Discovering Novel Proteoforms Using Proteogenomic Workflows Within the Galaxy Bioinformatics Platform.

Proteogenomics is a growing "multi-omics" research area that combines mass spectrometry-based proteomics and high-throughput nucleotide sequencing technologies. Proteogenomics has helped in genomic annotation for organisms whose complete genome sequences became available by using high-throughput DNA sequencing technologies. Apart from genome annotation, this multi-omics approach has also helped researchers confirm expression of variant proteins belonging to unique proteoforms that could have resulted from single-nucleotide polymorphism (SNP), insertion and deletions (Indels), splice isoforms, or other genome or transcriptome variations.A proteogenomic study depends on a multistep informatics workflow, requiring different software at each step. These integrated steps include creating an appropriate protein sequence database, matching spectral data against these sequences, and finally identifying peptide sequences corresponding to novel proteoforms followed by variant classification and functional analysis. The disparate software required for a proteogenomic study is difficult for most researchers to access and use, especially those lacking computational expertise. Furthermore, using them disjointedly can be error-prone as it requires setting up individual parameters for each software. Consequently, reproducibility suffers. Managing output files from each software is an additional challenge. One solution for these challenges in proteogenomics is the open-source Web-based computational platform Galaxy. Its capability to create and manage workflows comprised of disparate software while recording and saving all important parameters promotes both usability and reproducibility. Here, we describe a workflow that can perform proteogenomic analysis on a Galaxy-based platform. This Galaxy workflow facilitates matching of spectral data with a customized protein sequence database, identifying novel protein variants, assessing quality of results, and classifying variants along with visualization against the genome.

SARS-CoV-2 within-host population expansion, diversification and adaptation in zoo tigers, lions and hyenas.

SARS-CoV-2 rapidly adapts to new hosts following cross-species transmission; this is highly relevant as novel within-host variants have emerged following infection of susceptible wild and domestic animal species. Furthermore, SARS-CoV-2 transmission from animals (e.g., white-tailed deer, mink, domestic cats, and others) back to humans has also been observed, documenting the potential of novel animal-derived variants to infect humans. We investigated SARS-CoV-2 evolution and host-specific adaptation during an outbreak in Amur tigers (Panthera tigris altaica), African lions (Panthera leo), and spotted hyenas (Crocuta crocuta) at Denver Zoo in late 2021. SARS-CoV-2 genomes from longitudinal samples collected from 16 individuals were evaluated for within-host variation and genomic signatures of selection. The outbreak was likely initiated by a single spillover of a rare Delta sublineage subsequently transmitted from tigers to lions to hyenas. Within-host virus populations rapidly expanded and diversified. We detected signatures of purifying and positive selection, including strong positive selection in hyenas and in the nucleocapsid (N) gene in all animals. Four candidate species-specific adaptive mutations were identified: N A254V in lions and hyenas, and ORF1a E1724D, spike T274I, and N P326L in hyenas. These results reveal accelerated SARS-CoV-2 adaptation following host shifts in three non-domestic species in daily contact with humans.

HumanIslets.com: Improving accessibility, integration, and usability of human research islet data.

HumanIslets.com supports diabetes research by offering easy access to islet phenotyping data, analysis tools, and data download. It includes molecular omics, islet and cellular function assays, tissue processing metadata, and phenotypes from 547 donors. As it expands, the resource aims to improve human islet data quality, usability, and accessibility.

Ceftriaxone-resistant Neisseria gonorrhoeae detected in England, 2015-24: an observational analysis.

