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Coral reef development is intricately linked to both climate and the concentration of atmospheric CO2, specifically through temperature and carbonate chemistry in the upper ocean. In turn, the calcification of corals modifies the concentration of dissolved inorganic carbon (DIC) and total alkalinity in the ocean, impacting air–sea gas exchange, atmospheric CO2 concentration, and ultimately the climate. This feedback between atmospheric conditions and coral biogeochemistry can only be accounted for with a coupled coral–carbon–climate model. Here we present the implementation of a coral reef calcification module into an Earth system model. Simulated coral reef production of the calcium carbonate mineral aragonite depends on photosynthetically active radiation, nutrient concentrations, salinity, temperature, and the aragonite saturation state. An ensemble of 210 parameter perturbation simulations was performed to identify carbonate production parameter values that optimize the simulated distribution of coral reefs and associated carbonate production. The tuned model simulates the presence of coral reefs and regional-to-global carbonate production values in good agreement with data-based estimates, despite some limitations due to the imperfect simulation of climatic and biogeochemical fields driving the simulation of coral reef development. When used in association with methods accounting for bathymetry changes resulting from different sea levels, the model enables assessment of past and future coral–climate coupling on seasonal to millennial timescales, highlighting how climatic trends and variability may affect reef development and the resulting climate–carbon feedback.

Understanding cellular ultrastructure is tightly bound to microscopic resolution and the ability to identify individual components at that resolution. Expansion microscopy has revolutionised this topic. Here we present and compare two protocols of ultrastructure expansion microscopy that allow for 4.5-fold mostly isotropic expansion and the use of antibodies, metabolic labelling, and DNA stains to demarcate individual regions such as the endoplasmic reticulum, the nuclei, the peripheral endocytic compartments as well as the ventral disc and the cytoskeleton in Giardia lamblia. We present an optimised, shortened, and modular protocol that can be swiftly adjusted to the investigators needs in this important protozoan model organism.

The ubiquitous Torque teno virus (TTV) establishes a chronically persistent infection in the human host. TTV has not been associated with any apparent disease, but, as part of the human virome, it may confer a regulatory imprint on the human immune system with as yet unclear consequences. However, so far, only few studies have characterized the TTV-specific immune responses or the overall immunological imprints by TTV. Here, we reveal that TTV infection leads to a highly exhausted TTV-specific CD8+ T-cell response, hallmarked by decreased IFN-γ production and the expression of the inhibitory NKG2A-receptor. On a functional level, we identified a panel of highly polymorphic TTV-encoded peptides that lead to an expansion of regulatory NKG2A+ natural killer, NKG2A+CD4+, and NKG2A+CD8+ T cells via the stabilization of the non-classical HLA-E molecule. Our results thus demonstrate that TTV leads to a distinct imprint on the human immune system that may further regulate overall human immune responses in infectious, autoimmune, and malignant diseases.

Here, we present the case of a patient with a metastatic neuroendocrine tumor with cytologically negative ascites treated for spontaneous bacterial peritonitis (SBP). Ascitic cultures remained negative for bacterial growth but were positive for Candida albicans 8 days after SBP diagnosis. ß-D-glucan was only positive in ascites, while being negative in blood. Blood cultures remained negative throughout the whole admission. Fungal peritonitis presumably originated from an impending bowl perforation or an increasing vascular permeability caused by an increase in VEGF secondary to diffuse infiltration by the underlying malignant disease.

This study explores eight chief enrollment officers' lived experiences. Semi-structured interviews were conducted with the eight chief enrollment officers at Ohio's public and private four-year institutions addressing two research questions. First, "what experiences influence a chief enrollment officer's ability to lead," produced six key findings: taking on more responsibility in their previous job, recruiting during COVID-19, working through a technological revolution, benefitting from professional development, embracing mentorship, and having time spent in the role. Second, "what competencies are essential for a chief enrollment management officer to be successful," produced eight key findings: understanding enrollment management trends and practices, understanding and utilizing data, having an ability to lead, providing successful supervision, being technologically savvy, working with presidents and campus partners, and recognizing one's own weaknesses.

Introduction Discrimination is known to have different effects on health. In particular the mental health of affected people diminishes. Although it is known that marginalized groups are discriminated against more, at present only research on gender and ethnicity has been done. Further diversity domains like socioeconomic status, care responsibilities, sexual orientation, disability, mental and physical health, and their intersections have been scarcely looked at. Objectives The aim of the study was to determine the effects of discrimination on the mental health for employees and students of a university hospital taking diversity domains into account. Methods A web-based survey between June 22 to October 23 was conducted using the PHQ-4 and WHO-5 as well as innovative Diversity Minimal Item set to measure different diversity domains. Results Preliminary data shows that discrimination among employees and students is common, widespread and most frequent based on gender, ethnicity and health. The mental health of those who feel discriminated against tends to be poorer, especially looking at the intersectionality of diversity domains. Conclusions The results of this study suggest that both more measures to prevent discrimination in a university hospital have to be implemented and individuals from marginalized groups need special psychosocial support to ensure a safer working environment. In addition, greater attention to diversity and inclusion in medical research is needed to develop appropriate responses and interventions, including diversity policies. Disclosure of Interest None Declared

Introduction Herpes simplex virus (HSV) is frequently detected in the respiratory tract of mechanically ventilated patients and is associated with a worse outcome. The aim of this study is to determine whether antiviral therapy in HSV-positive patients improves outcome.Methods and analysis Prospective, multicentre, open-label, randomised, controlled trial in parallel-group design. Adult, mechanically ventilated patients with pneumonia and HSV type 1 detected in bronchoalveolar lavage (≥105 copies/mL) are eligible for participation and will be randomly allocated (1:1) to receive acyclovir (10 mg/kg body weight every 8 hours) for 10 days (or until discharge from the intensive care unit if earlier) or no intervention (control group). The primary outcome is mortality measured at day 30 after randomisation (primary endpoint) and will be analysed with Cox mixed-effects model. Secondary endpoints include ventilator-free and vasopressor-free days up to day 30. A total of 710 patients will be included in the trial.Ethics and dissemination The trial was approved by the responsible ethics committee and by Germany’s Federal Institute for Drugs and Medical Devices. The clinical trial application was submitted under the new Clinical Trials Regulation through CTIS (The Clinical Trials Information System). In this process, only one ethics committee, whose name is unknown to the applicant, and Germany’s Federal Institute for Drugs and Medical Devices are involved throughout the entire approval process. Results will be published in a journal indexed in MEDLINE and CTIS. With publication, de-identified, individual participant data will be made available to researchers.Trial registration number NCT06134492.

