An integrated metaproteomics workflow for studying host-microbe dynamics in bronchoalveolar lavage samples applied to cystic fibrosis disease.

Airway microbiota are known to contribute to lung diseases, such as cystic fibrosis (CF), but their contributions to pathogenesis are still unclear. To improve our understanding of host-microbe interactions, we have developed an integrated analytical and bioinformatic mass spectrometry (MS)-based metaproteomics workflow to analyze clinical bronchoalveolar lavage (BAL) samples from people with airway disease. Proteins from BAL cellular pellets were processed and pooled together in groups categorized by disease status (CF vs. non-CF) and bacterial diversity, based on previously performed small subunit rRNA sequencing data. Proteins from each pooled sample group were digested and subjected to liquid chromatography tandem mass spectrometry (MS/MS). MS/MS spectra were matched to human and bacterial peptide sequences leveraging a bioinformatic workflow using a metagenomics-guided protein sequence database and rigorous evaluation. Label-free quantification revealed differentially abundant human peptides from proteins with known roles in CF, like neutrophil elastase and collagenase, and proteins with lesser-known roles in CF, including apolipoproteins. Differentially abundant bacterial peptides were identified from known CF pathogens (e.g., Pseudomonas), as well as other taxa with potentially novel roles in CF. We used this host-microbe peptide panel for targeted parallel-reaction monitoring validation, demonstrating for the first time an MS-based assay effective for quantifying host-microbe protein dynamics within BAL cells from individual CF patients. Our integrated bioinformatic and analytical workflow combining discovery, verification, and validation should prove useful for diverse studies to characterize microbial contributors in airway diseases. Furthermore, we describe a promising preliminary panel of differentially abundant microbe and host peptide sequences for further study as potential markers of host-microbe relationships in CF disease pathogenesis.IMPORTANCEIdentifying microbial pathogenic contributors and dysregulated human responses in airway disease, such as CF, is critical to understanding disease progression and developing more effective treatments. To this end, characterizing the proteins expressed from bacterial microbes and human host cells during disease progression can provide valuable new insights. We describe here a new method to confidently detect and monitor abundance changes of both microbe and host proteins from challenging BAL samples commonly collected from CF patients. Our method uses both state-of-the art mass spectrometry-based instrumentation to detect proteins present in these samples and customized bioinformatic software tools to analyze the data and characterize detected proteins and their association with CF. We demonstrate the use of this method to characterize microbe and host proteins from individual BAL samples, paving the way for a new approach to understand molecular contributors to CF and other diseases of the airway.

The contribution of NaV1.6 to the efficacy of voltage-gated sodium channel inhibitors in wild type and NaV1.6 gain-of-function (GOF) mouse seizure control.

BACKGROUND AND PURPOSE: Inhibitors of voltage-gated sodium channels (NaVs) are important anti-epileptic drugs, but the contribution of specific channel isoforms is unknown since available inhibitors are non-selective. We aimed to create novel, isoform selective inhibitors of Nav channels as a means of informing the development of improved antiseizure drugs. EXPERIMENTAL APPROACH: We created a series of compounds with diverse selectivity profiles enabling block of NaV1.6 alone or together with NaV1.2. These novel NaV inhibitors were evaluated for their ability to inhibit electrically evoked seizures in mice with a heterozygous gain-of-function mutation (N1768D/+) in Scn8a (encoding NaV1.6) and in wild-type mice. KEY RESULTS: Pharmacologic inhibition of NaV1.6 in Scn8aN1768D/+ mice prevented seizures evoked by a 6-Hz shock. Inhibitors were also effective in a direct current maximal electroshock seizure assay in wild-type mice. NaV1.6 inhibition correlated with efficacy in both models, even without inhibition of other CNS NaV isoforms. CONCLUSIONS AND IMPLICATIONS: Our data suggest NaV1.6 inhibition is a driver of efficacy for NaV inhibitor anti-seizure medicines. Sparing the NaV1.1 channels of inhibitory interneurons did not compromise efficacy. Selective NaV1.6 inhibitors may provide targeted therapies for human Scn8a developmental and epileptic encephalopathies and improved treatments for idiopathic epilepsies.

Correction: Adaptation and validation of the Johnson-Lecci scale to assess anti-white bias among black UK minority group members.

[This corrects the article DOI: 10.1371/journal.pone.0277077.].

