused by sleep deprivation, but also the observed deficits in synaptic plasticity. In this study, we expressed the heterologousDrosophila melanogasterGαs-protein coupled octopamine receptor (DmOctβ1R) in mouse hippocampal neurons. This receptor is selectively activated by the systemically injected ligand (octopamine), thus allowing us to increase cAMP levels in hippocampal neurons during a five-hour sleep deprivation period. Our results show that chemogenetic enhancement of cAMP during the period of sleep deprivation prevents deficits in a persistent form of long-term potentiation (LTP) that is induced at the Schaffer collateral synapses in the hippocampal CA1 region. We also found that elevating cAMP levels only in the early or later half of sleep deprivation successfully prevented LTP deficits. These findings reveal that cAMP-dependent signaling pathways are key mediators of sleep deprivation at the synaptic level. Targeting these pathways could be useful in designing strategies to prevent the impact of sleep loss.

Significance statement

Insufficient sleep is an issue with significant health and socioeconomic implications. This includes a negative impact on memory consolidation. Previous studies in mice found that acute sleep deprivation leads to deficits in hippocampal synaptic plasticity and memory, which are associated with reduced levels of the signaling molecule cAMP. In this study, we used a chemogenetic strategy to enhance cAMP levels in specific hippocampal neurons during sleep deprivation. We found that this made synaptic plasticity resilient to the negative effects of sleep deprivation. These findings reveal that cAMP-dependent signaling pathways are key mediators of sleep deprivation and that targeting these pathways could be useful in designing strategies to prevent the impact of sleep loss.
A Microengineered Brain-Chip to Model Neuroinflammation in Humans
Authors: Pediaditakis I. , Kodella K. R. , Manatakis D. V. , Le C. Y. , Barthakur S. , Sorets A. , Gravanis A. , Ewart L. , Rubin L. L. , Manolakos E. S. , Hinojosa C. D. , Karalis K.
Published: March 2022

Species differences in the brain and the blood-brain barrier (BBB) biology hamper the translation from animal models to humans and impede the development of specific therapeutics for brain diseases. Here we present a human Brain-Chip engineered to recapitulate critical aspects of the complex brain cell-cell interactions that mediate neuroinflammation development. Our human organotypic microphysiological system (MPS) includes endothelial-like cells, pericytes, glia, and cortical neurons and maintains BBB permeability at in vivo relevant levels, providing a significant improvement in complexity and clinical mimicry compared to previous MPS models. This is the first report of a Brain-Chip with an RNA expression profile close to that of the adult human cortex and that demonstrates advantages over Transwell culture. Through perfusion of TNF-α, we recreated key inflammatory features, such as glia activation, the release of proinflammatory cytokines, and increased barrier permeability. Our model may provide a reliable tool for mechanistic studies in neuron-glial interactions and dysregulation of BBB function during neuroinflammation.
LIPA-driven hydrolysis of cholesteryl arachidonate promotes cancer metastasis via NF-κB
Authors: Chen Zhicong , Chen Fukai , Lee Hyeon Jeong , Zhang Meng , Yin Xianglin , Dai Mingji , Cheng Ji-Xin
Published: March 2022