Abstract Type IIA topoisomerases are essential DNA processing enzymes that must robustly and reliably relax DNA torsional stress. While cellular processes constantly create varying torsional stress, how this variation impacts type IIA topoisomerase function remains obscure. Using multiple single-molecule approaches, we examined the torsional dependence of eukaryotic topoisomerase II (topo II) activity on naked DNA and chromatin. We observed that topo II is ~50-fold more processive on buckled DNA than previously estimated. We further discovered that topo II relaxes supercoiled DNA prior to plectoneme formation, but with processivity reduced by ~100-fold. This relaxation decreases with diminishing torsion, consistent with topo II capturing transient DNA loops. Topo II retains high processivity on buckled chromatin (~10,000 turns) and becomes highly processive even on chromatin under low torsional stress (~1000 turns), consistent with chromatin’s predisposition to readily form DNA crossings. This work establishes that chromatin is a major stimulant of topo II function.

IntroductionAerobic exercise has been shown to improve cancer-associated cognitive decline (CACD) in breast cancer survivors (BCS), and recent findings suggest that one mechanism by which exercise may reduce cognitive decline is through alteration of the brain's functional organization. Many cognitive abilities and measures of functional brain organization change with age and disease, typically reflected in cognitive decline and reduced differentiation of brain networks, or “modularity.” Although previous research has identified associations between lifestyle interventions, such as exercise, and increased modularity, no studies have examined these relationships in cancer populations. The primary aim of this study was to investigate the preliminary effects of a 12-week aerobic exercise program on changes in brain network modularity in BCS. As a secondary aim, we explored correlations between changes in modularity with moderate-to-vigorous physical activity (MVPA) and cognitive function. Data were exploratory and used for hypothesis generation for a future, larger study.MethodsParticipants included a subsample of 10 BCS (M age = 65.9 ± 9.3 years) from a larger pilot study (N = 30 BCS) who were randomized to a 12-week aerobic exercise program (AE) or usual care (UC). The present study collected brain magnetic resonance imaging, Actigraph accelerometry, and cognitive task performance at baseline and 3-month follow-up (i.e., post-intervention; n = 4 AE, n = 6 UC). Intervention effects on modularity, MVPA, and cognition were quantified as magnitude of change between groups (Cohen's d). Changes in modularity were further explored via paired t-tests within groups. Associations between changes in modularity, MVPA, and cognitive performance were explored using Spearman's correlations.ResultsThe magnitude of changes in modularity between groups were small-to-moderate and favored the AE group (d = 0.23 to d = 0.67 across thresholds). Paired t-tests revealed a significant increase in modularity in the AE group from baseline to 3-month follow-up (t = 3.08, p = 0.03, d = 1.17), but not in the UC group. The correlation between changes in MVPA and changes in modularity were not statistically significant (r = 0.36, p = 0.39), and correlations between modularity and cognitive performance yielded mixed effects by cognitive domain.DiscussionFindings suggest that aerobic exercise may influence functional brain network organization and cognition in BCS. These data warrant further investigation in larger exercise trials.

Malignant transformation of human cells is associated with their re-programming which results in uncontrolled proliferation and in the same time biochemical activation of immunosuppressive pathways which form cancer immune evasion machinery. However, there is no conceptual understanding of whether immune evasion machinery pathways and expression of immune checkpoint proteins form a part of the process of malignant transformation or if they are triggered by T lymphocytes and natural killers (NK) attempting to attack cells which are undergoing or already underwent malignant transformation. To address this fundamental question, we performed experimental malignant transformation of BEAS-2B human bronchial epithelium cells and RC-124 non-malignant human kidney epithelial cells using bracken extracts containing carcinogenic alkaloid called ptaquiloside. This transformation led to a significant upregulation of cell proliferation velocity and in the same time led to a significant upregulation in expression of key immune checkpoint proteins – galectin-9, programmed death ligand 1 (PD-L1), indoleamine 2,3-dioxygenase (IDO1). Their increased expression levels were in line with upregulation of the levels and activities of HIF-1 transcription complex and transforming growth factor beta type 1 (TGF-β)-Smad3 signalling pathway. When co-cultured with T cells, transformed epithelial cells displayed much higher and more efficient immune evasion activity compared to original non-transformed cells. Therefore, this work resolved a very important scientific and clinical question and suggested that cancer immune evasion machinery is activated during malignant transformation of human cells regardless the presence of immune cells in microenvironment.