Longitudinal molecular analysis of clinical and fecal Escherichia coli isolates at a Veterans Affairs Medical Center in Minnesota, USA, 2012-2019.

Introduction: Extraintestinal Escherichia coli infections represent a growing public health threat, However, current studies often overlook important factors such as temporal patterns of infection, phylogenetic and clonal background, or the host gut E. coli population, despite their likely significance. Methods: In this study, we analyzed >7000 clinical E. coli isolates from patients at the Minneapolis Veterans Affairs Health Care System (2012-2019), and concurrent fecal E. coli from uninfected veterans. We assessed phylogenetic group distribution, membership in selected sequence types (STs), and subsets thereof-including the pandemic, resistance-associated ST131-H30R, and ST1193 lineages-and strain type, as defined by pulsed-field gel electrophoresis. We then analyzed these features alongside the temporal patterns of infection in individual hosts. Results: The H30R lineage emerged as the leading lineage, both overall and among fluoroquinolone-resistant isolates, with ST1193 following among fluoroquinolone-resistant isolates. Recurrences were common, occurring in 31% of subjects and 41% of episodes, and often multiple and delayed/prolonged (up to 23 episodes per subject; up to 2655d post-index). Remarkably, these recurrences typically involved the subject's index strain (63% of recurrences), even when affecting extra-urinary sites. ST131, H30R, ST1193, and fluoroquinolone-resistant strains generally caused significantly more recurrences than did other strains, despite similar recurrence intervals. ST131 strain types shifted significantly over the study period. Infection-causing strains were commonly detectable in host feces at times other than during an infection episode; the likelihood of detection varied with surveillance intensity and proximity to the infection. H30R and ST1193 were prominent causes of fecal-clinical clonal overlap. Discussion: These findings provide novel insights into the temporal and clonal characteristics of E. coli infections in veterans and support efforts to develop anti-colonization interventions.

An In-Vitro Comparison of Mixed Reality Navigation to Traditional Freehand and Patient Specific Instrumentation Techniques for Glenoid Guide Pin Insertion during Shoulder Arthroplasty.

BACKGROUND: Accurate insertion of the glenoid guide pin in shoulder arthroplasty (RSA) is important for obtaining optimized glenoid component position and orientation. The objective of this study was to evaluate and compare the accuracy of three glenoid guide pin insertion techniques: 1) traditional software planning using freehand guide pin insertion (freehand), 2) guide pin insertion utilizing patient-specific instrumentation (PSI), and 3) using a mixed reality navigation (MR-NAV) system. METHODS: Twenty (20) computer tomography (CT) scans were obtained from patients exhibiting glenoid erosion patterns according to the Walch and Favard classifications. Cases were planned using validated three-dimensional (3D) preoperative planning software. The CT data was then used to 3D print triplicate plastic models of each glenoid to evaluate the three guide pin insertion techniques. The first technique employed traditional software planning with freehand guide pin insertion. The second method used preoperatively planned PSI guides, while the third utilized a MR-NAV system, which provided real-time holographic guidance during guide pin insertion. Once all guide pins had been inserted into the models, an independent optical tracking system and custom digitization device was used to quantify the position and orientation of each guide pin relative to the glenoid. The outcomes for this study included the absolute mean error in guide pin inclination, version, and entry point relative to the preoperative plan. The absolute Total Global Error was also assessed, which was defined as the sum of the absolute guide pin orientation and position error relative to the preoperative plan. RESULTS: No statistically significant differences between MR-NAV and PSI were found for the inclination error (2±1° versus 2±1°; P=0.056), version error (1±1° versus 1±1°; P=1.000), and Total Global Error (5±1 [mm+deg] versus 5±1 [mm+deg], P=1.000), respectively. The freehand technique produced significantly greater error than MR-NAV and PSI for inclination (5±3°, P≤0.017), version (4±3°, P≤0.032) and Total Global Error (8±3 [mm+deg], P<0.001). No statistically significant differences in the entry point error were observed between all guide pin insertion methods (P≥0.058). DISCUSSION: These results demonstrate that the precision and accuracy of MR-NAV is comparable to PSI and superior to a freehand technique for glenoid guide pin insertion in-vitro. Further study is needed to compare the accuracy of these techniques intra-operatively, in addition to assessing a potential learning curve between surgeons of varying experience with the MR-NAV system.

The use of platelet rich plasma in the treatment of degenerative joint disease in cats: an exploratory case series.

