Plant-based transient overexpression systems enable rapid and scalable production of subunit vaccines. Previously, we have shown that cholera toxin B subunit (CTB), an oral cholera vaccine antigen, ...is N-glycosylated upon expression in transgenic Nicotiana benthamiana. Here, we found that overexpression of aglycosylated CTB by agroinfiltration of a tobamoviral vector causes massive tissue necrosis and poor accumulation unless retained in the endoplasmic reticulum (ER). However, the re-introduction of N-glycosylation to its original or an alternative site significantly relieved the necrosis and provided a high CTB yield without ER retention. Quantitative gene expression analysis of PDI, BiP, bZIP60, SKP1, 26Sα proteasome and PR1a, and the detection of ubiquitinated CTB polypeptides revealed that N-glycosylation significantly relieved ER stress and hypersensitive response, and facilitated the folding/assembly of CTB. The glycosylated CTB (gCTB) was characterized for potential vaccine use. Glycan profiling revealed that gCTB contained approximately 38% plant-specific glycans. gCTB retained nanomolar affinity to GM1-ganglioside with only marginal reduction of physicochemical stability and induced an anti-cholera holotoxin antibody response comparable to native CTB in a mouse oral immunization study. These findings demonstrated gCTB's potential as an oral immunogen and point to a potential role of N-glycosylation in increasing recombinant protein yields in plants.
Eccentric quasi-isometric (EQI) contractions (maintaining a yielding contraction for as long as possible, beyond task failure) have gained interest in research and applied settings. However, little ...is known regarding the biomechanical profile of EQIs. Fourteen well-trained males performed four maximal effort knee-extensor EQIs, separated by 180 seconds. Angular impulse, velocity, and time-under-tension through the 30-100º range of motion (ROM), and in eight ROM brackets were quantified. Statistical parametric mapping, analyses of variance, and standardised effects (Hedges' g (ES), %Δ) detected between-contraction joint-angle-specific differences in time-normalised and absolute variables. Mean velocity was 1.34º·s
−1
with most (62.5 ± 4.9%) of the angular impulse imparted between 40-70º. Most between-contraction changes occurred between 30-50º (p≤ 0.067, ES = 0.53 ± 0.31, 60 ± 52%), while measures remained constant between 50-100º (= 0.069-0.83, ES = 0.10 ± 0.26, 14.3 ± 24.6%). EQIs are a time-efficient means to impart high cumulative mechanical tension, especially at short to medium muscle lengths. However, angular impulse distribution shifts towards medium to long muscle lengths with repeat contractions. Practitioners may utilise EQIs to emphasize the initial portion of the ROM, and limit ROM, or apply EQIs in a fatigued state to emphasize longer muscle lengths.
Young exoplanets are snapshots of the planetary evolution process. Planets that orbit stars in young associations are particularly important because the age of the planetary system is well ...constrained. We present the discovery of a transiting planet larger than Neptune but smaller than Saturn in the 45 Myr Tucana-Horologium young moving group. The host star is a visual binary, and our follow-up observations demonstrate that the planet orbits the G6V primary component, DS Tuc A (HD 222259A, TIC 410214986). We first identified transits using photometry from the Transiting Exoplanet Survey Satellite (TESS; alerted as TOI 200.01). We validated the planet and improved the stellar parameters using a suite of new and archival data, including spectra from Southern Astrophysical Research/Goodman, South African Extremely Large Telescope/High Resolution Spectrograph and Las Cumbres Observatories/Network of Robotic Echelle Spectrographs; transit photometry from Spitzer; and deep adaptive optics imaging from Gemini/Gemini Planet Imager. No additional stellar or planetary signals are seen in the data. We measured the planetary parameters by simultaneously modeling the photometry with a transit model and a Gaussian process to account for stellar variability. We determined that the planetary radius is 5.70 0.17 R⊕ and that the orbital period is 8.1 days. The inclination angles of the host star's spin axis, the planet's orbital axis, and the visual binary's orbital axis are aligned within 15° to within the uncertainties of the relevant data. DS Tuc Ab is bright enough (V = 8.5) for detailed characterization using radial velocities and transmission spectroscopy.