Abstract Personalization of ICU nutrition is essential to future of critical care. Recommendations from American/European guidelines and practice suggestions incorporating recent literature are presented. Low-dose enteral nutrition (EN) or parenteral nutrition (PN) can be started within 48 h of admission. While EN is preferred route of delivery, new data highlight PN can be given safely without increased risk; thus, when early EN is not feasible, provision of isocaloric PN is effective and results in similar outcomes. Indirect calorimetry (IC) measurement of energy expenditure (EE) is recommended by both European/American guidelines after stabilization post-ICU admission. Below-measured EE (~ 70%) targets should be used during early phase and increased to match EE later in stay. Low-dose protein delivery can be used early (~ D1-2) (

Abstract Transposases are ubiquitous enzymes that catalyze DNA rearrangement events with broad impacts on gene expression, genome evolution, and the spread of drug-resistance in bacteria. Here, we use biochemical and structural approaches to define the molecular determinants by which IstA, a transposase present in the widespread IS21 family of mobile elements, catalyzes efficient DNA transposition. Solution studies show that IstA engages the transposon terminal sequences to form a high-molecular weight complex and promote DNA integration. A 3.4 Å resolution structure of the transposase bound to transposon ends corroborates our biochemical findings and reveals that IstA self-assembles into a highly intertwined tetramer that synapses two supercoiled terminal inverted repeats. The three-dimensional organization of the IstA•DNA cleaved donor complex reveals remarkable similarities with retroviral integrases and classic transposase systems, such as Tn7 and bacteriophage Mu, and provides insights into IS21 transposition.

ABSTRACTMalignant tumors often escape anticancer immune surveillance by suppressing the cytotoxic functions of T lymphocytes. While many of these immune evasion networks include checkpoint proteins, small molecular weight compounds, such as the amino acid L-kynurenine (LKU), could also substantially contribute to the suppression of anti-cancer immunity. However, the biochemical mechanisms underlying the suppressive effects of LKU on T-cells remain unclear. Here, we report for the first time that LKU suppresses T cell function as an aryl hydrocarbon receptor (AhR) ligand. The presence of LKU in T cells is associated with AhR activation, which results in competition between AhR and hypoxia-inducible factor 1 alpha (HIF-1α) for the AhR nuclear translocator, ARNT, leading to T cell exhaustion. The expression of indoleamine 2,3-dioxygenase 1 (IDO1, the enzyme that leads to LKU generation) is induced by the TGF-β-Smad-3 pathway. We also show that IDO-negative cancers utilize an alternative route for LKU production via the endogenous inflammatory mediator, the high mobility group box 1 (HMGB-1)-interferon-gamma (IFN-γ) axis. In addition, other IDO-negative tumors (like T-cell lymphomas) trigger IDO1 activation in eosinophils present in the tumor microenvironment (TME). These mechanisms suppress cytotoxic T cell function, and thus support the tumor immune evasion machinery.

Renal tumors comprise ~7% of all malignant pediatric tumors. Approximately 90% of pediatric kidney tumors comprise Wilms tumors, and the remaining 10% include clear cell sarcoma of the kidney, malignant rhabdoid tumor of the kidney, renal cell carcinoma and other rare renal tumors. Over the last 30 years, the role of cytokines and their receptors has been considerably investigated in both cancer progression and anti-cancer therapy. However, more effective immunotherapies require the cytokine profiling of each tumor type and comprehensive understanding of tumor biology. In this study, we aimed to investigate the activation of signaling pathways in response to cytokines in three pediatric kidney tumor cell lines, in WT-CLS1 and WT-3ab cells (both are Wilms tumors), and in G-401 cells (a rhabdoid kidney tumor, formerly classified as Wilms tumor). We observed that interferon-alpha (IFN-α) and interferon-gamma (IFN-γ) very strongly induced the activation of the STAT1 protein, whereas IL-6 and IFN-α activated STAT3 and IL-4 activated STAT6 in all examined tumor cell lines. STAT protein activation was examined by flow cytometry and Western blot using phospho-specific anti-STAT antibodies which recognize only activated (phosphorylated) STAT proteins. Nuclear translocation of phospho-STAT proteins upon activation with specific cytokines was furthermore confirmed by immunofluorescence. Our results also showed that both IFN-α and IFN-γ caused upregulation of major histocompatibility complex (MHC) class I proteins, however, these cytokines did not have any effect on the expression of MHC class II proteins. We also observed that pediatric kidney tumor cell lines exhibit the functional expression of an additional cytokine signaling pathway, the tumor necrosis factor (TNF)-α-mediated activation of nuclear factor kappa B (NF-κB). In summary, our data show that human pediatric renal tumor cell lines are responsive to stimulation with various human cytokines and could be used as in vitro models for profiling cytokine signaling pathways.

Generating biomass-at-age indices for fisheries stock assessments with acoustic data collected by uncrewed surface vessels (USVs) has been hampered by the need to resolve acoustic backscatter with contemporaneous biological (e.g., age) composition data. To address this limitation, Pacific hake (Merluccius productus; “hake”) acoustic data were gathered from a USV survey (in 2019) and acoustic-trawl survey (ATS; 2019 and eight previous years), and biological data were gathered from fishery-dependent and non-target (i.e., not specifically targeting hake) fishery-independent sources (2019 and eight previous years). To overcome the lack of contemporaneous biological sampling in the USV survey, age class compositions were estimated from a generalized linear mixed spatio-temporal model (STM) fit to the fishery-dependent and non-target fishery-independent data. The validity of the STM age composition estimation procedure was assessed by comparing estimates to age compositions from the ATS in each year. Hake biomass-at-age was estimated from all combinations of acoustics (USV or ATS in 2019, ATS only in other years) and age composition information (STM or ATS in all years). Across the survey area, proportional age class compositions derived from the best STM differed from ATS observations by 0.09 on average in 2019 (median relative error (MRE): 19.45%) and 0.14 across all years (MRE: 79.03%). In data-rich areas (i.e., areas with regular fishery operations), proportional age class compositions from the STM differed from ATS observations by 0.03 on average in 2019 (MRE: 11.46%) and 0.09 across years (MRE: 54.96%). On average, total biomass estimates derived using STM age compositions differed from ATS age composition-based estimates by approximately 7% across the study period (~ 3% in 2019) given the same source of acoustic data. When biomass estimates from different sources of acoustic data (USV or ATS) were compared given the same source of age composition data, differences were nearly ten-fold greater (22% or 27%, depending on if ATS or STM age compositions were used). STMs fit to non-contemporaneous data may provide suitable information for assigning population structure to acoustic backscatter in data-rich areas, but advancements in acoustic data processing (e.g., automated echo classification) may be needed to generate viable USV-based estimates of biomass-at-age.