Objective: To evaluate the effectiveness of intra-articular autologous Platelet Rich Plasma (PRP) in managing Degenerative Joint Disease (DJD) in cats. Design: Prospective pilot clinical trial. Methods: Six domestic cats with clinically and radiographically diagnosed DJD received intra-articular injections of autologous PRP. Clinical assessments pre and post intra-articular injections were conducted using the Feline Musculoskeletal Pain Index (FMPI, owner assessed) and Visual Analog Scale (VAS, clinician assessed) at baseline, Day 14, Day 28, and Day 42-45. Results: Significant improvements were noted in both FMPI and VAS scores at the end of the study period, indicating enhanced joint function and reduced pain. Conclusion and clinical relevance: The study suggests the potential of PRP therapy as a safe and effective treatment for feline DJD, warranting further research with larger cohorts and longer follow-up to establish comprehensive treatment guidelines.

A novel clinical metaproteomics workflow enables bioinformatic analysis of host-microbe dynamics in disease.

Clinical metaproteomics has the potential to offer insights into the host-microbiome interactions underlying diseases. However, the field faces challenges in characterizing microbial proteins found in clinical samples, usually present at low abundance relative to the host proteins. As a solution, we have developed an integrated workflow coupling mass spectrometry-based analysis with customized bioinformatic identification, quantification, and prioritization of microbial proteins, enabling targeted assay development to investigate host-microbe dynamics in disease. The bioinformatics tools are implemented in the Galaxy ecosystem, offering the development and dissemination of complex bioinformatic workflows. The modular workflow integrates MetaNovo (to generate a reduced protein database), SearchGUI/PeptideShaker and MaxQuant [to generate peptide-spectral matches (PSMs) and quantification], PepQuery2 (to verify the quality of PSMs), Unipept (for taxonomic and functional annotation), and MSstatsTMT (for statistical analysis). We have utilized this workflow in diverse clinical samples, from the characterization of nasopharyngeal swab samples to bronchoalveolar lavage fluid. Here, we demonstrate its effectiveness via analysis of residual fluid from cervical swabs. The complete workflow, including training data and documentation, is available via the Galaxy Training Network, empowering non-expert researchers to utilize these powerful tools in their clinical studies. IMPORTANCE: Clinical metaproteomics has immense potential to offer functional insights into the microbiome and its contributions to human disease. However, there are numerous challenges in the metaproteomic analysis of clinical samples, including handling of very large protein sequence databases for sensitive and accurate peptide and protein identification from mass spectrometry data, as well as taxonomic and functional annotation of quantified peptides and proteins to enable interpretation of results. To address these challenges, we have developed a novel clinical metaproteomics workflow that provides customized bioinformatic identification, verification, quantification, and taxonomic and functional annotation. This bioinformatic workflow is implemented in the Galaxy ecosystem and has been used to characterize diverse clinical sample types, such as nasopharyngeal swabs and bronchoalveolar lavage fluid. Here, we demonstrate its effectiveness and availability for use by the research community via analysis of residual fluid from cervical swabs.

A randomized open-label, observational study of the novel ketone ester, bis octanoyl (R)-1,3-butanediol, and its acute effect on ß-hydroxybutyrate and glucose concentrations in healthy older adults.

Bis-octanoyl (R)-1,3-butanediol (BO-BD) is a novel ketone ester (KE) ingredient which increases blood beta-hydroxybutyrate (BHB) concentrations rapidly after ingestion. KE is hypothesized to have beneficial metabolic effects on health and performance, especially in older adults. Whilst many studies have investigated the ketogenic effect of KE in young adults, they have not been studied in an exclusively older adult population, for whom age-related differences in body composition and metabolism may alter the effects. This randomized, observational, open-label study in healthy older adults (n = 30, 50% male, age = 76.5 years, BMI = 25.2 kg/m2) aimed to elucidate acute tolerance, blood BHB and blood glucose concentrations for 4 hours following consumption of either 12.5 or 25 g of BO-BD formulated firstly as a ready-to-drink beverage (n = 30), then as a re-constituted powder (n = 21), taken with a standard meal. Both serving sizes and formulations of BO-BD were well tolerated, and increased blood BHB, inducing nutritional ketosis (≥ 0.5mM) that lasted until the end of the study. Ketosis was dose responsive; peak BHB concentration (Cmax) and incremental area under the curve (iAUC) were significantly greater with 25 g compared to 12.5 g of BO-BD in both formulations. There were no significant differences in Cmax or iAUC between formulations. Blood glucose increased in all conditions following the meal; there were no consistent significant differences in glucose response between conditions. These results demonstrate that both powder and beverage formulations of the novel KE, BO-BD, induce ketosis in healthy older adults, facilitating future research on functional effects of this ingredient in aging.