OBJECTIVES: Since June 2022, there has been a rise in the number of ceftriaxone-resistant Neisseria gonorrhoeae cases detected in England (n = 15), of which a third were XDR. We describe the demographic and clinical details of the recent cases and investigate the phenotypic and molecular characteristics of the isolates. For a comprehensive overview, we also reviewed 16 ceftriaxone-resistant cases previously identified in England since December 2015 and performed a global genomic comparison of all publicly available ceftriaxone-resistant N. gonorrhoeae strains with mosaic penA alleles. METHODS: All N. gonorrhoeae isolates resistant to ceftriaxone (MIC > 0.125 mg/L) were whole-genome sequenced and compared with 142 global sequences of ceftriaxone-resistant N. gonorrhoeae. Demographic, behavioural and clinical data were collected. RESULTS: All cases were heterosexual, and most infections were associated with travel from the Asia-Pacific region. However, some had not travelled outside England within the previous few months. There were no ceftriaxone genital treatment failures, but three of five pharyngeal infections and the only rectal infection failed treatment. The isolates represented 13 different MLST STs, and most had the mosaic penA-60.001 allele. The global genomes clustered into eight major phylogroups, with regional associations. All XDR isolates belonged to the same phylogroup, represented by MLST ST16406. CONCLUSIONS: Most cases of ceftriaxone-resistant N. gonorrhoeae detected in England were associated with travel from the Asia-Pacific region. All genital infections were successfully treated with ceftriaxone, but there were extragenital treatment failures. Ceftriaxone resistance continues to be associated with the penA-60.001 allele within multiple genetic backgrounds and with widespread dissemination in the Asia-Pacific region.

Migratory strategies across an ecological barrier: is the answer blowing in the wind?

BACKGROUND: Ecological barriers can shape the movement strategies of migratory animals that navigate around or across them, creating migratory divides. Wind plays a large role in facilitating aerial migrations and can temporally or spatially change the challenge posed by an ecological barrier, with beneficial winds potentially converting a barrier into a corridor. Here, we explore the role wind plays in shaping initial southbound migration strategy among individuals breeding at two sites along an ecological barrier. METHODS: Using GPS satellite transmitters, we tracked the southbound migrations of Short-billed Dowitchers (Limnodromus griseus caurinus) from two breeding sites in Alaska to nonbreeding sites in coastal Mexico. The breeding sites were positioned in distinct regions along an ecological barrier - the Gulf of Alaska. We investigated potential differences in migratory timing, wind availability, and tailwind support en route across the Gulf of Alaska between individuals breeding at the two sites. RESULTS: Route choice and arrival timing to wintering sites differed markedly between the two breeding sites: individuals departing from the more westerly site left at the same time as those from further east but crossed the Gulf of Alaska farther west and arrived along the Pacific coast of Mexico an average of 19 days earlier than their counterparts. Dowitchers from both sites departed with slight tailwinds, but once aloft over the Gulf of Alaska, birds from the more westerly site had up to twelve times more tailwind assistance than birds from the more easterly one. CONCLUSIONS: The distinct migration strategies and degree of wind assistance experienced by birds at these two breeding sites demonstrates how differences in wind availability along migratory routes can form the basis for intraspecific variation in migration strategies with potential carryover effects. Future changes in wind regimes may therefore interact with changes in habitat availability to influence migration patterns and migratory bird conservation.

Exploratory functional and quality of life outcomes with daily consumption of the ketone ester bis-octanoyl (R)-1,3-butanediol in healthy older adults: a randomized, parallel arm, double-blind, placebo-controlled study.

Background: Ketone bodies are metabolites produced during fasting or on a ketogenic diet that have pleiotropic effects on the inflammatory and metabolic aging pathways underpinning frailty in in vivo models. Ketone esters (KEs) are compounds that induce hyperketonemia without dietary changes and that may impact physical and cognitive function in young adults. The functional effects of KEs have not been studied in older adults. Objectives: Our long-term goal is to examine if KEs modulate aging biology mechanisms and clinical outcomes relevant to frailty in older adults. Here, we report the exploratory functional and quality-of-life outcome measures collected during a 12-week safety and tolerability study of KE (NCT05585762). Design: Randomized, placebo-controlled, double-blinded, parallel-group, pilot trial of 12-weeks of daily KE ingestion. Setting: The Clinical Research Unit at the Buck Institute for Research on Aging, California. Participants: Community-dwelling older adults (≥ 65 years), independent in activities of daily living, with no unstable acute medical conditions (n = 30). Intervention: Subjects were randomly allocated (1:1) to consume 25 g daily of either KE (bis-octanoyl (R)-1,3-butanediol) or a taste, appearance, and calorie-matched placebo (PLA) containing canola oil. Measurements: Longitudinal change in physical function, cognitive function and quality of life were assessed as exploratory outcomes in n = 23 completers (n = 11 PLA, n = 12 KE). A composite functional outcome to describe the vigor-frailty continuum was calculated. Heart rate and activity was measured throughout the study using digital wearables. Results: There were no statistically significant longitudinal differences between groups in exploratory functional, activity-based or quality of life outcomes. Conclusion: Daily ingestion of 25 g of KE did not affect exploratory functional or quality-of-life end points in this pilot cohort of healthy older adults. Future work will address these endpoints as primary and secondary outcomes in a larger trial of pre-frail older adults.

