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Abstract High-throughput screening techniques are pivotal to unlocking the mysteries of biology. Yet, the promise of droplet microfluidics in enabling single-cell resolution, ultra-high-throughput screening remains largely unfulfilled. Droplet sorting errors caused by polydisperse droplet sizes that are often inevitable in multi-step assays have severely limited the effectiveness and utility of this technique, especially when screening large libraries. Even a relatively low 1% sorting error results in 10,000 false calls in a 1,000,000 droplet screen, imposing an unreasonably large burden on downstream validation. Here, we present NOVAsort (Next-generation Opto-Volume-based Accurate droplet sorter), a device capable of discerning droplets based on both size and fluorescence intensity. With a 1000- and 10,000-fold reduction in false positives and false negatives, respectively. NOVAsort addresses the challenges of conventional droplet sorting approaches and sets standards for accuracy and throughput in droplet microfluidic assays.

Introduction Current dietary assessment methods face challenges in accurately capturing individuals’ dietary habits, undermining the efficacy of public health strategies. The ‘Standardised and Objective Dietary Intake Assessment Tool’ (SODIAT)-1 study aims to assess the effectiveness of three emerging technologies (urine and capillary blood biomarkers, and wearable camera technology) and two online self-reporting dietary assessment tools to monitor dietary intake. Methods This randomised controlled crossover trial will recruit 30 participants (aged 18-70 years and BMI of 20-30 kg/m 2) from Imperial College London and the University of Reading. Exclusion criteria include recent weight change, food allergies/intolerances, following restrictive diets, certain health conditions and medication use. Interested volunteers will be directed to an online screening questionnaire via REDCap and eligible participants will attend a pre-study visit. Volunteers will consume, in a random order, two highly-controlled diets (compliant and non-compliant with UK guidelines) for four days each. Each study arm will be separated by at least one-week. During each test period, dietary intake will be monitored continuously using wearable cameras and self-recorded using eNutri (food frequency questionnaire) and Intake24 (24-hour dietary recall). Urine and capillary blood samples will be collected for biomarker analysis. Data analysis will assess the accuracy of dietary reporting across these methods using Lin’s concordance correlation coefficient. Discussion and ethical considerations This study introduces a novel approach to dietary assessment, addressing the limitations of traditional methods by reducing misreporting and enhancing inclusivity, particularly for underrepresented populations with literacy or language barriers. However, challenges persist, such as variability in biomarker data due to failure to adhere to sample storage requirements and the practicalities of continuously wearing cameras. To protect privacy, participants will be instructed to remove cameras at inappropriate times, and artificial intelligence will be used to blur all images captured apart from food.

Introduction Distinguishing patients with intracerebral haemorrhage (ICH) from other suspected stroke cases in the prehospital setting is crucial for determining the appropriate level of care and minimising the onset-to-treatment time, thereby potentially improving outcomes. Therefore, we developed prehospital prediction models to identify patients with ICH among suspected stroke cases.Methods Data were obtained from the Field Administration of Stroke Therapy-Magnesium prehospital stroke trial, where paramedics evaluated multiple variables in suspected stroke cases within the first 2 hours from the last known well time. A total of 19 candidate predictors were included to minimise overfitting and were subsequently refined through the backward exclusion of non-significant predictors. We used logistic regression and eXtreme Gradient Boosting (XGBoost) models to evaluate the performance of the predictors. Model performance was assessed using the area under the receiver operating characteristic curve (AUC), confusion matrix metrics and calibration measures. Additionally, models were internally validated and corrected for optimism through bootstrapping. Furthermore, a nomogram was built to facilitate paramedics in estimating the probability of ICH.Results We analysed 1649 suspected stroke cases, of which 373 (23%) were finally diagnosed with ICH. From the 19 candidate predictors, 9 were identified as independently associated with ICH (p