Daily consumption of ketone ester, bis-octanoyl (R)-1,3-butanediol, is safe and tolerable in healthy older adults, a randomized, parallel arm, double-blind, placebo-controlled, pilot study.

Objectives: Ketone bodies are endogenous metabolites produced during fasting or a ketogenic diet that have pleiotropic effects on aging pathways. Ketone esters (KEs) are compounds that induce ketosis without dietary changes, but KEs have not been studied in an older adult population. The primary objective of this trial was to determine tolerability and safety of KE ingestion in older adults. Design: Randomized, placebo-controlled, double-blinded, parallel-arm trial, with a 12-week intervention period ( NCT05585762 ). Setting: General community, Northern California, USA. Participants: Community-dwelling older adults, independent in activities of daily living, with no unstable acute medical conditions (n=30) were randomized and n=23 (M= 14, F=9) completed the protocol. Intervention: Participants were randomly allocated to consume either KE (bis-octanoyl (R)-1,3-butanediol) or a taste, appearance, and calorie-matched placebo (PLA) containing canola oil. Measurements: Tolerability was assessed using a composite score from a daily log for 2-weeks, and then via a bi-weekly phone interview. Safety was assessed by vital signs and lab tests at screening and weeks 0, 4 and 12, along with tabulation of adverse events. Results: There was no difference in the prespecified primary outcome of proportion of participants reporting moderate or severe nausea, headache, or dizziness on more than one day in a two-week reporting period (KE n =2 (14.3% [90% CI = 2.6 - 38.5]); PLA n=1 (7.1% [90% CI = 0.4 - 29.7]). Dropouts numbered four in the PLA group and two in the KE group. A greater number of symptoms were reported in both groups during the first two weeks; symptoms were reported less frequently between 2 - 12 weeks. There were no clinically relevant changes in safety labs or vital signs in either group. Conclusions: This KE was safe and well-tolerated in healthy older adults. These results provide a foundation for use of KEs in aging research. Highlights: Ketones esters induce ketosis without dietary changes and may target aging biologyStudies of ketone esters were limited in duration and focused on younger adultsWe found ketone esters were safe and tolerable for 12 weeks in healthy older adults.

The Kouvalchouk Procedure versus Distal Tibial Allograft for Treatment of Posterior Shoulder Instability: The Deltoid "Hammock" Effect Exists.

BACKGROUND: In 1993, Kouvalchouk described an acromial bone block with a pedicled deltoid flap for the treatment of posterior shoulder instability. This procedure provides a "double blocking" effect in that the acromial autograft restores posterior glenoid bone loss and the deltoid flap functions as a muscular "hammock" resembling the sling effect of the conjoint in the Latarjet procedure. The primary aim of this study was to compare the Kouvalchouk procedure to distal tibial allograft (DTA) reconstruction for the management of posterior shoulder instability with associated bone loss, while the secondary aim was to evaluate the deltoid hammock effect. s METHODS: Ten upper extremity cadavers were evaluated using a validated shoulder testing apparatus in 0° and 60° of glenohumeral abduction in the scapular plane. Testing was first performed on the normal shoulder state and was followed by the creation of a 20% posterior glenoid defect. Subsequently, the Kouvalchouk and DTA procedures were conducted. Forces of 0N, 5N, 10N and 15N were applied to the posterior deltoid tendinous insertion on the Kouvalchouk graft along the physiological muscle line-of-action to evaluate the 'hammock" effect of this procedure. Testing was additionally performed on the Kouvalchouk bone graft with the deltoid muscle sectioned from its bony attachment. For all test states, a posteriorly directed force was applied to the humeral head perpendicular to the direction of the glenoid bone defect, with the associated translation quantified using an optical tracking system. The outcome variable was posterior translation of the humeral head at an applied force magnitude of 30N. RESULTS: The Kouvalchouk procedure with the loaded deltoid flap (10N: P=0.039 and 15N: P<0.001) was significantly better at reducing posterior humeral head translation than the DTA. Overall, increased glenohumeral stability was observed with increased force applied to the posterior deltoid flap in the Kouvalchouk procedure. The 15N Kouvalchouk was most effective at preventing posterior humeral translation, and the difference was statistically significant compared with the 20% glenoid defect (P=0.003), detached Kouvalchouk (P<0.001), and 0N Kouvalchouk (P<0.001). The 15N Kouvalchouk procedure restored posterior shoulder joint stability to near normal levels, such that it was not significantly different from the intact state (P=0.203). CONCLUSIONS: The Kouvalchouk procedure with load applied to the deltoid was found to be biomechanically superior to the DTA for the management of posterior shoulder instability with associated bone loss. Additionally, the results confirmed the presence and effectiveness of the deltoid "hammock" effect.