Cholesteryl ester (CE) is an established marker in many types of aggressive cancers. Yet, the function of CE homeostasis during cancer progression is largely unknown. Here, enabled by Raman spectromicroscopy, pan-cancer bioinformatic analysis, and biocompatible Raman probe of cholesterol, LIPA was identified as the key regulator of CE hydrolysis in cancer cells. LIPA inhibition induced aberrant accumulation of CE-rich lipid droplet at single cell level. LIPA-driven CE hydrolysis was directly visualized by stimulated Raman scattering imaging of alkyne-tagged cholesterol. Cholesteryl arachidonate was identified as a dominant substrate that is rapidly hydrolyzed by LIPA. Inhibition of LIPA effectively suppressed cancer metastasis both in vitro and in vivo. Mechanistically, LIPA inhibition suppresses NF-κB signaling, while the NF-κB members positively regulate the expression level of LIPA, indicating regulation of CE homeostasis by a LIPA-CE-NFκB feedback loop. Collectively, our findings reveal that LIPA drives CE hydrolysis during cancer progression and is an important metabolic target for cancer therapy.
Asymmetrical lineage introgression and recombination in populations ofAspergillus flavus: implications for biological control
Authors: Molo Megan S. , White James B. , Cornish Vicki , Gell Richard M , Baars Oliver , Singh Rakhi , Carbone Mary Anna , Isakeit Thomas , Wise Kiersten A. , Woloshuk Charles P. , Bluhm Burton H. , Horn Bruce W. , Heiniger Ron W. , Carbone Ignazio
Published: March 2022

Aspergillus flavusis an agriculturally important fungus that causes ear rot of maize and produces aflatoxins (AFs), of which B1is the most potent carcinogen known. In the US, the management of AFs includes the deployment of biological control agents that comprise two nonaflatoxigenicA. flavusstrains, either Afla-Guard® (member of lineage IB) or AF36 (lineage IC). We used genotyping-by-sequencing to examine the influence of both biocontrol agents on native populations ofA. flavusin cornfields in Texas, North Carolina, Arkansas, and Indiana. This study examined up to 27,529 single-nucleotide polymorphisms (SNPs) in a total of 815A. flavusisolates, and 353 genome-wide haplotypes sampled before biocontrol application, three months after biocontrol application, and up to three years after initial application. Here, we report that the two distinctA. flavusevolutionary lineages IB and IC differ significantly in their frequency distributions across states. We provide evidence of increased unidirectional gene flow from lineage IB into IC, inferred to be due to the applied Afla-Guard biocontrol strain. Genetic exchange and recombination of biocontrol strains with native strains was detected in as little as three months after biocontrol application and up to one and three years later. There was limited inter-lineage migration in the untreated fields. These findings suggest that biocontrol products that include strains from lineage IB offer the greatest potential for sustained reductions in aflatoxin levels over several years. This knowledge has important implications for developing new biocontrol strategies.
High-throughput Production of Diverse Xenobiotic Metabolites with P450-transduced Huh7 Hepatoma Cell Lines
Authors: Lee Choon-Myung , Liu Ken H. , Singer Grant , Miller Gary W. , Li Shuzhao , Jones Dean P. , Morgan Edward T.
Published: March 2022

Precision medicine requires methods to assess drug metabolism and distribution, including the identification of known and undocumented drug and chemical exposures as well as their metabolites. Recent work demonstrated high-throughput generation of xenobiotic metabolites with human liver S-9 fractions and detection in human plasma and urine. Here, we developed a panel of lentivirally transduced human hepatoma cell lines (Huh7) that stably express individual cytochrome P450 (P450) enzymes and generate P450-specific xenobiotic metabolites. We verified protein expression by immunoblotting and demonstrated that the cell lines generate P450-specific metabolites from probe substrates. To increase analytical throughput, we used a pooling strategy where 36 chemicals were grouped into 12 unique mixtures, each mixture containing 6 randomly selected compounds, and each compound being present in two separate mixtures. Each mixture of compounds was incubated with 8 different P450 cell lines with cell extracts analyzed at 0 and 2 h. Extracts were analyzed using liquid chromatography-high resolution mass spectrometry. Cell lines selectively metabolized test substrates, with pazopanib metabolized by CYP3A4 and CYP2C8 cells, bupropion by CYP2B6, and β-naphthoflavone by CYP1A2 for example, showing substrate-enzyme specificity. Predicted metabolites from the remaining 33 compounds as well as many unidentified m/z features were detected. We also show that a specific metabolite generated by CYP2B6 cells, but not detected in the S9 system, was identified in human samples. Our data show that incubating these cell lines with chemical mixtures accelerated characterization of xenobiotic chemical space, while simultaneously allowing for the contributions of specific P450 enzymes to be identified.