SARS-CoV-2 outbreak in lions, tigers and hyenas at Denver Zoo.

In late 2019, SARS-CoV-2 spilled-over from an animal host into humans, where it efficiently spread, resulting in the COVID-19 pandemic. Through both natural and experimental infections, we learned that many animal species are susceptible to SARS-CoV-2. Importantly, animals in close proximity to humans, including companion, farmed, and those at zoos and aquariums, became infected, and many studies demonstrated transmission to/from humans in these settings. In this study, we first review the literature of SARS-CoV-2 infections in tigers and lions, and compare species, sex, age, virus and antibody detection assay, and types, frequency and length of clinical signs, demonstrating broad heterogeneity amongst infections. We then describe a SARS-CoV-2 outbreak in lions, tigers and hyenas at Denver Zoo in late 2021. Animals were tested for viral RNA (vRNA) for four months. Lions had significantly more viral RNA in nasal swabs than both tigers and hyenas, and many individual lions experienced viral recrudescence after weeks of undetectable vRNA. Infectious virus was correlated with high levels of vRNA and was more likely to be detected earlier during infection. Four months post-infection, all tested animals generated robust neutralizing antibody titers. Animals were infected with Delta lineage AY.20 identical to a variant circulating at less than 1% in Colorado humans at that time, suggesting a single spillover event from an infected human spread within and between species housed at the zoo. Better understanding of epidemiology and susceptibility of SARS-CoV-2 infections in animals is critical to limit the current and future spread and protect animal and human health.

Pyruvate dehydrogenase kinase 1 controls triacylglycerol hydrolysis in cardiomyocytes.

Pyruvate dehydrogenase kinase (PDK) 1 is one of four isozymes that inhibit the oxidative decarboxylation of pyruvate to acetyl-CoA via pyruvate dehydrogenase. PDK activity is elevated in fasting or starvation conditions to conserve carbohydrate reserves. PDK has also been shown to increase mitochondrial fatty acid utilization. In cardiomyocytes, metabolic flexibility is crucial for the fulfillment of high energy requirements. The PDK1 isoform is abundant in cardiomyocytes, but its specific contribution to cardiomyocyte metabolism is unclear. Here we show that PDK1 regulates cardiomyocyte fuel preference by mediating triacylglycerol turnover in differentiated H9c2 myoblasts using lentiviral shRNA to knockdown Pdk1. Somewhat surprisingly, PDK1 loss did not affect overall PDH activity, basal glycolysis, or glucose oxidation revealed by oxygen consumption rate experiments and 13C6 glucose labelling. On the other hand, we observed decreased triacylglycerol turnover in H9c2 cells with PDK1 knockdown, which was accompanied by decreased mitochondrial fatty acid utilization following nutrient deprivation. 13C16 palmitate tracing of uniformly labelled acyl chains revealed minimal acyl chain shuffling within triacylglycerol, indicating that the triacylglycerol hydrolysis, and not re-esterification, was dysfunctional in PDK1 suppressed cells. Importantly, PDK1 loss did not significantly impact the cellular lipidome or triacylglycerol accumulation following palmitic acid treatment, suggesting that effects of PDK1 on lipid metabolism were specific to the nutrient-deprived state. We validated that PDK1 loss decreased triacylglycerol turnover in Pdk1 knockout mice. Together, these findings implicate a novel role for PDK1 in lipid metabolism in cardiomyocytes, independent of its canonical roles in glucose metabolism.

Biomechanical Performance of 4-Leg Sustained Dynamic Compression Staples in First Tarsometatarsal Arthrodesis.