First-in-Human Assessment of Gut Permeability in Crohn's Disease Patients Using Fluorophore Technology.

Background and Aims: The dual sugar absorption test as a classic measure of human intestinal permeability has limited clinical utility due to lengthy and cumbersome urine collection, assay variability, and long turnaround. We aimed to determine if the orally administered fluorophore MB-102 (relmapirazin) (molecular weight [MW] = 372) compares to lactulose (L) (MW = 342) and rhamnose (R) (MW = 164)-based dual sugar absorption test as a measure of gut permeability in people with a spectrum of permeability including those with Crohn's disease (CD). Methods: We performed a single-center, randomized, open-label, crossover study comparing orally administered MB-102 (1.5 or 3.0 mg/kg) to L (1000 mg) and R (200 mg). Adults with active small bowel CD on magnetic resonance enterography (cases) and healthy adults (controls) were randomized to receive either MB-102 or L and R on study day 1, and the other tracer 3 to 7 days later. Urine was collected at baseline and 1, 2, 4, 6, 8, 10, and 12 hours after tracer ingestion to calculate the cumulative urinary percent excretion of MB-102 and L and R. Results: Nine cases and 10 controls completed the study without serious adverse events. Urinary recovery of administered MB-102 correlated with recovery of lactulose (r-squared = 0.83) for all participants. MB-102 urine recovery was also tracked with the L:R ratio urine recovery (r-squared = 0.57). In controls, the percentages of L and MB-102 recovered were similar within a narrow range, unlike in CD patients. Conclusion: This first-in-human study of an orally administered fluorophore to quantify gastrointestinal permeability in adults with CD demonstrates that MB-102 is well tolerated, and its recovery in urine mirrors that of percent L and the L:R ratio.

A secondary analysis of indices of hepatic and beta cell function following 12 weeks of carbohydrate and energy restriction vs. free-living control in adults with type 2 diabetes.

BACKGROUND: Substantial weight loss in people living with type 2 diabetes (T2D) can reduce the need for glucose-lowering medications while concurrently lowering glycemia below the diagnostic threshold for the disease. Furthermore, weight-loss interventions have also been demonstrated to improve aspects of underlying T2D pathophysiology related to ectopic fat in the liver and pancreatic beta-cell function. As such, the purpose of this secondary analysis was to explore the extent to which a low-carbohydrate and energy-restricted (LCER) diet intervention improves markers of beta-cell stress/function, liver fat accumulation, and metabolic related liver function in people with type 2 diabetes. METHODS: We conducted secondary analyses of blood samples from a larger pragmatic community-based parallel-group randomized controlled trial involving a 12-week pharmacist implemented LCER diet (Pharm-TCR: <50 g carbohydrates; ~850-1100 kcal/day; n = 20) versus treatment-as-usual (TAU; n = 16). Participants were people with T2D, using ≥ 1 glucose-lowering medication, and a body mass index of ≥ 30 kg/m2. Main outcomes were C-peptide to proinsulin ratio, circulating microRNA 375 (miR375), homeostatic model assessment (HOMA) beta-cell function (B), fatty liver index (FLI), hepatic steatosis index (HSI), HOMA insulin resistance (IR), and circulating fetuin-A and fibroblast growth factor 21 (FGF21). Data were analysed using linear regression with baseline as a covariate. RESULTS: There was no observed change in miR375 (p = 0.42), C-peptide to proinsulin ratio (p = 0.17) or HOMA B (p = 0.15). FLI and HSI were reduced by -25.1 (p < 0.0001) and - 4.9 (p < 0.0001), respectively. HOMA IR was reduced by -46.5% (p = 0.011). FGF21 was reduced by -161.2pg/mL (p = 0.035) with a similar tendency found for fetuin-A (mean difference: -16.7ng/mL; p = 0.11). These improvements in markers of hepatic function were accompanied by reductions in circulating metabolites linked to hepatic insulin resistance (e.g., diacylglycerols, ceramides) in the Pharm TCR group. CONCLUSIONS: The Pharm-TCR intervention did not improve fasting indices of beta-cell stress; however, markers of liver fat accumulation and and liver function were improved, suggesting that a LCER diet can improve some aspects of the underlying pathophysiology of T2D. TRIAL REGISTRATION: Clinicaltrials.gov (NCT03181165).