Abstract A multidisciplinary group of international physicians involved in the medical nutrition therapy (MNT) of adult critically ill patients met to discuss the value, role, and open questions regarding supplemental parenteral nutrition (SPN) along with oral or enteral nutrition (EN), particularly in the intensive care unit (ICU) setting. This manuscript summarizes the discussions and results to highlight the importance of SPN as part of a comprehensive approach to MNT in critically ill adults and for researchers to generate new evidence based on well-powered randomized controlled trials (RCTs). The experts agreed on several key points: SPN has shown clinical benefits, resulting in this strategy being included in American and European guidelines. Nevertheless, its use is heterogeneous across European countries, due to the persistence of uncertainties, such as the optimal timing and the risk of overfeeding in absence of indirect calorimetry (IC), which results in divergent opinions and barriers to SPN implementation. Education is also insufficient. The experts agreed on actions needed to increase evidence quality on SPN use in specific patients at a given time point during acute critical illness or recovery.

Epstein-Barr virus (EBV) is a ubiquitous herpesvirus, which infects over 90% of the adult human population worldwide. After primary infections, EBV is recurrently reactivating in most adult individuals. It is, however, unclear, why these EBV reactivations progress to EBV+ Hodgkin (EBV+HL) or non-Hodgkin lymphomas (EBV+nHL) only in a minority of EBV-infected individuals. The EBV LMP-1 protein encodes for a highly polymorphic peptide, which upregulates the immunomodulatory HLA-E in EBV-infected cells, thereby stimulating the inhibitory NKG2A-, but also the activating NKG2C-receptor on natural killer (NK) cells. Using a genetic-association approach and functional NK cell analyses, we now investigated, whether these HLA-E-restricted immune responses impact the development of EBV+HL and EBV+nHL. Therefore, we recruited a study cohort of 63 EBV+HL and EBV+nHL patients and 192 controls with confirmed EBV reactivations, but without lymphomas. Here, we demonstrate that in EBV+ lymphoma patients exclusively the high-affine LMP-1 GGDPHLPTL peptide variant-encoding EBV-strains reactivate. In EBV+HL and EBV+nHL patients, the high-expressing HLA-E*0103/0103 genetic variant was significantly overrepresented. Combined, the LMP-1 GGDPHLPTL and HLA-E*0103/0103 variants efficiently inhibited NKG2A+ NK cells, thereby facilitating the in vitro spread of EBV-infected tumor cells. In addition, EBV+HL and EBV+nHL patients, showed impaired pro-inflammatory NKG2C+ NK cell responses, which accelerated the in vitro EBV-infected tumor cells spread. In contrast, the blocking of NKG2A by monoclonal antibodies (Monalizumab) resulted in efficient control of EBV-infected tumor cell growth, especially by NKG2A+NKG2C+ NK cells. Thus, the HLA-E/LMP-1/NKG2A pathway and individual NKG2C+ NK cell responses are associated with the progression toward EBV+ lymphomas.

IntroductionUnderstanding how abundance, productivity and distribution of individual species may respond to climate change is a critical first step towards anticipating alterations in marine ecosystem structure and function, as well as developing strategies to adapt to the full range of potential changes.MethodsThis study applies the NOAA (National Oceanic and Atmospheric Administration) Fisheries Climate Vulnerability Assessment method to 64 federally-managed species in the California Current Large Marine Ecosystem to assess their vulnerability to climate change, where vulnerability is a function of a species’ exposure to environmental change and its biological sensitivity to a set of environmental conditions, which includes components of its resiliency and adaptive capacity to respond to these new conditions.ResultsOverall, two-thirds of the species were judged to have Moderate or greater vulnerability to climate change, and only one species was anticipated to have a positive response. Species classified as Highly or Very Highly vulnerable share one or more characteristics including: 1) having complex life histories that utilize a wide range of freshwater and marine habitats; 2) having habitat specialization, particularly for areas that are likely to experience increased hypoxia; 3) having long lifespans and low population growth rates; and/or 4) being of high commercial value combined with impacts from non-climate stressors such as anthropogenic habitat degradation. Species with Low or Moderate vulnerability are either habitat generalists, occupy deep-water habitats or are highly mobile and likely to shift their ranges.DiscussionAs climate-related changes intensify, this work provides key information for both scientists and managers as they address the long-term sustainability of fisheries in the region. This information can inform near-term advice for prioritizing species-level data collection and research on climate impacts, help managers to determine when and where a precautionary approach might be warranted, in harvest or other management decisions, and help identify habitats or life history stages that might be especially effective to protect or restore.

Background Galectin-9 is a member of the family of lectin proteins and crucially regulates human immune responses, particularly because of its ability to suppress the anticancer activities of T lymphocytes and natural killer cells. Recent evidence demonstrated that galectin-9 is highly expressed in a wide range of human malignancies including the most aggressive tumors, such as high-grade glioblastomas and pancreatic ductal adenocarcinomas, as well as common malignancies such as breast, lung and colorectal cancers. However, solid tumor cells at rest are known to secrete either very low amounts of galectin-9 or, in most of the cases, do not secrete it at all. Our aims were to elucidate whether T cells can induce galectin-9 secretion in human cancer cells derived from solid malignant tumors and whether this soluble form displays higher systemic immunosuppressive activity compared with the cell surface-based protein.Methods A wide range of human cancer cell lines derived from solid tumours, keratinocytes and primary embryonic cells were employed, together with helper and cytotoxic T cell lines and human as well as mouse primary T cells. Western blot analysis, ELISA, quantitative reverse transcriptase-PCR, on-cell Western and other measurement techniques were used to conduct the study. Results were validated using in vivo mouse model.Results We discovered that T lymphocytes induce galectin-9 secretion in various types of human cancer cells derived from solid malignant tumors. This was demonstrated to occur via two differential mechanisms: first by translocation of galectin-9 onto the cell surface followed by its proteolytic shedding and second due to autophagy followed by lysosomal secretion. For both mechanisms a protein carrier/trafficker was required, since galectin-9 lacks a secretion sequence. Secreted galectin-9 pre-opsonised T cells and, following interaction with other immune checkpoint proteins, their activity was completely attenuated. As an example, we studied the cooperation of galectin-9 and V-domain Ig-containing suppressor of T cell activation (VISTA) proteins in human cancer cells.Conclusion Our results underline a crucial role of galectin-9 in anticancer immune evasion. As such, galectin-9 and regulatory pathways controlling its production should be considered as key targets for immunotherapy in a large number of cancers.