Significance statement

High resolution mass spectrometry enables the identification of exposures to drugs and other xenobiotics in human samples. This paper demonstrates a workflow for high throughput production of xenobiotic metabolites using a panel of engineered cytochrome P450-expressing hepatoma cells. Active substrate-enzyme pairs can be identified using this workflow and generated metabolites can be used as surrogate standards to validate xenobiotic detection in humans.
Examination of diurnal variation and sex differences in hippocampal neurophysiology and spatial memory
Authors: Goode Lacy K. , Fusilier Allison R. , Remiszewski Natalie , Reeves Jacob M. , Abiraman Kavitha , Defenderfer Matthew , Paul Jodi R. , McMahon Lori L. , Gamble Karen L.
Published: March 2022

Circadian rhythms are biological processes that cycle across 24 hours and regulate many facets of neurophysiology, including learning and memory. Circadian variation in performance on spatial memory tasks is well-documented; however, the effect of sex across circadian time remains unclear. Additionally, little is known regarding the impact of time-of-day on hippocampal neuronal physiology. Here, we investigated the influence of both sex and time-of-day on hippocampal neurophysiology and memory.

Performance on the object location memory (OLM) task depended on both circadian time and sex, with memory enhanced at night in males but during the day in females. Long-term synaptic potentiation (LTP) magnitude at CA3-CA1 synapses was greater at night compared to day in both sexes. Next, we measured spontaneous synaptic excitation and inhibition onto CA1 pyramidal neurons. Frequency and amplitude of inhibition was greater during the day compared to night, regardless of sex. Frequency and amplitude of excitation was larger in females, compared to males, independent of time-of-day, although both time-of-day and sex influenced presynaptic release probability. At night, CA1 pyramidal neurons showed enhanced excitability (action potential firing and/or baseline potential) that was dependent on synaptic excitation and inhibition, regardless of sex. This study emphasizes the importance of sex and time-of-day in hippocampal physiology, especially given that many neurological disorders impacting the hippocampus are linked to circadian disruption and present differently in men and women. Knowledge about how sex and circadian rhythms affect hippocampal physiology can improve the translational relevancy of therapeutics and inform the appropriate timing of existing treatments.
The SARS-CoV-2 Delta variant induces an antibody response largely focused on class 1 and 2 antibody epitopes
Authors: Greaney Allison J. , Eguia Rachel T. , Starr Tyler N. , Khan Khadija , Franko Nicholas , Logue Jennifer K. , Lord Sandra M. , Speake Cate , Chu Helen Y. , Sigal Alex , Bloom Jesse D.
Published: March 2022

Exposure histories to SARS-CoV-2 variants and vaccinations will shape the specificity of antibody responses. To understand the specificity of Delta-elicited antibody immunity, we characterize the polyclonal antibody response elicited by primary or mRNA vaccine-breakthrough Delta infections. Both types of infection elicit a neutralizing antibody response focused heavily on the receptor-binding domain (RBD). We use deep mutational scanning to show that mutations to the RBD’s class 1 and class 2 epitopes, including sites 417, 478, and 484–486 often reduce binding of these Delta-elicited antibodies. The anti-Delta antibody response is more similar to that elicited by early 2020 viruses than the Beta variant, with mutations to the class 1 and 2, but not class 3 epitopes, having the largest effects on polyclonal antibody binding. In addition, mutations to the class 1 epitope (e.g., K417N) tend to have larger effects on antibody binding and neutralization in the Delta spike than in the D614G spike, both for vaccine- and Delta-infection-elicited antibodies. These results help elucidate how the antigenic impacts of SARS-CoV-2 mutations depend on exposure history.
ViReMa: A Virus Recombination Mapper of Next-Generation Sequencing data characterizes diverse recombinant viral nucleic acids
Authors: Sotcheff Stephanea , Zhou Yiyang , Sun Yan , Johnson John E. , Torbett Bruce E. , Routh Andrew L
Published: March 2022

