Quinone-mediated hydrogen anode for non-aqueous reductive electrosynthesis.

Electrochemical synthesis can provide more sustainable routes to industrial chemicals1-3. Electrosynthetic oxidations may often be performed 'reagent-free', generating hydrogen (H2) derived from the substrate as the sole by-product at the counter electrode. Electrosynthetic reductions, however, require an external source of electrons. Sacrificial metal anodes are commonly used for small-scale applications4, but more sustainable options are needed at larger scale. Anodic water oxidation is an especially appealing option1,5,6, but many reductions require anhydrous, air-free reaction conditions. In such cases, H2 represents an ideal alternative, motivating the growing interest in the electrochemical hydrogen oxidation reaction (HOR) under non-aqueous conditions7-12. Here we report a mediated H2 anode that achieves indirect electrochemical oxidation of H2 by pairing thermal catalytic hydrogenation of an anthraquinone mediator with electrochemical oxidation of the anthrahydroquinone. This quinone-mediated H2 anode is used to support nickel-catalysed cross-electrophile coupling (XEC), a reaction class gaining widespread adoption in the pharmaceutical industry13-15. Initial validation of this method in small-scale batch reactions is followed by adaptation to a recirculating flow reactor that enables hectogram-scale synthesis of a pharmaceutical intermediate. The mediated H2 anode technology disclosed here offers a general strategy to support H2-driven electrosynthetic reductions.

Shaping subcellular tubes through vesicle trafficking: Common and distinct pathways.

Cells with subcellular lumens form some of the most miniature tubes in the tubular organs of animals. These are often crucial components of the system, executing functions at remote body locations. Unlike tubes formed by intercellular or autocellular junctions, the cells with junctionless subcellular lumens face unique challenges in modifying the cell shape and plasma membrane organization to incorporate a membrane-bound tube within, often associated with dramatic cellular growth and extensions. Results in the recent years have shown that membrane dynamics, including both the primary delivery and recycling, is crucial in providing the cell with the flexibility to face these challenges. A significant portion of this information has come from two in vivo invertebrate models; the Drosophila tracheal terminal cells and the C. elegans excretory cell. This review focuses on the data obtained from these systems in the recent past about how trafficking pathways influence subcellular tube and branching morphogenesis. Given that such tubes occur in vertebrate vasculature, these insights are relevant to human health, and we contrast our conclusions with the less understood subcellular tubes of angiogenesis.

Heterogeneous M-N-C Catalysts for Aerobic Oxidation Reactions: Lessons from Oxygen Reduction Electrocatalysts.

Nonprecious metal heterogeneous catalysts composed of first-row transition metals incorporated into nitrogen-doped carbon matrices (M-N-Cs) have been studied for decades as leading alternatives to Pt for the electrocatalytic O2 reduction reaction (ORR). More recently, similar M-N-C catalysts have been shown to catalyze the aerobic oxidation of organic molecules. This Focus Review highlights mechanistic similarities and distinctions between these two reaction classes and then surveys the aerobic oxidation reactions catalyzed by M-N-Cs. As the active-site structures and kinetic properties of M-N-C aerobic oxidation catalysts have not been extensively studied, the array of tools and methods used to characterize ORR catalysts are presented with the goal of supporting further advances in the field of aerobic oxidation.

Inhibition of FOXP3 by stapled alpha-helical peptides dampens regulatory T cell function.

Despite continuing advances in the development of novel cellular-, antibody-, and chemotherapeutic-based strategies to enhance immune reactivity, the presence of regulatory T cells (Treg cells) remains a complicating factor for their clinical efficacy. To overcome dosing limitations and off-target effects from antibody-based Treg cell deletional strategies or small molecule drugging, we investigated the ability of hydrocarbon stapled alpha-helical (SAH) peptides to target FOXP3, the master transcription factor regulator of Treg cell development, maintenance, and suppressive function. Using the crystal structure of the FOXP3 homodimer as a guide, we developed SAHs in the likeness of a portion of the native FOXP3 antiparallel coiled-coil homodimerization domain (SAH-FOXP3) to block this key FOXP3 protein-protein interaction (PPI) through molecular mimicry. We describe the design, synthesis, and biochemical evaluation of single- and double-stapled SAHs covering the entire coiled-coil expanse. We show that lead SAH-FOXP3s bind FOXP3, are cell permeable and nontoxic to T cells, induce dose-dependent transcript and protein level alterations of FOXP3 target genes, impede Treg cell function, and lead to Treg cell gene expression changes in vivo consistent with FOXP3 dysfunction. These results demonstrate a proof of concept for rationally designed FOXP3-directed peptide therapeutics that could be used as approaches to amplify endogenous immune responsiveness.