Introduction: Coagulation disorders occur during the first 48 h after major burn injury, affecting mainly PT/INR and aPTT. Some authors consider them burn specific. The aim was to assess the magnitude of coagulation disorders during the first 10 days after severe burn injury and its eventual association with fluid intakes. Methods: Retrospective study of prospectively collected data including patients with burns ≥ 20%BSA (body surface area) who stayed ≥ 24 h at the burn-intensive care unit (ICU). Predetermined data for the analysis of fluid and blood products administration and laboratory values were extracted from the computerized information system. Descriptive statistics, data as median. Results: 167 patients included. The volumes of fluid resuscitation decreased over the first 10 days (median 6815 ml on day 1 decreasing to 4190 ml by day 10, p

The indications and contraindications of parenteral nutrition (PN) are discussed in view of recent clinical findings. For decades, PN has been restricted to patients unable to tolerate enteral nutrition (EN) intake owing to the perceived risk of severe side-effects. The evolution of the PN substrate composition and delivery of nutrition via all-in-one bags has dramatically improved the application prospects of PN. Recent studies show similar complication rates of nutrition therapy administered through enteral and intravenous routes. Therefore, indications of PN have, based on evidence, extended beyond complete gastrointestinal (GI) failure to include conditions such as insufficient EN generating persistent negative energy balance and insufficient protein intakes, malabsorption, or specific needs that are impossible to cover with EN feeds.

Today, suburban areas are home to an ever-increasing majority of the global population. Models indicate that the next generation of US metropolitan growth will rapidly continue outside of urban cores, where car-based development patterns have served as the dominant paradigm for more than a century. With the emergence of autonomous mobility technologies and services, the suburbs of the future offer key opportunities to tackle pressing environmental challenges, such as significant GHG emissions from private vehicle trips, underutilized and fragmented landscape spaces, and a high proportion of impervious surfaces. To leverage this opportunity, our research team employed a novel scenario-based parametric modeling framework to generate and optimize suburban land use patterns and block configurations that leverage autonomous mobility to optimize environmental performance and accessibility metrics. The framework performed through our project, NOGAS (Next Optimized Generation of Autonomous Suburbs), consists of five key parametric modules and a heuristic design process covering various planning and design decision-making stages including scenario generation, analysis, optimization, and visualization. It is the first of its kind tailored for suburban settings with emerging mobility systems, which, more importantly, prioritizes landscape performance and accessibility over the traditional automobile-centric approach in suburban development. One of the most significant findings from this research is that substantial enhancements to a neighborhood’s environmental performance and overall accessibility can be achieved by modifying existing suburban land use patterns and individual block configurations, without the necessity of increasing density. The results of the framework further suggest that a strategic atomized land use scheme, combined with an innovative clustered block typology, is favored for the anticipated widespread adoption of autonomous mobility systems and improved environmental performance. The innovative methods and findings introduced in this research illuminate an alternative approach to sustainable suburban development, offering valuable insights for city planners and developers to shape future suburban master plans, zoning regulations, and design guidelines.

Abstract Background Stroke survivors show deteriorated physical functioning and physical activity levels. Physical activity levels of stroke survivors are generally low. It is increasingly recognized that physical activity is a multidimensional construct that cannot be captured in a single outcome. In-depth insight into multidimensional physical activity patterns may guide the development and timing of targeted rehabilitation interventions. This longitudinal cohort study explored how multidimensional physical activity outcomes develop during recovery in the subacute phase after stroke and if changes in physical activity were correlated to recovery of lower limb motor function. Methods Patients were recruited during inpatient rehabilitation. At 3, 12, and 26 weeks post-onset, motor function was measured by the Fugl-Meyer Lower Extremity Assessment (FMA-LE). Physical activity was measured with the Activ8 accelerometer in multiple outcomes: counts per minute during walking (CPMwalking; a measure of Intensity), number of active bouts (Frequency), mean length of active bouts (Distribution) and % of waking time in upright positions (Duration). Generalized estimating equations (GEE) were used to study changes in physical activity over time and the relation with the change in lower limb motor recovery. Results Thirty-nine patients (age 56 ± 9, 77% male, 89% ischemic stroke) were included. GEE models showed a significant main effect of time for PA Intensity (+ 13%, p = 0.007) and Duration (+ 64%, p = 0.012) between 3 and 12 weeks. Motor function did not show a significant effect in all PA models across the 3 timepoints (p > 0.020). A significant interaction effect of time × motor function was observed (p

Abstract The preferential use of the oral/enteral route in critically ill patients over gut rest is uniformly recommended and applied. This article provides practical guidance on enteral nutrition in compliance with recent American and European guidelines. Low-dose enteral nutrition can be safely started within 48 h after admission, even during treatment with small or moderate doses of vasopressor agents. A percutaneous access should be used when enteral nutrition is anticipated for ≥ 4 weeks. Energy delivery should not be calculated to match energy expenditure before day 4–7, and the use of energy-dense formulas can be restricted to cases of inability to tolerate full-volume isocaloric enteral nutrition or to patients who require fluid restriction. Low-dose protein (max 0.8 g/kg/day) can be provided during the early phase of critical illness, while a protein target of > 1.2 g/kg/day could be considered during the rehabilitation phase. The occurrence of refeeding syndrome should be assessed by daily measurement of plasma phosphate, and a phosphate drop of 30% should be managed by reduction of enteral feeding rate and high-dose thiamine. Vomiting and increased gastric residual volume may indicate gastric intolerance, while sudden abdominal pain, distension, gastrointestinal paralysis, or rising abdominal pressure may indicate lower gastrointestinal intolerance.

