Carbon monoxide gas produced by a giant impact in the inner region of a young system.

Models of terrestrial planet formation predict that the final stages of planetary assembly-lasting tens of millions of years beyond the dispersal of young protoplanetary disks-are dominated by planetary collisions. It is through these giant impacts that planets like the young Earth grow to their final mass and achieve long-term stable orbital configurations1. A key prediction is that these impacts produce debris. So far, the most compelling observational evidence for post-impact debris comes from the planetary system around the nearby 23-million-year-old A-type star HD 172555. This system shows large amounts of fine dust with an unusually steep size distribution and atypical dust composition, previously attributed to either a hypervelocity impact2,3 or a massive asteroid belt4. Here we report the spectrally resolved detection of a carbon monoxide gas ring co-orbiting with dusty debris around HD 172555 between about six and nine astronomical units-a region analogous to the outer terrestrial planet region of our Solar System. Taken together, the dust and carbon monoxide detections favour a giant impact between large, volatile-rich bodies. This suggests that planetary-scale collisions, analogous to the Moon-forming impact, can release large amounts of gas as well as debris, and that this gas is observable, providing a window into the composition of young planets.

Adaptive nonequilibrium design of actin-based metamaterials: Fundamental and practical limits of control.

The adaptive and surprising emergent properties of biological materials self-assembled in far-from-equilibrium environments serve as an inspiration for efforts to design nanomaterials. In particular, controlling the conditions of self-assembly can modulate material properties, but there is no systematic understanding of either how to parameterize external control or how controllable a given material can be. Here, we demonstrate that branched actin networks can be encoded with metamaterial properties by dynamically controlling the applied force under which they grow and that the protocols can be selected using multi-task reinforcement learning. These actin networks have tunable responses over a large dynamic range depending on the chosen external protocol, providing a pathway to encoding "memory" within these structures. Interestingly, we obtain a bound that relates the dissipation rate and the rate of "encoding" that gives insight into the constraints on control-both physical and information theoretical. Taken together, these results emphasize the utility and necessity of nonequilibrium control for designing self-assembled nanostructures.

Climate change determines the sign of productivity trends in US forests.

Forests are integral to the global land carbon sink, which has sequestered ~30% of anthropogenic carbon emissions over recent decades. The persistence of this sink depends on the balance of positive drivers that increase ecosystem carbon storage-e.g., CO2 fertilization-and negative drivers that decrease it-e.g., intensifying disturbances. The net response of forest productivity to these drivers is uncertain due to the challenge of separating their effects from background disturbance-regrowth dynamics. We fit non-linear models to US forest inventory data (113,806 plot remeasurements in non-plantation forests from ~1999 to 2020) to quantify productivity trends while accounting for stand age, tree mortality, and harvest. Productivity trends were generally positive in the eastern United States, where climate change has been mild, and negative in the western United States, where climate change has been more severe. Productivity declines in the western United States cannot be explained by increased mortality or harvest; these declines likely reflect adverse climate-change impacts on tree growth. In the eastern United States, where data were available to partition biomass change into age-dependent and age-independent components, forest maturation and increasing productivity (likely due, at least in part, to CO2 fertilization) contributed roughly equally to biomass carbon sinks. Thus, adverse effects of climate change appear to overwhelm any positive drivers in the water-limited forests of the western United States, whereas forest maturation and positive responses to age-independent drivers contribute to eastern US carbon sinks. The future land carbon balance of forests will likely depend on the geographic extent of drought and heat stress.

Adaptation of SARS-CoV-2 to ACE2H353K mice reveals new spike residues that drive mouse infection.