Global Proteomic Analysis Reveals Alterations in Differentially Expressed Proteins between Cardiopathic Lamin A/C Mutations.

Lamin A/C (LMNA) is an important component of nuclear lamina. Mutations cause arrhythmia, heart failure, and sudden cardiac death. While LMNA-associated cardiomyopathy typically has an aggressive course that responds poorly to conventional heart failure therapies, there is variability in severity and age of penetrance between and even within specific mutations, which is poorly understood at the cellular level. Further, this heterogeneity has not previously been captured to mimic the heterozygous state, nor have the hundreds of clinical LMNA mutations been represented. Herein, we have overexpressed cardiopathic LMNA variants in HEK cells and utilized state-of-the-art quantitative proteomics to compare the global proteomic profiles of (1) aggregating Q353 K alone, (2) Q353 K coexpressed with WT, (3) aggregating N195 K coexpressed with WT, and (4) nonaggregating E317 K coexpressed with WT to help capture some of the heterogeneity between mutations. We analyzed each data set to obtain the differentially expressed proteins (DEPs) and applied gene ontology (GO) and KEGG pathway analyses. We found a range of 162 to 324 DEPs from over 6000 total protein IDs with differences in GO terms, KEGG pathways, and DEPs important in cardiac function, further highlighting the complexity of cardiac laminopathies. Pathways disrupted by LMNA mutations were validated with redox, autophagy, and apoptosis functional assays in both HEK 293 cells and in induced pluripotent stem cell derived cardiomyocytes (iPSC-CMs) for LMNA N195 K. These proteomic profiles expand our repertoire for mutation-specific downstream cellular effects that may become useful as druggable targets for personalized medicine approach for cardiac laminopathies.

Visualization of Cerebral Pressure Autoregulatory Insults in Traumatic Brain Injury.

OBJECTIVES: The first aim was to investigate the combined effect of insult intensity and duration of the pressure reactivity index (PRx) and deviation from the autoregulatory cerebral perfusion pressure target (∆CPPopt = actual CPP - optimal CPP [CPPopt]) on outcome in traumatic brain injury. The second aim was to determine if PRx influenced the association between intracranial pressure (ICP), CPP, and ∆CPPopt with outcome. DESIGN: Observational cohort study. SETTING: Neurocritical care unit, Cambridge, United Kingdom. PATIENTS: Five hundred fifty-three traumatic brain injury patients with ICP and arterial blood pressure monitoring and 6-month outcome data (Glasgow Outcome Scale [GOS]). INTERVENTION: None. MEASUREMENTS AND MAIN RESULTS: The insult intensity (mm Hg or PRx coefficient) and duration (minutes) of ICP, PRx, CPP, and ∆CPPopt were correlated with GOS and visualized in heatmaps. In these plots, there was a transition from favorable to unfavorable outcome when PRx remained positive for 30 minutes and this was also the case for shorter durations when the intensity was higher. In a similar plot of ∆CPPopt, there was a gradual transition from favorable to unfavorable outcome when ∆CPPopt went below -5 mm Hg for 30-minute episodes of time and for shorter durations for more negative ∆CPPopt. Furthermore, the percentage of monitoring time with certain combinations of PRx with ICP, CPP, and ∆CPPopt were correlated with GOS and visualized in heatmaps. In the combined PRx/ICP heatmap, ICP above 20 mm Hg together with PRx above 0 correlated with unfavorable outcome. In a PRx/CPP heatmap, CPP below 70 mm Hg together with PRx above 0.2-0.4 correlated with unfavorable outcome. In the PRx-/∆CPPopt heatmap, ∆CPPopt below 0 together with PRx above 0.2-0.4 correlated with unfavorable outcome. CONCLUSIONS: Higher intensities for longer durations of positive PRx and negative ∆CPPopt correlated with worse outcome. Elevated ICP, low CPP, and negative ∆CPPopt were particularly associated with worse outcomes when the cerebral pressure autoregulation was concurrently impaired.