Type II topoisomerases modulate chromosome supercoiling, condensation, and catenation by moving one double-stranded DNA segment through a transient break in a second duplex. How DNA strands are chosen and selectively passed to yield appropriate topological outcomes – for example, decatenation vs. catenation – is poorly understood. Here, we show that at physiological enzyme concentrations, eukaryotic type IIA topoisomerases (topo IIs) readily coalesce into condensed bodies. DNA stimulates condensation and fluidizes these assemblies to impart liquid-like behavior. Condensation induces both budding yeast and human topo IIs to switch from DNA unlinking to active DNA catenation, and depends on an unstructured C-terminal region, the loss of which leads to high levels of knotting and reduced catenation. Our findings establish that local protein concentration and phase separation can regulate how topo II creates or dissolves DNA links, behaviors that can account for the varied roles of the enzyme in supporting transcription, replication, and chromosome compaction.

OBJECTIVES:. Bivalirudin, an IV direct thrombin inhibitor, and unfractionated heparin (UFH) are frequently used anticoagulants in the pediatric critical care setting. An accurate, specific, point-of-care test to quantify and detect anticoagulation resistance is not currently available. This study evaluates the ability of a rapid (< 10 min), micro-volume (< 50 uL) coagulation test to detect and quantify the anticoagulation effect of bivalirudin and UFH using a functional, clot time endpoint in pediatric critical care patients. DESIGN:. Single-site retrospective laboratory sample analysis and chart review. SETTING:. A 105-bed pediatric and cardiac ICUs delivering extracorporeal membrane oxygenation. SUBJECTS:. Forty-one citrated, frozen, biobanked plasma specimens comprising 21 with bivalirudin and 20 with UFH from 15 anticoagulated pediatric patients were analyzed. Thirteen patients were on extracorporeal membrane oxygenation, one had a submassive pulmonary embolism, and one was on a left ventricular assist device. INTERVENTIONS:. None. MEASUREMENT AND MAIN RESULTS:. A Clotting Time Score (CTS) was derived on each sample. The CTS detected patients that had developed a pathologic clotting event with 100% sensitivity and 82% specificity compared with prothrombin time with 25% sensitivity/76% specificity and activated partial thromboplastin time with 0% sensitivity/0% specificity. Additionally, the CTS detected subtherapeutic anticoagulation in response to UFH in patients that were clinically determined to be UFH resistant requiring alternative anticoagulation with bivalirudin. CONCLUSIONS:. The CTS appears to be a clinically valuable indicator of coagulation status in patients treated with either UFH or bivalirudin. Results outside of the therapeutic range due to inadequate dosing or anticoagulation resistance appeared to be associated with clot formation. CTS testing may reduce the risk of anticoagulation-related complications via the rapid identification of patients at high risk for pathologic thrombotic events.

Transcranial current stimulation (tCS) techniques have been shown to induce cortical plasticity. As an important relay in the motor system, the cerebellum is an interesting target for plasticity induction using tCS, aiming to modulate its excitability and connectivity. However, until now it remains unclear, which is the most effective tCS method for inducing plasticity in the cerebellum. Thus, in this study, the effects of anodal transcranial direct current stimulation (tDCS), 50 Hz transcranial alternating current stimulation (50 Hz tACS), and high frequency transcranial random noise stimulation (tRNS) were compared with sham stimulation in 20 healthy subjects in a within-subject design. tCS was applied targeting the cerebellar lobe VIIIA using neuronavigation. We measured corticospinal excitability, short-interval intracortical inhibition (SICI), short-latency afferent inhibition (SAI), and cerebellar brain inhibition (CBI) and performed a sensor-based movement analysis at baseline and three times after the intervention (post1 = 15 min; post2 = 55 min; post3 = 95 min). Corticospinal excitability increased following cerebellar tACS and tRNS compared to sham stimulation. This effect was most pronounced directly after stimulation but lasted for at least 55 min after tACS. Cortico-cortical and cerebello-cortical conditioning protocols, as well as sensor-based movement analyses, did not change. Our findings suggest that cerebellar 50 Hz tACS is the most effective protocol to change corticospinal excitability.

Electroencephalography-neurofeedback (EEG-NF) has become a valuable tool in the field of psychology, e.g., to improve cognitive function. Nevertheless, a large percentage of NF users seem to be unable to control their own brain activation. Therefore, the aim of this study was to examine whether a different kind of visual feedback could positively influence NF performance after one training session. Virtual reality (VR) seems to have beneficial training effects and has already been reported to increase motivational training aspects. In the present study, we tested 61 young healthy adults (mean age: 23.48 years; 28 female) to investigate, whether 3D VR-based NF training has a more beneficial effect on the sensorimotor rhythm (SMR, 12–15 Hz) power increase than a mere 2D conventional NF paradigm. In the 3D group, participants had to roll a ball along a predefined path in an immersive virtual environment, whereas the 2D group had to increase the height of a bar. Both paradigms were presented using VR goggles. Participants completed one baseline and six feedback runs with 3 min each, in which they should try to increase SMR power over Cz. Half of the participants received real feedback whereas the other half received sham feedback. Participants receiving 3D VR-based feedback showed a linear increase in SMR power over the feedback runs within one training session. This was the case for the real as well as for the sham 3D feedback group and might be related to more general VR-related effects. The 2D group receiving the conventional bar feedback showed no changes in SMR power over the feedback runs. The present study underlines that the visual feedback modality has differential effects on the NF training performance and that 3D VR-based feedback has advantages over conventional 2D feedback.