The Coronavirus Disease 2019 (COVID-19) pandemic continues to cause extraordinary loss of life and economic damage. Animal models of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) infection are needed to better understand disease pathogenesis and evaluate preventive measures and therapies. While mice are widely used to model human disease, mouse angiotensin converting enzyme 2 (ACE2) does not bind the ancestral SARS-CoV-2 spike protein to mediate viral entry. To overcome this limitation, we "humanized" mouse Ace2 using CRISPR gene editing to introduce a single amino acid substitution, H353K, predicted to facilitate S protein binding. While H353K knockin Ace2 (mACE2H353K) mice supported SARS-CoV-2 infection and replication, they exhibited minimal disease manifestations. Following 30 serial passages of ancestral SARS-CoV-2 in mACE2H353K mice, we generated and cloned a more virulent virus. A single isolate (SARS2MA-H353K) was prepared for detailed studies. In 7-11-month-old mACE2H353K mice, a 104 PFU inocula resulted in diffuse alveolar disease manifested as edema, hyaline membrane formation, and interstitial cellular infiltration/thickening. Unexpectedly, the mouse-adapted virus also infected standard BALB/c and C57BL/6 mice and caused severe disease. The mouse-adapted virus acquired five new missense mutations including two in spike (K417E, Q493K), one each in nsp4, nsp9, and M and a single nucleotide change in the 5' untranslated region. The Q493K spike mutation arose early in serial passage and is predicted to provide affinity-enhancing molecular interactions with mACE2 and further increase the stability and affinity to the receptor. This new model and mouse-adapted virus will be useful to evaluate COVID-19 disease and prophylactic and therapeutic interventions.IMPORTANCEWe developed a new mouse model with a humanized angiotensin converting enzyme 2 (ACE2) locus that preserves native regulatory elements. A single point mutation in mouse ACE2 (H353K) was sufficient to confer in vivo infection with ancestral severe acute respiratory syndrome-coronavirus-2 virus. Through in vivo serial passage, a virulent mouse-adapted strain was obtained. In aged mACE2H353K mice, the mouse-adapted strain caused diffuse alveolar disease. The mouse-adapted virus also infected standard BALB/c and C57BL/6 mice, causing severe disease. The mouse-adapted virus acquired five new missense mutations including two in spike (K417E, Q493K), one each in nsp4, nsp9, and M and a single nucleotide change in the 5' untranslated region. The Q493K spike mutation arose early in serial passage and is predicted to provide affinity-enhancing molecular interactions with mACE2 and further increase the stability and affinity to the receptor. This new model and mouse-adapted virus will be useful to evaluate COVID-19 disease and prophylactic and therapeutic interventions.

Regulation of phosphoribosyl ubiquitination by a calmodulin-dependent glutamylase.

The bacterial pathogen Legionella pneumophila creates an intracellular niche permissive for its replication by extensively modulating host-cell functions using hundreds of effector proteins delivered by its Dot/Icm secretion system1. Among these, members of the SidE family (SidEs) regulate several cellular processes through a unique phosphoribosyl ubiquitination mechanism that bypasses the canonical ubiquitination machinery2-4. The activity of SidEs is regulated by another Dot/Icm effector known as SidJ5; however, the mechanism of this regulation is not completely understood6,7. Here we demonstrate that SidJ inhibits the activity of SidEs by inducing the covalent attachment of glutamate moieties to SdeA-a member of the SidE family-at E860, one of the catalytic residues that is required for the mono-ADP-ribosyltransferase activity involved in ubiquitin activation2. This inhibition by SidJ is spatially restricted in host cells because its activity requires the eukaryote-specific protein calmodulin (CaM). We solved a structure of SidJ-CaM in complex with AMP and found that the ATP used in this reaction is cleaved at the α-phosphate position by SidJ, which-in the absence of glutamate or modifiable SdeA-undergoes self-AMPylation. Our results reveal a mechanism of regulation in bacterial pathogenicity in which a glutamylation reaction that inhibits the activity of virulence factors is activated by host-factor-dependent acyl-adenylation.

Adaptive Monte Carlo augmented with normalizing flows.

SignificanceMonte Carlo methods, tools for sampling data from probability distributions, are widely used in the physical sciences, applied mathematics, and Bayesian statistics. Nevertheless, there are many situations in which it is computationally prohibitive to use Monte Carlo due to slow "mixing" between modes of a distribution unless hand-tuned algorithms are used to accelerate the scheme. Machine learning techniques based on generative models offer a compelling alternative to the challenge of designing efficient schemes for a specific system. Here, we formalize Monte Carlo augmented with normalizing flows and show that, with limited prior data and a physically inspired algorithm, we can substantially accelerate sampling with generative models.