Genetic recombination is a tremendous source of intra-host diversity in viruses and is critical for their ability to rapidly adapt to new environments or fitness challenges. While viruses are routinely characterized using high-throughput sequencing techniques, characterizing the genetic products of recombination in next-generation sequencing data remains a challenge. Viral recombination events can be highly diverse and variable in nature, including simple duplications and deletions, or more complex events such as copy/snap-back recombination, inter-virus or inter-segment recombination and insertions of host nucleic acids. Due to the variable mechanisms driving virus recombination and the different selection pressures acting on the progeny, recombination junctions rarely adhere to simple canonical sites or sequences. Furthermore, numerous different events may be present simultaneously in a viral population, yielding a complex mutational landscape. We have previously developed an algorithm called ViReMa (Virus Recombination Mapper) that bootstraps the bowtie short-read aligner to capture and annotate a wide-range of recombinant species found within virus populations. Here, we have updated ViReMa to provide an ‘error-density’ function designed to accurately detect recombination events in the longer reads now routinely generated by the Illumina platforms and provide output reports for multiple types of recombinant species using standardized formats. We demonstrate the utility and flexibility of ViReMa in different settings to report deletion events in simulated data from Flock House virus, copy-back RNA species in Sendai viruses, short duplication events in HIV, and virus to host recombination in an archaeal DNA virus.
Deep brain stimulation pulse sequences to optimally modulate frequency-specific neural activity
Authors: Farooqi Hafsa , Vitek Jerrold L. , Sanabria David Escobar
Published: March 2022

Precise neuromodulation systems are needed to identify the role of neural oscillatory dynamics in brain function, and to advance the development of electrical stimulation therapies tailored to each patient’s signature of brain dysfunction. Low-frequency, local field potentials (LFPs) are of increasing interest for the development of these systems because they reflect the synaptic inputs to a recorded neuronal population and can be chronically recorded in humans. Here, we identified stimulation pulse sequences to optimally minimize or maximize the 2-norm of frequency-specific LFP oscillations using a generalized mathematical model of spontaneous and stimulation-evoked LFP activity, and a subject-specific model of neural dynamics in the pallidum of a Parkinson’s disease patient. We leveraged convex and mixed-integer optimization tools to identify the pulse patterns, and employed constraints on the pulse frequency and amplitude that are required to keep electrical stimulation within its safety envelope. Our analysis revealed that a combination of phase, amplitude, and frequency pulse modulation is needed to attain optimal suppression or amplification of the targeted oscillations. Phase modulation is sufficient to modulate oscillations with a constant amplitude envelope. To attain optimal modulation for oscillations with a time-varying envelope, a trade-off between frequency and amplitude pulse modulation is needed. The optimized pulse sequences identified here were invariant to changes in the dynamics of stimulation-evoked neural activity, including the damping and natural frequency or complexity (i.e., generalized vs. patient-specific model). Our results reveal the structure of pulse sequences that can be used in closed-loop brain stimulation devices to control neural activity in real-time.

Significance Statement

This computational study reveals the structure of electrical pulse sequences that optimally modulate spontaneous, synaptic-related neural oscillations measured using local field potentials (LFPs). We considered constraints on the pulse amplitude and frequency, relevant to keep stimulation pulses within the safety envelope. Our mathematical analysis indicate that these optimized pulse sequences are invariant to changes in the studied neural dynamics, and a trade-off between amplitude and frequency pulse modulation is critical when the pulse amplitude is constrained. The pulse sequences identified here provide the rationale for the development of closed-loop neuromodulation systems that employ stimulation-evoked responses to control neural activity. These closed-loop systems will be key to characterize the role of neural activity in brain function and ultimately advance personalized neuromodulation devices.
A maximum-entropy model to predict 3D structural ensembles of chromatins from pairwise distances: Applications to Interphase Chromosomes and Structural Variants
Authors: Shi Guang , Thirumalai D.
Published: March 2022