E3 ligases constitute a large and diverse family of proteins that play a central role in regulating protein homeostasis by recruiting substrate proteins via recruitment domains to the proteasomal degradation machinery. Small molecules can either inhibit, modulate or hijack E3 function. The latter class of small molecules led to the development of selective protein degraders, such as PROTACs (PROteolysis TArgeting Chimeras), that recruit protein targets to the ubiquitin system leading to a new class of pharmacologically active drugs and to new therapeutic options. Recent efforts have focused on the E3 family of Baculovirus IAP Repeat (BIR) domains that comprise a structurally conserved but diverse 70 amino acid long protein interaction domain. In the human proteome, 16 BIR domains have been identified, among them promising drug targets such as the Inhibitors of Apoptosis (IAP) family, that typically contain three BIR domains (BIR1, BIR2, and BIR3). To date, this target area lacks assay tools that would allow comprehensive evaluation of inhibitor selectivity. As a consequence, the selectivity of current BIR domain targeting inhibitors is unknown. To this end, we developed assays that allow determination of inhibitor selectivity in vitro as well as in cellulo. Using this toolbox, we have characterized available BIR domain inhibitors. The characterized chemical starting points and selectivity data will be the basis for the generation of new chemical probes for IAP proteins with well-characterized mode of action and provide the basis for future drug discovery efforts and the development of PROTACs and molecular glues.

The NavDx® blood test analyzes tumor tissue modified viral (TTMV)-HPV DNA to provide a reliable means of detecting and monitoring HPV-driven cancers. The test has been clinically validated in a large number of independent studies and has been integrated into clinical practice by over 1000 healthcare providers at over 400 medical sites in the US. This Clinical Laboratory Improvement Amendments (CLIA), high complexity laboratory developed test, has also been accredited by the College of American Pathologists (CAP) and the New York State Department of Health. Here, we report a detailed analytical validation of the NavDx assay, including sample stability, specificity as measured by limits of blank (LOBs), and sensitivity illustrated via limits of detection and quantitation (LODs and LOQs). LOBs were 0–0.32 copies/μL, LODs were 0–1.10 copies/μL, and LOQs were 2 = 1) across a broad range of analyte concentrations. These results demonstrate that NavDx accurately and reproducibly detects circulating TTMV-HPV DNA, which has been shown to aid in the diagnosis and surveillance of HPV-driven cancers.

The meat industry is typically using a mixture of fresh and frozen meat batters for minced meat production. Our goal was to find the exact threshold for fresh to frozen meat ratio capable of controlling the meat temperature during processing, but without having an adverse effect on the sensory quality of minced pork. To achieve this, the percentage of frozen meat used for the minced pork production was increased from 0% (control) to 50% (maximum) in 10% increments. To keep the minced meat temperature in control and make the processing resistant to fat smearing, the addition of 30% of frozen meat to the meat batter is sufficient. The soluble protein content, instrumental cutting force, and the sensory perceived firmness, juiciness, and inner cohesion were not affected by the addition of frozen meat. However, it has contributed to a significant increase of the drip loss and the amount of non-intact cells (ANIC). With the addition of frozen meat into the minced pork, the compliance to ANIC regulation by the German regulatory authorities is technologically (practically) almost impossible.

Through their ability to regulate gene expression in most organs, glucocorticoid (GC) hormones influence numerous physiological processes and are therefore key regulators of organismal homeostasis. In bone, GC hormones inhibit expression of the hormone Osteocalcin for poorly understood reasons. Here, we show that in a classical endocrine feedback loop, osteocalcin in return enhanced the biosynthesis of GC as well as mineralocorticoid hormones (adrenal steroidogenesis) in rodents and primates. Conversely, inactivation of osteocalcin signaling in adrenal glands significantly impaired adrenal growth and steroidogenesis in mice. Embryo-made osteocalcin was necessary for normal Sf1 expression in fetal adrenal cells and adrenal cell steroidogenic differentiation and therefore determined the number of steroidogenic cells present in the adrenal glands of adult animals. Embryonic, not postnatal, osteocalcin also governed adrenal growth, adrenal steroidogenesis, blood pressure, electrolyte equilibrium, and the rise in circulating corticosterone levels during the acute stress response in adult offspring. This osteocalcin-dependent regulation of adrenal development and steroidogenesis occurred even in the absence of a functional hypothalamus/pituitary/adrenal axis and explains why osteocalcin administration during pregnancy promoted adrenal growth and steroidogenesis and improved the survival of adrenocorticotropic hormone signaling–deficient animals. This study reveals that a bone-derived embryonic hormone influences lifelong adrenal functions and organismal homeostasis in the mouse.

Immune checkpoint proteins play crucial roles in human embryonic development but are also used by cancer cells to escape immune surveillance. These proteins and biochemical pathways associated with them form a complex machinery capable of blocking the ability of cytotoxic immune lymphoid cells to attack cancer cells and, ultimately, to fully suppress anti-tumor immunity. One of the more recently discovered immune checkpoint proteins is V-domain Ig-containing suppressor of T cell activation (VISTA), which plays a crucial role in anti-cancer immune evasion pathways. The biochemical mechanisms underlying regulation of VISTA expression remain unknown. Here, we report for the first time that VISTA expression is controlled by the transforming growth factor beta type 1 (TGF-β)-Smad3 signaling pathway. However, in T lymphocytes, we found that VISTA expression was differentially regulated by TGF-β depending on their immune profile. Taken together, our results demonstrate the differential biochemical control of VISTA expression in human T cells and various types of rapidly proliferating cells, including cancer cells, fetal cells and keratinocytes.