Changes in Inhaled Nitric Oxide Use Across ICUs After Implementation of a Standard Pathway.

OBJECTIVES: Inhaled nitric oxide (iNO) is a selective pulmonary vasodilator. It is expensive, frequently used, and not without risk. There is limited evidence supporting a standard approach to initiation and weaning. Our objective was to optimize the use of iNO in the cardiac ICU (CICU), PICU, and neonatal ICU (NICU) by establishing a standard approach to iNO utilization. DESIGN: A quality improvement study using a prospective cohort design with historical controls. SETTING: Four hundred seven-bed free standing quaternary care academic children's hospital. PATIENTS: All patients on iNO in the CICU, PICU, and NICU from January 1, 2017 to December 31, 2022. INTERVENTIONS: Unit-specific standard approaches to iNO initiation and weaning. MEASUREMENTS AND MAIN RESULTS: Sixteen thousand eighty-seven patients were admitted to the CICU, PICU, and NICU with 9343 in the pre-iNO pathway era (January 1, 2017 to June 30, 2020) and 6744 in the postpathway era (July 1, 2020 to December 31, 2022). We found a decrease in the percentage of CICU patients initiated on iNO from 17.8% to 11.8% after implementation of the iNO utilization pathway. We did not observe a change in iNO utilization between the pre- and post-iNO pathway eras in either the PICU or NICU. Based on these data, we estimate 564 total days of iNO (-24%) were saved over 24 months in association with the standard pathway in the CICU, with associated cost savings. CONCLUSIONS: Implementation of a standard pathway for iNO use was associated with a statistically discernible reduction in total iNO usage in the CICU, but no change in iNO use in the NICU and PICU. These differential results likely occurred because of multiple contextual factors in each care setting.

Passive high explosive neutron inspection (PHENIX): a new method to confirm the presence or absence of high explosives for nuclear treaty verification.

Advanced instruments and methods need to be developed now to create a technical basis to support the negotiation of future nuclear arms control treaties. One new capability that is anticipated is the ability to confirm either the declared presence or declared absence of high explosive (HE) material in the presence of special nuclear material (SNM). Towards this goal, Passive HE Neutron Inspection (PHENIX) has been developed and demonstrated as a method for confirming the presence or absence of HE in the presence of plutonium. The method exploits the inherent presence of neutrons associated with the decay of plutonium as an internal probe source for performing prompt gamma-ray neutron activation analysis (PGNAA), searching for the presence of HE as revealed by the emission of characteristic gamma rays following neutron absorption in hydrogen and nitrogen which are building blocks of present-day, military-grade HE. Tests using stoichiometrically-correct hemishells of mock HE with plutonium show that a system can be expected to positively confirm the presence or absence of these signatures, supporting determination of HE presence or absence with Pu, in a few hours. To protect other potentially sensitive gamma-ray signatures from a treaty accountable item, an analog information barrier has been conceptualized and tested which physically prevents the collection of gamma-ray spectral data outside of user selected energy windows strategically chosen to view only narrow spectral regions corresponding to the hydrogen (2223.2 keV) and nitrogen (9807.2 keV, 10,318.2 keV, and 10,829.2 keV) PGNAA signatures.

Peripheral HMGB1 is linked to O3 pathology of disease-associated astrocytes and amyloid.