US Hispanic/Latino individuals are diverse in genetic ancestry, culture, and environmental exposures. Here, we characterized and controlled for this diversity in genome-wide association studies ...(GWASs) for the Hispanic Community Health Study/Study of Latinos (HCHS/SOL). We simultaneously estimated population-structure principal components (PCs) robust to familial relatedness and pairwise kinship coefficients (KCs) robust to population structure, admixture, and Hardy-Weinberg departures. The PCs revealed substantial genetic differentiation within and among six self-identified background groups (Cuban, Dominican, Puerto Rican, Mexican, and Central and South American). To control for variation among groups, we developed a multi-dimensional clustering method to define a “genetic-analysis group” variable that retains many properties of self-identified background while achieving substantially greater genetic homogeneity within groups and including participants with non-specific self-identification. In GWASs of 22 biomedical traits, we used a linear mixed model (LMM) including pairwise empirical KCs to account for familial relatedness, PCs for ancestry, and genetic-analysis groups for additional group-associated effects. Including the genetic-analysis group as a covariate accounted for significant trait variation in 8 of 22 traits, even after we fit 20 PCs. Additionally, genetic-analysis groups had significant heterogeneity of residual variance for 20 of 22 traits, and modeling this heteroscedasticity within the LMM reduced genomic inflation for 19 traits. Furthermore, fitting an LMM that utilized a genetic-analysis group rather than a self-identified background group achieved higher power to detect previously reported associations. We expect that the methods applied here will be useful in other studies with multiple ethnic groups, admixture, and relatedness.
To test the hypothesis that a longer length of time between diagnosis of hypertensive disorders of pregnancy and delivery is associated with increased risk of cardiovascular morbidity in the years ...after delivery.
This is a retrospective cohort study based in the New York State Inpatient Database. The first delivery for all patients from 2005 to 2014 who delivered preterm with an International Classification of Diseases, 9th Revision, Clinical Modification code for hypertensive disorders of pregnancy (excluding isolated chronic hypertension) was included. The duration between diagnosis and delivery was divided into 7 days or less or more than 7 days. The primary outcome was admission for a composite of cardiovascular disease, stroke, or death after the index delivery through December 31, 2014.
There were 22,594 patients with a median follow-up period of 5.2 years: 19,750 (87.4%) were delivered within 7 days of diagnosis and 2,844 (12.6%) were delivered more than 7 days from diagnosis. The primary outcome occurred in 216 (1.1%) patients in the 0-7 days group (21 events/10,000 person-years) and 67 (2.4%) patients in the more than 7 days group (46 events/10,000 person-years), adjusted hazard ratio 1.45 (95% CI 1.09 to 1.93). The findings were robust in a number of sensitivity analyses.
Prolonged expectant management of preterm hypertensive disorders of pregnancy is associated with an increased risk of maternal cardiac disease in the ensuing years.
The atherosclerotic plaque microenvironment is highly complex, and selective agents that modulate plaque stability are not yet available. We sought to develop a scRNA-seq analysis workflow to ...investigate this environment and uncover potential therapeutic approaches. We designed a user-friendly, reproducible workflow that will be applicable to other disease-specific scRNA-seq datasets.
Here we incorporated automated cell labeling, pseudotemporal ordering, ligand-receptor evaluation, and drug-gene interaction analysis into a ready-to-deploy workflow. We applied this pipeline to further investigate a previously published human coronary single-cell dataset by Wirka et al. Notably, we developed an interactive web application to enable further exploration and analysis of this and other cardiovascular single-cell datasets.
We revealed distinct derivations of fibroblast-like cells from smooth muscle cells (SMCs), and showed the key changes in gene expression along their de-differentiation path. We highlighted several key ligand-receptor interactions within the atherosclerotic environment through functional expression profiling and revealed several avenues for future pharmacological development for precision medicine. Further, our interactive web application, PlaqView (www.plaqview.com), allows lay scientists to explore this and other datasets and compare scRNA-seq tools without prior coding knowledge.
This publicly available workflow and application will allow for more systematic and user-friendly analysis of scRNA datasets in other disease and developmental systems. Our analysis pipeline provides many hypothesis-generating tools to unravel the etiology of coronary artery disease. We also highlight potential mechanisms for several drugs in the atherosclerotic cellular environment. Future releases of PlaqView will feature more scRNA-seq and scATAC-seq atherosclerosis-related datasets to provide a critical resource for the field, and to promote data harmonization and biological interpretation.
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•Automated cell annotation and pseudotemporal analyses reveal expression changes in de-differentiated smooth muscle cells (SMC).•Ligand-receptor profiling reveals potential drug targets that modulate SMC and fibroblast signaling during atherosclerosis.•Reproducible workflow allows for detailed evaluation of scRNA-seq datasets with minimal coding and eliminates potential bias.•PlaqView web application allows for interactive analysis of scRNA-seq datasets and facilitates data sharing and reanalysis.
As our understanding of the impact of specific molecular properties on applications in discovery‐based disciplines improves, the extent to which published synthetic methods meet (or do not meet) ...desirable criteria is ever clearer. Herein, we show how the application of simple (and in many cases freely available) computational tools can be used to develop a semiquantitative understanding of the potential of new methods to support molecular discovery. This analysis can, among other things, inform the design of improved substrate scoping studies; direct the prioritization of specific exemplar structures for synthesis; and substantiate claims of potential future applications for new methods.