Evaluating the impact of alcohol minimum unit pricing on deaths and hospitalisations in Scotland: a controlled interrupted time series study.

BACKGROUND: Since May 1, 2018, every alcoholic drink sold in Scotland has had minimum unit pricing (MUP) of £0·50 per unit. Previous studies have indicated that the introduction of this policy reduced alcohol sales by 3%. We aimed to assess whether this has led to reductions in alcohol-attributable deaths and hospitalisations. METHODS: Study outcomes, wholly attributable to alcohol consumption, were defined using routinely collected data on deaths and hospitalisations. Controlled interrupted time series regression was used to assess the legislation's impact in Scotland, and any effect modification across demographic and socioeconomic deprivation groups. The pre-intervention time series ran from Jan 1, 2012, to April 30, 2018, and for 32 months after the policy was implemented (until Dec 31, 2020). Data from England, a part of the UK where the intervention was not implemented, were used to form a control group. FINDINGS: MUP in Scotland was associated with a significant 13·4% reduction (95% CI -18·4 to -8·3; p=0·0004) in deaths wholly attributable to alcohol consumption. Hospitalisations wholly attributable to alcohol consumption decreased by 4·1% (-8·3 to 0·3; p=0·064). Effects were driven by significant improvements in chronic outcomes, particularly alcoholic liver disease. Furthermore, MUP legislation was associated with a reduction in deaths and hospitalisations wholly attributable to alcohol consumption in the four most socioeconomically deprived deciles in Scotland. INTERPRETATION: The implementation of MUP legislation was associated with significant reductions in deaths, and reductions in hospitalisations, wholly attributable to alcohol consumption. The greatest improvements were in the four most socioeconomically deprived deciles, indicating that the policy is positively tackling deprivation-based inequalities in alcohol-attributable health harm. FUNDING: Scottish Government.

Evaluating the impact of minimum unit pricing for alcohol in Scotland: a theory-based synthesis of the evidence.

BACKGROUND: In May 2018, the Scottish Government set a minimum unit price (MUP) of £0·50 per unit of alcohol sold in Scotland to reduce alcohol-related health harms. We synthesised evidence to establish the effects of MUP on alcohol-related health and social harms, at population level and within specific societal groups. METHODS: We did a theory-based synthesis of academic and grey research evidence about impacts of MUP in Scotland, including compliance, price, consumption, health outcomes, social outcomes, public attitudes, and the alcoholic drinks industry. We searched the Public Health Scotland's MUP evaluation portfolio and relevant grey and academic literature for studies published between Jan 1, 2018, and Jan 31, 2023. We conducted systematic searches and screening of bibliographic databases (Scopus, Public Health Database, EconLit, MEDLINE, ProQuest Public Health, Social Policy and Practice, NHS Scotland Knowledge Network Library Search, medRxiv, bioRxiv, SSRN, Idox Knowledge Exchange, Social Policy & Practice, and Google Search). Search terms were tailored to specific databases but included variants of the terms "minimum unit pricing", "alcohol", and "policy". Eligibility literature included English-language research into impacts of MUP on either the population of Scotland or a specific subpopulation. We excluded conference abstracts, literature reviews, articles that did not report research, and research based solely on data from before the introduction of MUP. FINDINGS: We included 40 reports in our analysis. On the balance of evidence, MUP improved population-level health outcomes, demonstrated most starkly by a 13·4% reduction in alcohol-attributable deaths in Scotland compared with England. There was no evidence of substantial negative effects on the alcoholic drinks industry or social harms at the population level. While population-level outcomes were predominantly positive, some qualitative evidence suggests that MUP might have exacerbated health and social harms for some individuals or groups, especially those with alcohol dependence who were financially vulnerable. INTERPRETATION: MUP in Scotland has been effective in reducing alcohol-related health harms, with little evidence of any effect on social harms. If MUP continues, policymakers should consider raising the £0·50 per unit threshold and supplementing the intervention with policies or services to address any unintended negative effects experienced by specific groups. The synthesis is persuasive due to the prospective, theory-based design of the evaluation portfolio and the quality and comprehensiveness of the evidence. FUNDING: Scottish Government.