BACKGROUND: This study investigates the biomechanical efficacy of new 4-leg Sustained Dynamic Compression (SDC) NiTiNOL staples, hypothesized to offer superior stability and resilience to loading before fusion completion, compared with conventional hardware. METHODS: Twenty sawbones left full foot models were divided into 4 treatment groups: (1) 4-leg Inline Staple, (2) 4-leg Inline Staple + 2-leg Staple, (3) 4-leg Inline Staple + Screw, and (4) Plate + Screw. An osteotomy was performed to simulate a Lapidus procedure, and the respective fixation methods were applied. Mechanical testing was conducted using a servo-hydraulic testing machine to evaluate constructs' load, contact force, contact area, and plantar gap. RESULTS: The 4-leg Inline Staple + Screw group demonstrated significantly increased joint contact force, joint contact area, and decreased plantar gap compared with the Plate + Screw group, both before and after cyclic testing. All SDC-containing constructs exhibited post-cyclic joint contact areas that were 2.36×, 3.87×, and 5.49× greater than the post-cyclic plate + screw group. Most notably, the 4-leg Inline Staple + Screw group maintained a plantar gap of less than 3 mm throughout the testing, unlike other groups. CONCLUSIONS: The 4-leg Inline SDC Staple, particularly when combined with a static screw, demonstrated biomechanical superiority over traditional plate and screw constructs in Lapidus procedures. These findings suggest a promising avenue for enhanced post-operative stability, which could translate into quicker patient recovery, improved fusion rates, and potentially lower non-union rates. Further clinical trials are warranted to validate these biomechanical advantages in patient outcomes. LEVEL OF EVIDENCE: Therapeutic, Level V: Bench Testing.

Association between delayed outbreak identification and SARS-CoV-2 infection and mortality among long-term care home residents, Ontario, Canada, March to November 2020: a cohort study.

BackgroundLate outbreak identification is a common risk factor mentioned in case reports of large respiratory infection outbreaks in long-term care (LTC) homes.AimTo systematically measure the association between late SARS-CoV-2 outbreak identification and secondary SARS-CoV-2 infection and mortality in residents of LTC homes.MethodsWe studied SARS-CoV-2 outbreaks across LTC homes in Ontario, Canada from March to November 2020, before the COVID-19 vaccine rollout. Our exposure (late outbreak identification) was based on cumulative infection pressure (the number of infectious resident-days) on the outbreak identification date (early: ≤ 2 infectious resident-days, late: ≥ 3 infectious resident-days), where the infectious window was -2 to +8 days around onset. Our outcome consisted of 30-day incidence of secondary infection and mortality, based on the proportion of at-risk residents with a laboratory-confirmed SARS-CoV-2 infection with onset within 30 days of the outbreak identification date.ResultsWe identified 632 SARS-CoV-2 outbreaks across 623 LTC homes. Of these, 36.4% (230/632) outbreaks were identified late. Outbreaks identified late had more secondary infections (10.3%; 4,437/42,953) and higher mortality (3.2%; 1,374/42,953) compared with outbreaks identified early (infections: 3.3%; 2,015/61,714; p < 0.001, mortality: 0.9%; 579/61,714; p < 0.001). After adjustment for 12 LTC home covariates, the incidence of secondary infections in outbreaks identified late was 2.90-fold larger than that of outbreaks identified early (OR: 2.90; 95% CI: 2.04-4.13).ConclusionsThe timeliness of outbreak identification could be used to predict the trajectory of an outbreak, plan outbreak measures and retrospectively provide feedback for quality improvement, with the objective of reducing the impacts of respiratory infections in LTC home residents.

Rationale and protocol for a safety, tolerability and feasibility randomized, parallel arm, double-blind, placebo-controlled, pilot study of a novel ketone ester targeting frailty via immunometabolic geroscience mechanisms.