The property ladder: Careful control of molecular properties is closely linked to success in small‐molecule‐led discovery ventures. Computational tools to calculate molecular properties are now widely available, many of them as freeware. These tools can be used to help guide the application of new synthetic methods and to evaluate alignment with future discovery needs.
AbstractObjectiveTo develop and validate a pragmatic risk score to predict mortality in patients admitted to hospital with coronavirus disease 2019 (covid-19).DesignProspective observational cohort ...study.SettingInternational Severe Acute Respiratory and emerging Infections Consortium (ISARIC) World Health Organization (WHO) Clinical Characterisation Protocol UK (CCP-UK) study (performed by the ISARIC Coronavirus Clinical Characterisation Consortium—ISARIC-4C) in 260 hospitals across England, Scotland, and Wales. Model training was performed on a cohort of patients recruited between 6 February and 20 May 2020, with validation conducted on a second cohort of patients recruited after model development between 21 May and 29 June 2020.ParticipantsAdults (age ≥18 years) admitted to hospital with covid-19 at least four weeks before final data extraction.Main outcome measureIn-hospital mortality.Results35 463 patients were included in the derivation dataset (mortality rate 32.2%) and 22 361 in the validation dataset (mortality rate 30.1%). The final 4C Mortality Score included eight variables readily available at initial hospital assessment: age, sex, number of comorbidities, respiratory rate, peripheral oxygen saturation, level of consciousness, urea level, and C reactive protein (score range 0-21 points). The 4C Score showed high discrimination for mortality (derivation cohort: area under the receiver operating characteristic curve 0.79, 95% confidence interval 0.78 to 0.79; validation cohort: 0.77, 0.76 to 0.77) with excellent calibration (validation: calibration-in-the-large=0, slope=1.0). Patients with a score of at least 15 (n=4158, 19%) had a 62% mortality (positive predictive value 62%) compared with 1% mortality for those with a score of 3 or less (n=1650, 7%; negative predictive value 99%). Discriminatory performance was higher than 15 pre-existing risk stratification scores (area under the receiver operating characteristic curve range 0.61-0.76), with scores developed in other covid-19 cohorts often performing poorly (range 0.63-0.73).ConclusionsAn easy-to-use risk stratification score has been developed and validated based on commonly available parameters at hospital presentation. The 4C Mortality Score outperformed existing scores, showed utility to directly inform clinical decision making, and can be used to stratify patients admitted to hospital with covid-19 into different management groups. The score should be further validated to determine its applicability in other populations.Study registrationISRCTN66726260
Technology utilizing human induced pluripotent stem cells (iPS cells) has enormous potential to provide improved cellular models of human disease. However, variable genetic and phenotypic ...characterization of many existing iPS cell lines limits their potential use for research and therapy. Here we describe the systematic generation, genotyping and phenotyping of 711 iPS cell lines derived from 301 healthy individuals by the Human Induced Pluripotent Stem Cells Initiative. Our study outlines the major sources of genetic and phenotypic variation in iPS cells and establishes their suitability as models of complex human traits and cancer. Through genome-wide profiling we find that 5-46% of the variation in different iPS cell phenotypes, including differentiation capacity and cellular morphology, arises from differences between individuals. Additionally, we assess the phenotypic consequences of genomic copy-number alterations that are repeatedly observed in iPS cells. In addition, we present a comprehensive map of common regulatory variants affecting the transcriptome of human pluripotent cells.
Abstract
The spread of neurofibrillary tau pathology in Alzheimer disease (AD) mostly follows a stereotypical pattern of topographical progression but atypical patterns associated with ...interhemispheric asymmetry have been described. Because histopathological studies that used bilateral sampling are limited, this study aimed to assess interhemispheric tau pathology differences and the presence of topographically atypical cortical spreading patterns. Immunohistochemical staining for detection of tau pathology was performed in 23 regions of interest in 57 autopsy cases comparing bilateral cortical regions and hemispheres. Frequent mild (82% of cases) and occasional moderate (32%) interhemispheric density discrepancies were observed, whereas marked discrepancies were uncommon (7%) and restricted to occipital regions. Left and right hemispheric tau pathology dominance was observed with similar frequencies, except in Braak Stage VI that favored a left dominance. Interhemispheric Braak stage differences were observed in 16% of cases and were more frequent in advanced (IV–VI) versus early (I–III) stages. One atypical lobar topographical pattern in which occipital tau pathology density exceeded frontal lobe scores was identified in 4 cases favoring a left dominant asymmetry. We speculate that asymmetry and atypical topographical progression patterns may be associated with atypical AD clinical presentations and progression characteristics, which should be tested by comprehensive clinicopathological correlations.
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