Abstract Vaccine development targeting SARS-CoV-2 in 2020 was of critical importance in reducing COVID-19 severity and mortality. In the U.K. during the initial roll-out most individuals either received two doses of Pfizer COVID-19 vaccine (BNT162b2) or the adenovirus-based vaccine from Oxford/AstraZeneca (ChAdOx1-nCoV-19). There are conflicting data as to the impact of age, sex and body habitus on cellular and humoral responses to vaccination, and most studies in this area have focused on determinants of mRNA vaccine immunogenicity. Here, we studied a cohort of participants in a population-based longitudinal study (COVIDENCE UK) to determine the influence of age, sex, body mass index (BMI) and pre-vaccination anti-Spike (anti-S) antibody status on vaccine-induced humoral and cellular immune responses to two doses of BNT162b2 or ChAdOx-n-CoV-19 vaccination. Younger age and pre-vaccination anti-S seropositivity were both associated with stronger antibody responses to vaccination. BNT162b2 generated higher neutralising and anti-S antibody titres to vaccination than ChAdOx1-nCoV-19, but cellular responses to the two vaccines were no different. Irrespective of vaccine type, increasing age was also associated with decreased frequency of cytokine double-positive CD4+T cells. Increasing BMI was associated with reduced frequency of SARS-CoV-2-specific TNF+CD8% T cells for both vaccines. Together, our findings demonstrate that increasing age and BMI are associated with attenuated cellular and humoral responses to SARS-CoV-2 vaccination. Whilst both vaccines induced T cell responses, BNT162b2 induced significantly elevated humoral immune response as compared to ChAdOx-n-CoV-19.

Objective To determine whether weekly oral vitamin D supplementation influences grip strength, explosive leg power, cardiorespiratory fitness or spirometric lung volumes in Mongolian schoolchildren.Methods Multicentre, randomised, placebo-controlled clinical trial conducted in children aged 6–13 years at baseline attending 18 schools in Ulaanbaatar. The intervention was weekly oral doses of 14 000 IU vitamin D3 (n=4418) or placebo (n=4433) for 3 years. Outcome measures were grip strength, standing long jump distance and serum 25-hydroxyvitamin D (25(OH)D) concentrations (determined in all participants), peak oxygen uptake (VO2peak, determined in a subset of 632 participants using 20 m multistage shuttle run tests) and spirometric outcomes (determined in a subset of 1343 participants).Results 99.8% of participants had serum 25(OH)D concentrations

Objective To determine whether vitamin D supplementation influences grip strength, explosive leg power, cardiorespiratory fitness and risk of exercise-induced bronchoconstriction (EIB) in South African schoolchildren.Methods Substudy (n=450) in Cape Town schoolchildren aged 8–11 years nested within a phase 3 randomised placebo-controlled trial (ViDiKids). The intervention was weekly oral doses of 10 000 IU vitamin D3 (n=228) or placebo (n=222) for 3 years. Outcome measures were serum 25-hydroxyvitamin D3 (25(OH)D3) concentrations, grip strength, standing long jump distance, peak oxygen uptake (VO2peak, determined using 20 m multistage shuttle run tests) and the proportion of children with EIB, measured at end-study.Results 64.7% of participants had serum 25(OH)D3concentrations

Summary: Obesity and fatty liver diseases—metabolic dysfunction-associated steatotic liver disease (MASLD) and metabolic dysfunction-associated steatohepatitis (MASH)—affect over one-third of the global population and are exacerbated in individuals with reduced functional aldehyde dehydrogenase 2 (ALDH2), observed in approximately 560 million people. Current treatment to prevent disease progression to cancer remains inadequate, requiring innovative approaches. We observe that Aldh2−/− and Aldh2−/−Sptbn1+/− mice develop phenotypes of human metabolic syndrome (MetS) and MASH with accumulation of endogenous aldehydes such as 4-hydroxynonenal (4-HNE). Mechanistic studies demonstrate aberrant transforming growth factor β (TGF-β) signaling through 4-HNE modification of the SMAD3 adaptor SPTBN1 (β2-spectrin) to pro-fibrotic and pro-oncogenic phenotypes, which is restored to normal SMAD3 signaling by targeting SPTBN1 with small interfering RNA (siRNA). Significantly, therapeutic inhibition of SPTBN1 blocks MASH and fibrosis in a human model and, additionally, improves glucose handling in Aldh2−/− and Aldh2−/−Sptbn1+/− mice. This study identifies SPTBN1 as a critical regulator of the functional phenotype of toxic aldehyde-induced MASH and a potential therapeutic target.

Abstract Magnetic hysteresis measurements are routinely made in the Earth and planetary sciences to identify geologically meaningful magnetic recorders, and to study variations in present and past environments. Interpreting magnetic hysteresis data in terms of domain state and paleomagnetic stability are major motivations behind undertaking these measurements, but the interpretations remain fraught with challenges and ambiguities. To shed new light on these ambiguities, we have undertaken a systematic micromagnetic study to quantify the magnetic hysteresis behavior of room‐temperature magnetite as a function of particle size (45–195 nm; equivalent spherical volume diameter) and shape (oblate, prolate and equant); our models span uniformly magnetized single domain (SD) to non‐uniformly magnetized single vortex (SV) states. Within our models the reduced magnetization associated with SV particles marks a clear boundary between SD (≥0.5) and SV (