BACKGROUND: Frailty is a geriatric syndrome characterized by chronic inflammation and metabolic insufficiency that creates vulnerability to poor outcomes with aging. We hypothesize that interventions which target common underlying mechanism of aging could ameliorate frailty. Ketone bodies are metabolites produced during fasting or on a ketogenic diet that have pleiotropic effects on inflammatory and metabolic aging pathways in laboratory animal models. Ketone esters (KEs) are compounds that induce ketosis without dietary changes, but KEs have not been studied in an older adult population. Our long-term goal is to examine if KEs modulate aging biology mechanisms and clinical outcomes relevant to frailty in older adults. OBJECTIVES: The primary objective of this randomized, placebo-controlled, double-blinded, parallel-group, pilot trial is to determine tolerability of 12-weeks of KE ingestion in a broad population of older adults (≥ 65 years). Secondary outcomes include safety and acute blood ketone kinetics. Exploratory outcomes include physical function, cognitive function, quality of life, aging biomarkers and inflammatory measures. METHODS: Community-dwelling adults who are independent in activities of daily living, with no unstable acute medical conditions (n = 30) will be recruited. The study intervention is a KE or a taste, appearance, and calorie matched placebo beverage. Initially, acute 4-hour ketone kinetics after 12.5g or 25g of KE consumption will be assessed. After collection of baseline safety, functional, and biological measurements, subjects will randomly be allocated to consume KE 25g or placebo once daily for 12-weeks. Questionnaires will assess tolerability daily for 2-weeks, and then via phone interview at bi-monthly intervals. Safety assessments will be repeated at week 4. All measures will be repeated at week 12. CONCLUSION: This study will evaluate feasibility, tolerability, and safety of KE consumption in older adults and provide exploratory data across a range of aging-related endpoints. This data will inform design of larger trials to rigorously test KE effects on aging mechanisms and clinical outcomes relevant to frailty.

Efficacy of Self-Natural Posture Exercise (SNPE) programs on chronic low back pain: A randomized controlled feasibility trial with waitlist control.

BACKGROUND: Low back pain (LBP) is a significant source of disability and decreased quality of life. The Self-Natural Posture Exercise (SNPE) intervention can be used effectively in many cases, but feasibility and impact has not been fully explored. OBJECTIVE: The current study explores the feasibility and efficacy of face-to-face (FtF) and virtual (Vir) SNPE programs on chronic low back pain. METHODS: This is a randomized single-blinded waitlist control study with 10 participants in the FtF group (age 45.8 ± 2.89) and 9 in the Vir group (age 52.2 ± 2.3). Participants had low to moderate low back pain for > 3 months. Those who would eventually become the Vir group served initially as a waitlist control (Con) group. The FtF group received 12 weeks of in-person exercise training sessions conducted and the Vir group received weekly recorded training videos created by and featuring the same instructor. Measured outcomes included feasibility assessed by rate of retention and rate of attendance, level of low back pain (Oswestry Disability Index, Visual Analogue Scale), quality of life (36-Item Short Form Health Survey 1.0), muscular pressure pain threshold, and muscle tone. Analysis was done via repeated measures ANOVA and Wilcoxon tests. RESULTS: Rates of retention were 80% in the FtF group and 78% in the Vir group. Attendance rates were 88% in the FtF group and 60% in the Vir group. ODI, VAS, and some SF-36 domains improved in both the FtF and Vir groups. Overall, the FtF group improved in more domains and by a larger degree than the Vir group and both groups improved relative to Con. CONCLUSION: A 12-week SNPE program, done virtually or in person, shows promise in improving ODI, VAS, and some SF-36 domains. FtF appears to be more effective. Future studies would benefit from sampling a larger and more diverse population.

Innovating Flexor Tendon Repair Training with a Three-dimensional Printed Model.

Background: Flexor tendon repair is a technically demanding procedure, with functional outcome directly proportional to skillful execution. A repair must be strong to manage early mobilization and precise to allow for gliding through the tendon sheath. As a result, junior residents face a steep learning curve that may be mitigated by exposure to surgical simulators. Methods: To facilitate flexor tendon repair training, a surgical training device and accompanying instructional video were developed. Simulation workshops were held for junior orthopedic and plastic surgery residents (n = 11). To objectively assess validity of the curriculum, study participants performed cadaveric flexor tendon repairs before and after the workshop. Anonymous recordings of these repairs were graded by two certified hand surgeons. Additionally, a tensometer was used to measure strength of repair. Results: Model realism, educational utility, and overall usefulness rated high: 4.6 ±â€…0.52 95% confidence interval (CI) for realism, 4.9 ±â€…0.42 95% CI for device, 4.7 ±â€…0.96 95% CI for video, and 4.9 ±â€…0.66 95% CI overall. Subjective confidence increased after the training session (73% ranked "moderately" or "extremely"). Likewise, scores given by the surgeons grading the repairs improved for overall quality and time of repair (pre: 2.77 ±â€…0.61, post: 4.22 ±â€…0.56, P= 0.0002). Strength of repair did not change (P = 0.87). Conclusions: The proposed three-dimensional surgical simulator for flexor tendon repair is realistic and useful, with improved surgical technique and improved confidence demonstrated after use. This design can be three-dimensionally printed en masse and provide value to hand surgery training curriculum.