The principles that govern the organization of genomes, which are needed for a deeper understanding of how chromosomes are packaged and function in eukaryotic cells, could be deciphered if the three dimensional (3D) structures are known. Recently, single-cell imaging experiments have determined the 3D coordinates of a number of loci in a chromosome. Here, we introduce a computational method (Distance Matrix to Ensemble of Structures, DIMES), based on the maximum entropy principle, with experimental pair-wise distances between loci as constraints, to generate a unique ensemble of 3D chromatin structures. Using the ensemble of structures, we quantitatively account for the distribution of pair-wise distances, three-body co-localization and higher-order interactions. We demonstrate that the DIMES method can be applied to both small length-scale and chromosome-scale imaging data to quantify the extent of heterogeneity and fluctuations in the shapes on various length scales. We develop a perturbation method that is used in conjunction with DIMES to predict the changes in 3D structures from structural variations. Our method also reveals quantitative differences between the 3D structures inferred from Hi-C and the ones measured in imaging experiments. Finally, the physical interpretation of the parameters extracted from DIMES provides insights into the origin of phase separation between euchromatin and heterochromatin domains.
A broad and potent neutralization epitope in SARS-related coronaviruses
Authors: Yuan Meng , Zhu Xueyong , He Wan-ting , Zhou Panpan , Kaku Chengzi I. , Capozzola Tazio , Zhu Connie Y. , Yu Xinye , Liu Hejun , Yu Wenli , Hua Yuanzi , Tien Henry , Peng Linghang , Song Ge , Cottrell Christopher A. , Schief William R. , Nemazee David , Walker Laura M. , Andrabi Raiees , Burton Dennis R. , Wilson Ian A.
Published: March 2022

Many neutralizing antibodies (nAbs) elicited to ancestral SARS-CoV-2 through natural infection and vaccination generally have reduced effectiveness to SARS-CoV-2 variants. Here we show therapeutic antibody ADG20 is able to neutralize all SARS-CoV-2 variants of concern (VOCs) including Omicron (B.1.1.529) as well as other SARS-related coronaviruses. We delineate the structural basis of this relatively escape-resistant epitope that extends from one end of the receptor binding site (RBS) into the highly conserved CR3022 site. ADG20 can then benefit from high potency through direct competition with ACE2 in the more variable RBS and interaction with the more highly conserved CR3022 site. Importantly, antibodies that are able to target this site generally neutralize all VOCs, albeit with reduced potency against Omicron. Thus, this highly conserved and vulnerable site can be exploited for design of universal vaccines and therapeutic antibodies.
Direct colorimetry of imipenem decomposition as a novel cost effective method for detecting carabapenamase producing bacteria
Authors: Kotsakis Stathis D. , Lambropoulou Anastasia , Miliotis Georgios , Tzelepi Eva , Miriagou Vivi , Tzouvelekis Leonidas S.
Published: March 2022

In the absence of a molecule that would collectively inhibit both metallo-β-lactamases and serine reactive carbapenemases, containment of their genes’ spreading is the main weapon currently available for confronting carbepenem resistance in hospitals. Cost effective methodologies rapidly detecting carbapenemase producing enterobacteria (CPE) would facilitate such measures. Herein a low cost CPE detection method was developed that was based on the direct colorimetry of the yellow shift caused by the accumulation of diketopiperazines – products of the acid catalyzed imipenem oligomerization – induced by carbapenemase action on dense solutions of imipenem/cilastatin. The reactions were studied by spectrophotometry in the visible spectrum using preparations of β-lactamases from the four molecular classes. The effects of various buffers on reactions containing the potent carbapenemases NDM-1 and NMC-A were monitored at 405 nm. Optimal conditions were used for the analysis of cell suspensions and the assay was evaluated using 38 selected enterobacteria including 29 CPE as well as nine carbapenemase-negative strains overexpressing other β-lactamases. The development of the yellow color was specific for carbapenemase containing enzyme preparations and the maximum intensity was achieved in acidic or un-buffered conditions in the presence of zinc. When applied on bacterial cell suspensions the assay could detect CPE with 96.7 % sensitivity and 100 % specificity with results being comparable to those obtained with the CARBA NP technique. Direct colorimetry of carbapenemase-induced imipenem decomposition required minimum reagents while exhibited high accuracy in detecting CPE. Therefore it should be considered for screening purposes after further clinical evaluation.