Abstract The ability of rocks to hold a reliable record of the ancient geomagnetic field depends on the structure and stability of magnetic domain‐states contained within constituent particles. In paleomagnetic studies, the Day plot is an easily constructed graph of magnetic hysteresis parameters that is frequently used to estimate the likely magnetic recording stability of samples. Often samples plot in the region of the Day plot attributed to so‐called pseudo‐single‐domain particles with little understanding of the implications for domain‐states or recording fidelity. Here we use micromagnetic models to explore the hysteresis parameters of magnetite particles with idealized prolate and oblate truncated‐octahedral geometries containing single domain (SD), single‐vortex and occasionally multi‐vortex states. We show that these domain states exhibit a well‐defined trend in the Day plot that extends from the SD region well into the multi‐domain region, all of which are likely to be stable remanence carriers. We suggest that although the interpretation of the Day plot and its variants might be subject to ambiguities, if the magnetic mineralogy is known, it can still provide some useful insights about paleomagnetic specimens' dominant domain state, average particle sizes and, consequently, their paleomagnetic stability.

Objective To determine whether weekly oral vitamin D supplementation influences growth, body composition, pubertal development or spirometric outcomes in South African schoolchildren.Design Phase 3 double-blind randomised placebo-controlled trial.Setting Socioeconomically disadvantaged peri-urban district of Cape Town, South Africa.Participants 1682 children of black African ancestry attending government primary schools and aged 6–11 years at baseline.Interventions Oral vitamin D3 (10 000 IU/week) versus placebo for 3 years.Main outcome measures Height-for-age and body mass index-for-age, measured in all participants; Tanner scores for pubertal development, spirometric lung volumes and body composition, measured in a subset of 450 children who additionally took part in a nested substudy.Results Mean serum 25-hydroxyvitamin D3 concentration at 3-year follow-up was higher among children randomised to receive vitamin D versus placebo (104.3 vs 64.7 nmol/L, respectively; mean difference (MD) 39.7 nmol/L, 95% CI 37.6 to 41.9 nmol/L). No statistically significant differences in height-for-age z-score (adjusted MD (aMD) −0.08, 95% CI −0.19 to 0.03) or body mass index-for-age z-score (aMD −0.04, 95% CI −0.16 to 0.07) were seen between vitamin D versus placebo groups at follow-up. Among substudy participants, allocation to vitamin D versus placebo did not influence pubertal development scores, % predicted forced expiratory volume in 1 s (FEV1), % predicted forced vital capacity (FVC), % predicted FEV1/FVC, fat mass or fat-free mass.Conclusions Weekly oral administration of 10 000 IU vitamin D3 boosted vitamin D status but did not influence growth, body composition, pubertal development or spirometric outcomes in South African schoolchildren.Trial registration numbers ClinicalTrials.gov NCT02880982, South African National Clinical Trials Register DOH-27-0916-5527.

Abstract Micromagnetic modeling allows the systematic study of the effects of particle size and shape on the first‐order reversal curve (FORC) magnetic hysteresis response for magnetite particles in the single‐domain (SD) and pseudo‐single domain (PSD) particle size range. The interpretation of FORCs, though widely used, has been highly subjective. Here, we use micromagnetics to model randomly oriented distributions of particles to allow more physically meaningful interpretations. We show that one commonly found type of PSD particle—namely the single vortex (SV) particle—has far more complex signals than SD particles, with multiple peaks and troughs in the FORC distribution, where the peaks have higher switching fields for larger SV particles. Particles in the SD to SV transition zone have the lowest switching fields. Symmetrical and prolate particles display similar behavior, with distinctive peaks forming near the vertical axis of the FORC diagram. In contrast, highly oblate particles produce “butterfly” structures, suggesting that these are potentially diagnostic of particle morphology. We also consider FORC diagrams for distributions of particle sizes and shapes and produce an online application that users can use to build their own FORC distributions. There is good agreement between the model predictions for distributions of particle sizes and shapes, and the published experimental literature.