Inhibition of RHOA activity preserves the survival and hemostasis function of long-term cold-stored platelets.

ABSTRACT: Patients with thrombocytopenia require platelet transfusion to prevent and stop hemorrhage. Cold storage of platelets results in complex molecular lesions, including changes in membrane microdomains that are recognized by host macrophages and hepatocyte counter-receptors, resulting in phagocytosis and clearance upon transfusion. For this reason, platelets are stored at room temperature, a method that confers increased risk of bacterial contamination. By applying signaling analysis and genetic and pharmacological approaches, we identified that cold-induced activation of RAS homolog family, member A (RHOA) GTPase causes the major hallmarks of platelet cold storage lesions. RHOA deficiency renders murine platelets insensitive to cold storage-induced damage, and pharmacological inhibition by a RHOA activation inhibitor, R-G04, can prevent the cold storage-induced lesions. RHOA inhibition prevents myosin activation and clathrin-independent formation and internalization of lipid rafts enriched in active glycosyltransferases as well as abnormal distribution of GPIbα. RHOA inhibition further prevents the metabolic reprogramming of cold storage-induced lesions and allows the maintenance of glycolytic flux and mitochondria-dependent respiration. Importantly, human platelets transfused in mice after cold storage, in the presence of R-G04 or its more potent enantiomer S-G04, can circulate in vivo at similar levels as room temperature-stored platelets while retaining their hemostatic activity in vivo, as assessed by bleeding time correction in aspirin-treated mice. Our studies provide a mechanism-based translational approach to prevent cold storage-induced damage, which is useful for human platelet transfusion in patients with thrombocytopenia.

Superior Capsular Reconstruction using the Long Head of Biceps Tendon: A Biomechanical Assessment of Tenodesis Location and Angle of Fixation.

BACKGROUND: Massive irreparable rotator cuff tears (MIRCT) treated with superior capsular reconstruction (SCR) using the long head of the biceps tendon have shown satisfactory early results. Different techniques and positions for biceps tenodesis have been described. This study aimed to evaluate the effect of tenodesis location and glenohumeral fixation angle for graft tensioning on the biomechanics of a SCR using a single strand biceps technique. METHODS: Eight cadaveric shoulders were mounted to a custom biomechanical simulator which employed static tone loads to the deltoid and rotator cuff muscles. All cadavers were first tested in the intact condition, and then in the simulated MIRCT condition by sectioning the tendinous insertions of the supraspinatus and upper border of the infraspinatus. SCR using the long head of the biceps tendon was then evaluated. Three biceps tenodesis locations relative to the greater tuberosity (anterior, middle, and posterior) and two glenohumeral fixation angles (0° and 30°) for graft tensioning were tested. An optical tracking system was used to quantify superior-inferior (SI) and anterior-posterior (AP) humeral head translation relative to the glenoid, while the functional abduction force was quantified using a load sensor. All tests were conducted at 0°, 30° and 60° of glenohumeral abduction in a randomized fashion. RESULTS: When assessing isolated superior humeral head migration, all biceps tenodesis locations were effective at decreasing superior migration, with no tenodesis location significantly better than the other (P=0.213). However, biceps grafts tensioned at 30° of glenohumeral abduction were significantly better at reducing proximal humeral migration as compared to graft tensioning at 0° abduction (P=0.008). Posterior humeral head translation observed in the MIRCT condition was significantly reduced when tensioning the biceps tendon at 30° of glenohumeral abduction compared to 0° for all tenodesis locations (P≤0.043). Tenodesis location also significantly influenced posterior humeral head translation (P=0.001), with middle and posterior positions restoring near normal humeral head position when fixed at 30° glenohumeral abduction. All SCR techniques using the biceps tendon improved the functional abduction force relative to the MIRCT condition, although no statistically significant differences were observed relative to the intact condition (P≥0.448). DISCUSSION: SCR using the long head biceps tendon is biomechanically effective in reducing posterosuperior translation of the humeral head in the setting of a MIRCT. Graft tensioning and fixation at 30° of glenohumeral abduction combined with either a middle or posterior tenodesis location on the greater tuberosity most effectively restores near normal time-zero humeral head kinematics.