INTRODUCTION: Ozone (O3) is an air pollutant associated with Alzheimer's disease (AD) risk. The lung-brain axis is implicated in O3-associated glial and amyloid pathobiology; however, the role of disease-associated astrocytes (DAAs) in this process remains unknown. METHODS: The O3-induced astrocyte phenotype was characterized in 5xFAD mice by spatial transcriptomics and proteomics. Hmgb1fl/fl LysM-Cre+ mice were used to assess the role of peripheral myeloid cell high mobility group box 1 (HMGB1). RESULTS: O3 increased astrocyte and plaque numbers, impeded the astrocyte proteomic response to plaque deposition, augmented the DAA transcriptional fingerprint, increased astrocyte-microglia contact, and reduced bronchoalveolar lavage immune cell HMGB1 expression in 5xFAD mice. O3-exposed Hmgb1fl/fl LysM-Cre+ mice exhibited dysregulated DAA mRNA markers. DISCUSSION: Astrocytes and peripheral myeloid cells are critical lung-brain axis interactors. HMGB1 loss in peripheral myeloid cells regulates the O3-induced DAA phenotype. These findings demonstrate a mechanism and potential intervention target for air pollution-induced AD pathobiology. HIGHLIGHTS: Astrocytes are part of the lung-brain axis, regulating how air pollution affects plaque pathology. Ozone (O3) astrocyte effects are associated with increased plaques and modified by plaque localization. O3 uniquely disrupts the astrocyte transcriptomic and proteomic disease-associated astrocyte (DAA) phenotype in plaque associated astrocytes (PAA). O3 changes the PAA cell contact with microglia and cell-cell communication gene expression. Peripheral myeloid cell high mobility group box 1 regulates O3-induced transcriptomic changes in the DAA phenotype.

Stabilizing effect of an elbow orthosis with an adjustable hinge axis after lateral collateral ligament injury: A biomechanical study.

Background: Current commercial elbow braces have a straight hinge that does not account for the native carrying angle of the elbow. The objective of this study was to determine the effectiveness of a custom-designed hinged elbow orthosis (HEO) with variable valgus angulations in stabilizing a lateral collateral ligament (LCL) deficient elbow. Methods: Eight cadaveric upper extremities were mounted in an elbow motion simulator in the abducted varus gravity-loaded position. The specimens were examined before and after simulated LCL injury and then with the addition of the custom-designed HEO with 0°, 10°, and 20° of valgus angulation. Kinematic data were recorded using an electromagnetic tracking system. Results: The LCL injured state with or without the brace resulted in significant increases in varus angulation of the elbow compared to the intact state in both pronation and supination (P < 0.05). There were no significant differences in varus-valgus angulation or ulnohumeral rotation between any of the brace angles and the LCL injured state with the forearm pronated and supinated. Discussion: The custom-designed HEO did not provide any additional stability to the LCL injured elbow. The varus arm position should be avoided during the rehabilitation of an LCL injured elbow even when an HEO is used.

Blood flow restriction training does not negatively alter the mechanical strength or histomorphology of uninjured equine superficial digital flexor tendons.

BACKGROUND: Low load exercise training with blood flow restriction (BFR) has become increasingly used by human physical therapists to prescribe controlled exercise following orthopaedic injury; its effects on the equine superficial digital flexor tendon (SDFT), however, are unknown. OBJECTIVE: To investigate outcomes of pressure specific BFR walking exercise on uninjured equine SDFT biomechanics and histomorphology. STUDY DESIGN: Controlled in vivo experiment. METHODS: Four forelimbs of four horses were exposed to 40 BFR-walk sessions (10-min interval walking) on a treadmill over a 56-day study period with their contralateral forelimbs serving as untreated controls. Similarly, four forelimbs of four control horses were exposed to 40 sham cuff walk sessions. On study Day 56, all horses (n = 8) were humanely euthanised and forelimb SDFTs underwent non-destructive biomechanical testing and corresponding histomorphological analysis. Significance in biomechanical parameters between treatment groups was analysed using a mixed-effects ANOVA with Tukey's post-hoc tests. RESULTS: Statistically significant differences in SDFT stiffness for both first (p = 0.02) and last cycles (p = 0.03) were appreciated within the BFR treated group only, with BFR exposed forelimbs being significantly stiffer than the contralateral unexposed forelimbs. When normalised to cross-sectional area, no significant differences were appreciated among treatment groups in elastic modulus for the first (p = 0.5) or last cycles (p = 0.4). No histological differences were appreciated among treatment groups according to Bonar, Movin, or musculotendinous junction evaluation criteria. MAIN LIMITATIONS: Short-term comparisons were performed in a small sample population without correlation to performance outcome measures. Optimal occlusion percentages and walk protocols remain unknown. CONCLUSIONS: This study demonstrated no negative impact of BFR on mechanical strength of the equine SDFT; however, evidence suggests that BFR results in increased tendon stiffness based on biomechanical testing and subsequent calculations. No consistent detrimental histomorphological changes were seen.