Abstract The active rift zones in Iceland provide unique insight into the geodynamic processes of divergent plate boundaries. The geodynamics of Iceland are studied intensively, particularly, by geophysical methods sensitive to active and/or visible structures such as earthquake seismic and Synthetic Aperture Radar observations or aerial photographs. However, older and less active structures, that may exert a strong control on the presently active geodynamics, are often buried beneath recent volcanic or sedimentary deposits and are—due to their passive mode—overseen by the typical geophysical investigations. Aeromagnetic surveys provide spatial information about subsurface magnetization contrasts relating to both active and inactive structures. However, the aeromagnetic data in Iceland were collected in the 1970-80s and are relevant only to large-scale regional rift studies. With the availability of reliable drones and light-weight atomic scalar sensors, high-quality drone magnetic surveys can provide an unprecedented spatial resolution of both active and passive structures of rift systems as compared to conventional airborne surveys. Here, we present the results of a drone-towed magnetic scalar field and scalar gradiometry study of the north-northeast trending Bárðarbunga spreading center to the north of the Vatnajökull ice cap, Iceland. Our results provide new information about the structural complexity of rift zones with evidence of densely-spaced, conjugate and oblique faults throughout the area. Evidence is shown of a hitherto unknown and prominent east-northeast trending fault structure that coincides with the northern tip of the main eruption edifice of the 1797 and 2014-15 Holuhraun volcanic events. We suggest that this pre-existing structure controlled the locus of vertical magma migration during the two Holuhraun events.

Abstract Drugs that target pre-mRNA splicing hold great therapeutic potential, but the quantitative understanding of how these drugs work is limited. Here we introduce mechanistically interpretable quantitative models for the sequence-specific and concentration-dependent behavior of splice-modifying drugs. Using massively parallel splicing assays, RNA-seq experiments, and precision dose-response curves, we obtain quantitative models for two small-molecule drugs, risdiplam and branaplam, developed for treating spinal muscular atrophy. The results quantitatively characterize the specificities of risdiplam and branaplam for 5’ splice site sequences, suggest that branaplam recognizes 5’ splice sites via two distinct interaction modes, and contradict the prevailing two-site hypothesis for risdiplam activity at SMN2 exon 7. The results also show that anomalous single-drug cooperativity, as well as multi-drug synergy, are widespread among small-molecule drugs and antisense-oligonucleotide drugs that promote exon inclusion. Our quantitative models thus clarify the mechanisms of existing treatments and provide a basis for the rational development of new therapies.

Abstract Tailwater recovery (TWR) systems, which divert phosphorus‐rich drainage water from cranberry (Vaccinium macrocarpon Ait.) farms into reservoirs, have the potential to improve water quality of freshwater lakes in Massachusetts. However, residents and environmentalists have questioned the potential water quality benefits of TWR systems. In the southeastern United States, research shows that TWR systems decrease agricultural inputs of phosphorus (P) to surface water by 23%–92%. Additionally, a case study in Massachusetts demonstrated the efficacy of TWR and other best management practices in reducing P concentrations in White Island Pond. Although TWR systems appear effective as part of a P management strategy, more research is needed to quantify their environmental benefits and allay public concerns. We propose filling three critical research gaps to strengthen and broaden support for TWR systems in cranberry agriculture in Massachusetts: (1) quantifying physical properties, (2) quantifying water storage potential, and (3) quantifying P retention capacity. Core Ideas Excess phosphorus (P) from cranberry farms may contribute to the eutrophication of freshwater lakes. Agricultural tailwater recovery (TWR) systems are increasingly used to conserve water and improve water quality. Earth removal, critical to create TWR ponds and provide sand for cranberry farms, has drawn public opposition. Research and case studies indicate TWR systems may decrease P inputs from cranberry farms to surface water. Long‐term monitoring, research, and stakeholder engagement are needed to assess efficacy of cranberry TWR systems.

Summary: Receptor tyrosine kinases (RTKs) control stem cell maintenance vs. differentiation decisions. Casitas B-lineage lymphoma (CBL) family ubiquitin ligases are negative regulators of RTKs, but their stem cell regulatory roles remain unclear. Here, we show that Lgr5+ intestinal stem cell (ISC)-specific inducible Cbl-knockout (KO) on a Cblb null mouse background (iDKO) induced rapid loss of the Lgr5Hi ISCs with transient expansion of the Lgr5Lo transit-amplifying population. LacZ-based lineage tracing revealed increased ISC commitment toward enterocyte and goblet cell fate at the expense of Paneth cells. Functionally, Cbl/Cblb iDKO impaired the recovery from radiation-induced intestinal epithelial injury. In vitro, Cbl/Cblb iDKO led to inability to maintain intestinal organoids. Single-cell RNA sequencing in organoids identified Akt-mTOR (mammalian target of rapamycin) pathway hyperactivation upon iDKO, and pharmacological Akt-mTOR axis inhibition rescued the iDKO defects. Our results demonstrate a requirement for Cbl/Cblb in the maintenance of ISCs by fine-tuning the Akt-mTOR axis to balance stem cell maintenance vs. commitment to differentiation.