Protein prenylation by farnesyltransferase (FTase) is often described as the targeting of a cysteine-containing motif (CaaX) that is enriched for aliphatic amino acids at the a1 and a2 positions, while quite flexible at the X position. Prenylation prediction methods often rely on these features despite emerging evidence that FTase has broader target specificity than previously considered. Using a machine learning approach and training sets based on canonical (prenylated, proteolyzed, and carboxymethylated) and recently identified shunted motifs (prenylation only), this study aims to improve prenylation predictions with the goal of determining the full scope of prenylation potential among the 8000 possible Cxxx sequence combinations. Further, this study aims to subdivide the prenylated sequences as either shunted (i.e., uncleaved) or cleaved (i.e., canonical). Predictions were determined for Saccharomyces cerevisiae FTase and compared to results derived using currently available prenylation prediction methods. In silico predictions were further evaluated using in vivo methods coupled to two yeast reporters, the yeast mating pheromone a-factor and Hsp40 Ydj1p, that represent proteins with canonical and shunted CaaX motifs, respectively. Our machine learning-based approach expands the repertoire of predicted FTase targets and provides a framework for functional classification.

Summary: This protocol is used to profile the engagement of kinase inhibitors across nearly 200 kinases in a live-cell context. This protocol utilizes one single kinase tracer (NanoBRET(TM) Tracer K10) that operates quantitatively at four different concentrations. Minimizing the number of tracers offers a significant workflow improvement over the previous protocol that utilized a combination of 6 tracers. Each NanoBRET(TM) kinase assay is built using commercially available plasmids and has been optimized for NanoLuc tagging orientation, diluent DNA, and tracer concentration.For complete details on the use and execution of this protocol, please refer to Vasta et al. (2018).

Liquid–liquid phase separation (LLPS) of intrinsically disordered regions (IDRs) in proteins can drive the formation of membraneless compartments in cells. Phase-separated structures enrich for specific partner proteins and exclude others. Previously, we showed that the IDRs of metazoan DNA replication initiators drive DNA-dependent phase separation in vitro and chromosome binding in vivo, and that initiator condensates selectively recruit replication-specific partner proteins (Parker et al., 2019). How initiator IDRs facilitate LLPS and maintain compositional specificity is unknown. Here, using Drosophila melanogaster (Dm) Cdt1 as a model initiation factor, we show that phase separation results from a synergy between electrostatic DNA-bridging interactions and hydrophobic inter-IDR contacts. Both sets of interactions depend on sequence composition (but not sequence order), are resistant to 1,6-hexanediol, and do not depend on aromaticity. These findings demonstrate that distinct sets of interactions drive condensate formation and specificity across different phase-separating systems and advance efforts to predict IDR LLPS propensity and partner selection a priori.

Food products often consist of several phases. Comminuted meat products, for example, are multiphase systems consisting of structured meat particles and unstructured batter-like substance. To develop and understand the processing of these products, it is important to understand the sensory and mechanical perception principles. To this end, two-phase food prototypes consisting of mixtures of ground beef and beef batter were prepared and subjected to sensory, texture, and oral processing analysis. The oral processing analysis focused on the biomechanical data of the chewing process, namely the kinematics of jaw movement and electromyographic activity. The ground meat served as the anisotropic phase and the meat dough as the isotropic phase. A significant increase in muscle activity, duration per bite, and occlusion time with increasing proportion of fibrous particles was demonstrated (p < 0.05). In contrast, a higher proportion of isotropic substance resulted in significantly higher amplitudes of jaw movement and faster jaw kinetics (p < 0.05). In mixed regimes, the system responded mainly according to the dominant phase, with sensory or mechanical response changing at a critical point. In combination with texture and sensory data, a holistic characterization of the food models could be performed.

Larger processing equipment to produce minced meat could affect its structure due to intensive processing and a high energy intake in the meat mass. To assess if this would result in alterations in the minced meat quality, finely chopped meat (FCM) was added in different concentrations (15, 30, 45, 60, 75, 90, and 100%) to minced meat and quality parameters were analyzed. FCM was used to simulate different intensity of an unintended destruction of meat cells due to various processes. The amount of non-intact cells (ANIC) was determined histologically and furthermore, soluble protein content, water holding capacity, mechanical and sensory texture, and scanning electron and confocal laser scanning microscopy was applied to analyze the meat structure and quality. ANIC indicated that even adding 15% FCM was statistically (p < 0.05) distinguishable from 100% minced meat and 30% FCM had already 50 Vol.-% ANIC. In contrast, the addition of 15% or 30% FCM did not result in significant differences in drip loss of raw and cooked meat as well as mechanical and sensory texture analysis. This study showed that intensive processing might be detectable via ANIC, but that the minced meat quality was not affected.

In medical tasks such as human motion analysis, computer-aided auxiliary systems have become the preferred choice for human experts for their high efficiency. However, conventional approaches are typically based on user-defined features such as movement onset times, peak velocities, motion vectors, or frequency domain analyses. Such approaches entail careful data post-processing or specific domain knowledge to achieve a meaningful feature extraction. Besides, they are prone to noise and the manual-defined features could hardly be re-used for other analyses. In this paper, we proposed probabilistic movement primitives (ProMPs), a widely-used approach in robot skill learning, to model human motions. The benefit of ProMPs is that the features are directly learned from the data and ProMPs can capture important features describing the trajectory shape, which can easily be extended to other tasks. Distinct from previous research, where classification tasks are mostly investigated, we applied ProMPs together with a variant of Kullback-Leibler (KL) divergence to quantify the effect of different transcranial current stimulation methods on human motions. We presented an initial result with 10 participants. The results validate ProMPs as a robust and effective feature extractor for human motions.