Threshold and Laser Conversion in Nanostructured-Resonator Parametric Oscillators.

We explore optical parametric oscillation (OPO) in nanophotonic resonators, enabling arbitrary, nonlinear phase matching and nearly lossless control of energy conversion. Such pristine OPO laser converters are determined by nonlinear light-matter interactions, making them both technologically flexible and broadly reconfigurable. We utilize a nanostructured inner-wall modulation in the resonator to achieve universal phase matching for OPO-laser conversion, but coherent backscattering also induces a counterpropagating pump laser. This depletes the intraresonator optical power in either direction, increasing the OPO threshold power and limiting laser-conversion efficiency, the ratio of optical power in target signal and idler frequencies to the pump. We develop an analytical model of this system that emphasizes an understanding of optimal laser-conversion and threshold behaviors, and we use the model to guide experiments with nanostructured-resonator OPO laser-conversion circuits, fully integrated on chip and unlimited by group-velocity dispersion. Our Letter demonstrates the fundamental connection between OPO laser-conversion efficiency and the resonator coupling rate, subject to the relative phase and power of counterpropagating pump fields. We achieve (40±4) mW of on-chip power, corresponding to (41±4)% conversion efficiency, and discover a path toward near-unity OPO laser-conversion efficiency.

Is breakfast consumption detrimental, unnecessary, or an opportunity for health promotion? A review of cardiometabolic outcomes and functional food choices.

Breakfast consumption is generally considered a health-promoting habit for cardiometabolism, particularly with regard to chrononutrition. Glucose uptake is enhanced by proper insulin secretion triggered by the pancreatic clock, averting metabolic dysregulation related to insulin resistance. Breakfast skipping, in turn, is often considered a behaviour detrimental to health, in part due to putative inverse metabolic actions compared to breakfast consumption, such that breakfast skipping may promote circadian desynchrony. However, most ill health concerns about breakfast skipping are inferred from observational research, and recent well-controlled randomized clinical trials have shown benefits of breakfast skipping for cardiovascular risk factors. Accordingly, this review describes the effects of breakfast consumption versus breakfast skipping on cardiovascular risk factors (blood pressure and glycaemic and lipid indices). In addition, the view of breakfast consumption as an opportunity for functional food ingestion is considered to provide further insights into decision-making practice. Collectively, both breakfast consumption and breakfast skipping can be considered viable habits, but they depend on individual preferences, planning, and the specific foods being consumed or omitted. When consumed, breakfast should consist primarily of functional foods typical for this meal (e.g., eggs, dairy products, nuts, fruits, whole grains, coffee, tea, etc.). While breakfast consumption aligns with chrononutrition principles, breakfast skipping can contribute to a calorie deficit over time, which has the potential for widespread cardiometabolic benefits for patients with overweight/obesity. The concepts and practical considerations discussed in the present review may aid health care personnel in personalising breakfast consumption recommendations for diverse patient populations.

Remembering the Work of Phillip L. Geissler: A Coda to His Scientific Trajectory.

Phillip L. Geissler made important contributions to the statistical mechanics of biological polymers, heterogeneous materials, and chemical dynamics in aqueous environments. He devised analytical and computational methods that revealed the underlying organization of complex systems at the frontiers of biology, chemistry, and materials science. In this retrospective we celebrate his work at these frontiers.

Microscopic origin of tunable assembly forces in chiral active environments.

Across a variety of spatial scales, from nanoscale biological systems to micron-scale colloidal systems, equilibrium self-assembly is entirely dictated by-and therefore limited by-the thermodynamic properties of the constituent materials. In contrast, nonequilibrium materials, such as self-propelled active matter, expand the possibilities for driving the assemblies that are inaccessible in equilibrium conditions. Recently, a number of works have suggested that active matter drives or accelerates self-organization, but the emergent interactions that arise between solutes immersed in actively driven environments are complex and poorly understood. Here, we analyze and resolve two crucial questions concerning actively driven self-assembly: (i) how, mechanistically, do active environments drive self-assembly of passive solutes? (ii) Under which conditions is this assembly robust? We employ the framework of odd hydrodynamics to theoretically explain numerical and experimental observations that chiral active matter, i.e., particles driven with a directional torque, produces robust and long-ranged assembly forces. Together, these developments constitute an important step towards a comprehensive theoretical framework for controlling self-assembly in nonequilibrium environments.