Daily consumption of ketone ester, bis-octanoyl (R)-1,3-butanediol, is safe and tolerable in healthy older adults in 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 assess the tolerability and safety of KE ingestion in a cohort of older adults. DESIGN: Randomized, placebo-controlled, double-blinded, parallel-arm trial (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; M = 15, F = 15; age = 76 y, range 65-90 y) were randomized and n = 23 (M = 14, F = 9) completed the protocol. INTERVENTION: Participants were randomly allocated to consume either KE (25 g bis-octanoyl (R)-1,3-butanediol) or a taste, appearance, and calorie-matched placebo (PLA) containing canola oil daily for 12 weeks. 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 and 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 this study of healthy older adults. These results provide an initial foundation for use of KEs in clinical research with older adults.

Stemless reverse shoulder arthroplasty neck shaft angle influences humeral component time-zero fixation and survivorship: a cadaveric biomechanical assessment.

Background: Stemless humeral components are being clinically investigated for reverse shoulder arthroplasty (RSA) procedures. There is, however, a paucity of basic science literature on the surgical parameters that influence the success of these procedures. Therefore, this cadaveric biomechanical study evaluated the neck shaft angle (NSA) of implantation on the survivability and performance of stemless RSA humeral components during cyclical loading. Methods: Twelve paired cadaveric humeri were implanted with stemless RSA humeral components at NSAs of 135° and 145°. Implant-bone motion at the periphery of the implant was measured with 3 optical machine vision USB3 cameras outfitted with c-mount premium lenses and quantified with ProAnalyst software. A custom 3-dimensional loading apparatus was used to cyclically apply 3 loading directions representative of physiological states at 5 progressively increasing loading magnitudes. Stemless 135° and 145° implants were compared based on the maximum implant-bone relative distraction detected, as well as the survivorship of the implants throughout the loading protocol. Results: Primary fixation and implant biomechanical survivorship were substantially better in the 145° NSA implants. The 135° NSA implants elicited significantly higher implant-bone distractions during cyclical loading (P = .001), and implant survivorship was considerably lower in the 135° NSA specimens when compared to the 145° NSA specimens (135° NSA: 0%, 145° NSA: 50%) (P < .001). Conclusion: NSA is a modifiable parameter that influences time-zero implant stability, as well as the early survivorship of the stemless RSA humeral components tested in this study. NSA resections of 145° appear to promote better stability than those utilizing 135° NSAs during early postoperative eccentric loads. Further studies are required to assess if other stemless reversed humeral implant designs have improved time-zero fixation at higher NSAs.

Adult Human Brain Tissue Cultures to Study NeuroHIV.

HIV-associated neurocognitive disorders (HAND) persist under antiretroviral therapy as a complex pathology that has been difficult to study in cellular and animal models. Therefore, we generated an ex vivo human brain slice model of HIV-1 infection from surgically resected adult brain tissue. Brain slice cultures processed for flow cytometry showed >90% viability of dissociated cells within the first three weeks in vitro, with parallel detection of astrocyte, myeloid, and neuronal populations. Neurons within brain slices showed stable dendritic spine density and mature spine morphologies in the first weeks in culture, and they generated detectable activity in multi-electrode arrays. We infected cultured brain slices using patient-matched CD4+ T-cells or monocyte-derived macrophages (MDMs) that were exposed to a GFP-expressing R5-tropic HIV-1 in vitro. Infected slice cultures expressed viral RNA and developed a spreading infection up to 9 days post-infection, which were significantly decreased by antiretrovirals. We also detected infected myeloid cells and astrocytes within slices and observed minimal effect on cellular viability over time. Overall, this human-centered model offers a promising resource to study the cellular mechanisms contributing to HAND (including antiretroviral toxicity, substance use, and aging), infection of resident brain cells, and new neuroprotective therapeutics.