CONTEXTO: Exercício de baixa carga com restrição do fluxo sanguíneo (RFS) tem sido cada vez mais utilizado por fisioterapeutas humanos para tratar lesões ortopédicas. Porém, seus efeitos no tendão flexor digital superficial (TFDS) de equinos não é conhecida. OBJETIVOS: O objetivo deste estudo foi investigar o efeito de específicas pressões com RFS durante o passo em cavalos sem lesão no TFDS, por meio de histologia e análise biomecânica. DELINEAMENTO DO ESTUDO: Estudo controlado. MÉTODOS: Quatro membros torácicos de quatro cavalos foram expostos a 40 sessões de RFS durante o passo (10 minutos de caminhada intervalada), ao longo de 56 dias. O membro contralateral foi utilizado como controle. Da mesma forma, quatro membros de quatro cavalos controle foram expostos a 40 sessões simuladas de caminhada com torniquete. No dia 56, todos os cavalos (n = 8) foram eutanasiados, e os TFDS foram submetidos a testes biomecânicos não destrutivos e análise histológica. A significância dos parâmetros biomecânicos entre tratamentos foi analisada utilizando ANOVA de efeitos mistos, seguida pelo teste de Tukey. RESULTADOS: A rigidez do TFDS foi estatisticamente diferente nos primeiros (p = 0.02) e últimos (p = 0.03) ciclos no grupo submetido à RFS, sendo os membros tratados significativamente mais rígidos do que os membros contralaterais não expostos ao tratamento. Quando normalizado para a área transversal, não foi observada diferença significativa entre os grupos de tratamento no módulo de elasticidade para os primeiros (p = 0.5) e últimos (p = 0.4) ciclos. Não foram identificadas diferenças histológicas nos diferentes tipos de tratamento, de acordo com os critérios de avaliação Bonar, Movin e de junção musculo­tendínea. PRINCIPAIS LIMITAÇÕES: Comparações de curto prazo foram realizadas em uma amostra pequena da população, sem correlação com medidas de resultados de desempenho. As porcentagens ideais de oclusão e os protocolos de caminhada permanecem desconhecidos. CONCLUSÕES: Este estudo não demonstrou impacto negativo do RFS na resistência mecânica do TFDS equino; no entanto, as evidências sugerem que a RFS resulta em aumento da rigidez do tendão com base em testes biomecânicos e cálculos subsequentes. Nenhuma alteração histológica prejudicial consistente foi observada.

Pragmatic Approach to In Situ Simulation to Identify Latent Safety Threats Before Moving to a Newly Built ICU.

OBJECTIVES: Transitions to new care environments may have unexpected consequences that threaten patient safety. We undertook a quality improvement project using in situ simulation to learn the new patient care environment and expose latent safety threats before transitioning patients to a newly built adult ICU. DESIGN: Descriptive review of a patient safety initiative. SETTING: A newly built 24-bed neurocritical care unit at a tertiary care academic medical center. SUBJECTS: Care providers working in neurocritical care unit. INTERVENTIONS: We implemented a pragmatic three-stage in situ simulation program to learn a new patient care environment, transitioning patients from an open bay unit to a newly built private room-based ICU. The project tested the safety and efficiency of new workflows created by new patient- and family-centric features of the unit. We used standardized patients and high-fidelity mannequins to simulate patient scenarios, with "test" patients created through all electronic databases. Relevant personnel from clinical and nonclinical services participated in simulations and/or observed scenarios. We held a debriefing after each stage and scenario to identify safety threats and other concerns. Additional feedback was obtained via a written survey sent to all participants. We prospectively surveyed for missed latent safety threats for 2 years following the simulation and fixed issues as they arose. MEASUREMENTS AND MAIN RESULTS: We identified and addressed 70 latent safety threats, including issues concerning physical environment, infection prevention, patient workflow, and informatics before the move into the new unit. We also developed an orientation manual that highlighted new physical and functional features of the ICU and best practices gleaned from the simulations. All participants agreed or strongly agreed that simulations were beneficial. Two-year follow-up revealed only two missed latent safety threats. CONCLUSIONS: In situ simulation effectively identifies latent safety threats surrounding the transition to new ICUs and should be considered before moving into new units.