Comparisons between dual-energy X-ray absorptiometry and bioimpedance devices for appendicular lean mass and muscle quality in Hispanic adults.

The purpose of this study was to compare single- and multi-frequency bioimpedance (BIA) devices against dual-energy X-ray absorptiometry (DXA) for appendicular lean mass (ALM) and muscle quality index (MQI) metrics in Hispanic adults. One hundred thirty-one Hispanic adults (18-55 years) participated in this study. ALM was measured with single-frequency bioimpedance analysis (SFBIA), multi-frequency bioimpedance analysis (MFBIA) and DXA. ALMTOTAL (left arm + right arm + left leg + right leg) and ALMARMS (left arm + right arm) were computed for all three devices. Handgrip strength (HGS) was measured using a dynamometer. The average HGS was used for all MQI models (highest left hand + highest right hand)/2. MQIARMS was defined as the ratio between HGS and ALMARMS. MQITOTAL was established as the ratio between HGS and ALMTOTAL. SFBIA and MFBIA had strong correlations with DXA for all ALM and MQI metrics (Lin's concordance correlation coefficient values ranged from 0·86 (MQIMFBIA-ARMS) to 0·97 (Arms LMSFBIA); all P < 0·001). Equivalence testing varied between methods (e.g. SFBIA v. DXA) when examining the different metrics (i.e. ALMTOTAL, ALMARMS, MQITOTAL and MQIARMS). MQIARMS was the only metric that did not differ from the line of identity and had no proportional bias when comparing all the devices against each other. The current study findings demonstrate good overall agreement between SFBIA, MFBIA and DXA for ALMTOTAL and ALMARMS in a Hispanic population. However, SFBIA and MFBIA have better agreement with DXA when used to compute MQIARMS than MQITOTAL.

Rhodiola rosea as an adaptogen to enhance exercise performance: a review of the literature.

Rhodiola rosea (RR) is a plant whose bioactive components may function as adaptogens, thereby increasing resistance to stress and improving overall resilience. Some of these effects may influence exercise performance and adaptations. Based on studies of rodents, potential mechanisms for the ergogenic effects of RR include modulation of energy substrate stores and use, reductions in fatigue and muscle damage and altered antioxidant activity. At least sixteen investigations in humans have explored the potential ergogenicity of RR. These studies indicate acute RR supplementation (∼200 mg RR containing ∼1 % salidroside and ∼3 % rosavin, provided 60 min before exercise) may prolong time-to-exhaustion and improve time trial performance in recreationally active males and females, with limited documented benefits of chronic supplementation. Recent trials providing higher doses (∼1500 to 2400 mg RR/d for 4­30 d) have demonstrated ergogenic effects during sprints on bicycle ergometers and resistance training in trained and untrained adults. The effects of RR on muscle damage, inflammation, energy system modulation, antioxidant activity and perceived exertion are presently equivocal. Collectively, it appears that adequately dosed RR enhances dimensions of exercise performance and related outcomes for select tasks. However, the current literature does not unanimously show that RR is ergogenic. Variability in supplementation dose and duration, concentration of bioactive compounds, participant characteristics, exercise tests and statistical considerations may help explain these disparate findings. Future research should build on the longstanding use of RR and contemporary clinical trials to establish the conditions in which supplementation facilitates exercise performance and adaptations.

Clinical specificity of two assays for immunoglobulin kappa and lambda free light chains.