Importance

Currently, spread of multi-drug resistant (MDR) carbapenemase-producing enterobacteria (CPE), mostly in the clinical setting, is among the most pressing public health problems worldwide. In order to effectively control CPE, use of reliable and affordable methods detecting carbapenemase genes or the respective β-lactamases is of vital importance. Herein we developed a novel method, based on a previously undescribed phenomenon, which can detect CPE with few reagents by direct colorimetry of bacterial suspensions and imipenem/cilastatin mixtures.
Modulation of glucose metabolism by 2-Deoxy-D-Glucose (2DG) promotes IL-17 producing human T cell subsets
Authors: Chen Xin , Kozhaya Lina , Wang Chuxi , Placek Lindsey , Karhan Ece , Unutmaz Derya
Published: March 2022

Activation and differentiation of T cells are closely linked to their cellular metabolic programs. Glycolysis and mitochondrial metabolism are thought to be critical in modulating T cell function. Here we asked to what extent inhibition of glycolysis, using 2-Deoxy-D-Glucose (2DG), regulate activation, effector function, or differentiation of human T cell subsets. We found that glycolysis is required for T cell receptor (TCR) -mediated activation and proliferation of human naive CD4+ T cells but had less of an impact on memory subsets. CD4+ T cells cultured in the presence of 2DG displayed higher level of IL-17-secreting cells (Th17) from memory or in vitro differentiated naive regulatory T cell (Tregs) subsets. Moreover, the mucosal associated invariant T (MAIT) cell subset survived or expanded better and secreted higher IL-17 in the presence of 2DG. Remarkably, we found that the 2DG effect was reversed by mannose but not by glucose. Collectively, these findings suggest that 2DG could enrich IL-17 secreting human effector T cell subsets and their cellular functions. Our finding provides a framework to manipulate glycolytic pathways in human T cells in infectious diseases such as COVID19 and in enhancing cancer immunotherapy.
How large and diverse are field populations of fungal plant pathogens? The case of Zymoseptoria tritici
Authors: McDonald Bruce A. , Suffert Frederic , Bernasconi Alessio , Mikaberidze Alexey
Published: March 2022

Pathogen populations differ in the amount of genetic diversity they contain. Populations carrying higher genetic diversity are thought to have a greater evolutionary potential than populations carrying less diversity. We used published studies to estimate the range of values associated with two critical components of genetic diversity, the number of unique pathogen genotypes and the number of spores produced during an epidemic, for the septoria tritici blotch pathogen Zymoseptoria tritici. We found that wheat fields experiencing typical levels of infection are likely to carry between 3.2 and 14.9 million pathogen genotypes per hectare and produce at least 2.3 to 10.5 trillion pycnidiospores per hectare. Given the experimentally derived mutation rate of 3 × 10−10 substitutions per site per cell division, we estimate that between 28 and 130 million pathogen spores carrying adaptive mutations to counteract fungicides and resistant cultivars will be produced per hectare during a growing season. This suggests that most of the adaptive mutations that have been observed in Z. tritici populations can emerge through local selection from standing genetic variation that already exists within each field. The consequences of these findings for disease management strategies are discussed.