OBJECTIVES: Free light chain (FLC) assays and the ratio of κ/λ are recommended for diagnosis, prognosis and monitoring of plasma cell dyscrasias (PCD). Limited data exists on FLC clinical specificity in patients diagnosed with other conditions. METHODS: We assessed the κ, λ, and κ/λ FLC ratio using the FreeLite assay and the Sebia FLC ELISA assay in 176 patients with clinical presentations of fatigue, anemia, polyclonal hypergammaglobulinemia, joint disorders, kidney disease and non PCD-cancers with no monoclonal protein observed on serum protein electrophoresis or MASS-FIX immunoglobulin isotyping. Manufacturer defined reference intervals (RI) and glomerular filtration rate (GFR) specific RI (renal RI) were utilized. RESULTS: For the κ/λ ratio, 68.7 % (121/176) of specimens on the FreeLite and 87.5 % (154/176) of specimens on the Sebia assay were within RI. For κ, 68.2 % (120/176) and 72.2 % (127/176) of results were outside RI for FreeLite and Sebia respectively. For λ, 37.5 % (66/176) and 84.1 % (148/176) of FreeLite and Sebia results were outside RI. With FreeLite and Sebia, patients with kidney disease (n=25) had the highest κ/λ ratios. 44 patients (25.0 %) had GFR <60 mL/min/BSA. When renal RI were applied, 13.6 % had a FLCr outside the renal RI with FreeLite, and 4.5 % with Sebia. CONCLUSIONS: In a cohort of patients with signs and symptoms suggestive of PCDs, but ultimately diagnosed with other conditions, Sebia FLC had improved clinical specificity relative to FreeLite, if one was using an abnormal κ/λ ratio as a surrogate for monoclonality.

Liposomal delivery enhances absorption of vitamin C into plasma and leukocytes: a double-blind, placebo-controlled, randomized trial.

PURPOSE: L-Ascorbic acid (vitamin C) is an essential water-soluble vitamin that plays an important role in various physiological functions, including immune health. The stability of vitamin C in the gastrointestinal tract its bioavailability is limited. This study aimed to investigate if a liposomal form of vitamin C can increase absorption compared to standard vitamin C. METHODS: In a randomized, double-blind, placebo-controlled, crossover fashion, 19 males and 8 females (n = 27; 36.0 ± 5.1 years, 165.0 ± 6.9 cm, 70.6 ± 7.1 kg) ingested a single-dose of placebo (PLA), 500 mg vitamin C (VIT C), and 500 mg liposomal vitamin C (LV-VIT C, LipoVantage®, Specnova, LLC, Tyson Corner, VA, USA). Venous blood samples were collected 0, 0.5-, 1-, 1.5-, 2-, 3-, 4-, 6-, 8-, 12-, and 24-hours after ingestion and were analyzed for plasma and leukocyte vitamin C concentration. RESULTS: VIT C and LV-VIT C demonstrated significantly greater Cmax and AUC0 - 24 in plasma and in leukocytes compared to placebo (p < 0.001). Additionally, LV-VIT C had significantly higher Cmax (plasma + 27%, leukocytes + 20%, p < 0.001) and AUC0 - 24 (plasma + 21%, leukocytes + 8%, p < 0.001) values as compared to VIT C. CONCLUSION: Liposomal formulation of vitamin C increases absorption into plasma and leukocytes. TRIAL REGISTRATION: Clinical Trials Registry - India (CTRI/2023/04/051789).

Mutant phenotypes for thousands of bacterial genes of unknown function.

One-third of all protein-coding genes from bacterial genomes cannot be annotated with a function. Here, to investigate the functions of these genes, we present genome-wide mutant fitness data from 32 diverse bacteria across dozens of growth conditions. We identified mutant phenotypes for 11,779 protein-coding genes that had not been annotated with a specific function. Many genes could be associated with a specific condition because the gene affected fitness only in that condition, or with another gene in the same bacterium because they had similar mutant phenotypes. Of the poorly annotated genes, 2,316 had associations that have high confidence because they are conserved in other bacteria. By combining these conserved associations with comparative genomics, we identified putative DNA repair proteins; in addition, we propose specific functions for poorly annotated enzymes and transporters and for uncharacterized protein families. Our study demonstrates the scalability of microbial genetics and its utility for improving gene annotations.

Primary care usage at the end of life: a retrospective cohort study of cancer patients using linked primary and hospital care data.

PURPOSE: Health service use is most intensive in the final year of a person's life, with 80% of this expenditure occurring in hospital. Close involvement of primary care services has been promoted to enhance quality end-of-life care that is appropriate to the needs of patients. However, the relationship between primary care involvement and patients' use of hospital care is not well described. This study aims to examine primary care use in the last year of life for cancer patients and its relationship to hospital usage. METHODS: Retrospective cohort study in Victoria, Australia, using linked routine care data from primary care, hospital and death certificates. Patients were included who died related to cancer between 2008 and 2017. RESULTS: A total of 758 patients were included, of whom 88% (n = 667) visited primary care during the last 6 months (median 9.1 consultations). In the last month of life, 45% of patients were prescribed opioids, and 3% had imaging requested. Patients who received home visits (13%) or anticipatory medications (15%) had less than half the median bed days in the last 3 months (4 vs 9 days, p < 0.001, 5 vs 10 days, p = 0.001) and 1 month of life (0 vs 2 days, p = 0.002, 0 vs 3 days, p < 0.001), and reduced emergency department presentations (32% vs 46%, p = 0.006, 31% vs 47% p < 0.001) in the final month. CONCLUSION: This study identifies two important primary care processes-home visits and anticipatory medication-associated with reduced hospital usage and intervention at the end of life.

Computing equilibrium free energies through a nonequilibrium quench.

Many methods to accelerate sampling of molecular configurations are based on the idea that temperature can be used to accelerate rare transitions. These methods typically compute equilibrium properties at a target temperature using reweighting or through Monte Carlo exchanges between replicas at higher temperatures. A recent paper [G. M. Rotskoff and E. Vanden-Eijnden, Phys. Rev. Lett. 122, 150602 (2019)] demonstrated that accurate equilibrium densities of states can also be computed through a nonequilibrium "quench" process, where sampling is performed at a higher temperature to encourage rapid mixing and then quenched to lower energy states with dissipative dynamics. Here, we provide an implementation of the quench dynamics in LAMMPS and evaluate a new formulation of nonequilibrium estimators for the computation of partition functions or free energy surfaces (FESs) of molecular systems. We show that the method is exact for a minimal model of N-independent harmonic springs and use these analytical results to develop heuristics for the amount of quenching required to obtain accurate sampling. We then test the quench approach on alanine dipeptide, where we show that it gives an FES that is accurate near the most stable configurations using the quench approach but disagrees with a reference umbrella sampling calculation in high FE regions. We then show that combining quenching with umbrella sampling allows the efficient calculation of the free energy in all regions. Moreover, by using this combined scheme, we obtain the FES across a range of temperatures at no additional cost, making it much more efficient than standard umbrella sampling if this information is required. Finally, we discuss how this approach can be extended to solute tempering and demonstrate that it is highly accurate for the case of solvated alanine dipeptide without any additional modifications.

Controlled Stiffness of Direct-Write, Near-Field Electrospun Gelatin Fibers Generates Differences in Tenocyte Morphology and Gene Expression.

Tendinopathy is a leading cause of mobility issues. Currently, the cell-matrix interactions involved in the development of tendinopathy are not fully understood. In vitro tendon models provide a unique tool for addressing this knowledge gap as they permit fine control over biochemical, micromechanical, and structural aspects of the local environment to explore cell-matrix interactions. In this study, direct-write, near-field electrospinning of gelatin solution was implemented to fabricate micron-scale fibrous scaffolds that mimic native collagen fiber size and orientation. The stiffness of these fibrous scaffolds was found to be controllable between 1 MPa and 8 MPa using different crosslinking methods (EDC, DHT, DHT+EDC) or through altering the duration of crosslinking with EDC (1 h to 24 h). EDC crosslinking provided the greatest fiber stability, surviving up to 3 weeks in vitro. Differences in stiffness resulted in phenotypic changes for equine tenocytes with low stiffness fibers (∼1 MPa) promoting an elongated nuclear aspect ratio while those on high stiffness fibers (∼8 MPa) were rounded. High stiffness fibers resulted in the upregulation of matrix metalloproteinase (MMPs) and proteoglycans (possible indicators for tendinopathy) relative to low stiffness fibers. These results demonstrate the feasibility of direct-written gelatin scaffolds as tendon in vitro models and provide evidence that matrix mechanical properties may be crucial factors in cell-matrix interactions during tendinopathy formation.