Metabologenomic characterization uncovers a clinically aggressive IDH mutant glioma subtype.

Mutations in the pivotal metabolic isocitrate dehydrogenase (IDH) enzymes are recognized to drive the molecular footprint of diffuse gliomas, and patients with IDH mutant gliomas have overall favorable outcomes compared to patients with IDH wild-type tumors. However, survival still varies widely among patients with IDH mutated tumors. Here, we aimed to characterize molecular signatures that explain the range of IDH mutant gliomas. By integrating matched epigenome-wide methylome, transcriptome, and global metabolome data in 154 patients with gliomas, we identified a group of IDH mutant gliomas with globally altered metabolism that resembled IDH wild-type tumors. IDH-mutant gliomas with altered metabolism have significantly shorter overall survival from their IDH mutant counterparts that is not fully accounted for by recognized molecular prognostic markers of CDKN2A/B loss and glioma CpG Island Methylator Phenotype (GCIMP) status. IDH-mutant tumors with dysregulated metabolism harbored distinct epigenetic alterations that converged to drive proliferative and stem-like transcriptional profiles, providing a window to target novel dependencies in gliomas.

Platelet Function is Independent of Sphingolipid Manipulation.

INTRODUCTION: Previous literature suggests that sphingolipids may impact systemic coagulation and platelet aggregation, thus modulating the risks of thrombotic events. The goal of this investigation was to evaluate the role of serum sphingolipids on intrinsic platelet function to assess whether pharmacologic manipulation of sphingolipid metabolites would impact platelet aggregability. METHODS: C57BL/6J mice were injected with either normal saline, 1 mg/kg FTY720 (synthetic sphingosine-1-phosphate [S1P] receptor analog), or 5 mg/kg SLM6031434 (sphingosine kinase two inhibitor). Mice were sacrificed at 6 h and whole blood (WB) was collected for impedance aggregometry assessing platelet responsiveness to arachidonic acid or adenosine diphosphate. Ex vivo studies utilized WB or platelet-rich plasma that was pretreated with S1P, FTY720, amitriptyline, or d-sphingosine then analyzed by aggregability and flow cytometry for platelet and platelet-derived microvesicle characteristics. RESULTS: FTY720 and SLM6031434 pretreated induced similar arachidonic acid and adenosine diphosphate-mediated platelet aggregation as controls. Ex vivo WB and platelet-rich plasma treatment with S1P, FTY720, amitriptyline and d-sphingosine did not impact platelet aggregation. The percentages of CD41+, CD62P+ and CD41+/ceramide+, CD62P+/ceramide + platelets, and platelet-derived microvesicle were not significantly different between amitriptyline-treated and normal saline-treated cohorts. CONCLUSIONS: Sphingolipid modulating agents, such as FTY720, SLM6031434, S1P, amitriptyline, ceramide, and d-sphingosine do not appear to independently impact platelet aggregation in murine models.

High-fat diet, microbiome-gut-brain axis signaling, and anxiety-like behavior in male rats.

Obesity, associated with the intake of a high-fat diet (HFD), and anxiety are common among those living in modern urban societies. Recent studies suggest a role of microbiome-gut-brain axis signaling, including a role for brain serotonergic systems in the relationship between HFD and anxiety. Evidence suggests the gut microbiome and the serotonergic brain system together may play an important role in this response. Here we conducted a nine-week HFD protocol in male rats, followed by an analysis of the gut microbiome diversity and community composition, brainstem serotonergic gene expression (tph2, htr1a, and slc6a4), and anxiety-related defensive behavioral responses. We show that HFD intake decreased alpha diversity and altered the community composition of the gut microbiome in association with obesity, increased brainstem tph2, htr1a and slc6a4 mRNA expression, including in the caudal part of the dorsomedial dorsal raphe nucleus (cDRD), a subregion previously associated with stress- and anxiety-related behavioral responses, and, finally, increased anxiety-related defensive behavioral responses. The HFD increased the Firmicutes/Bacteroidetes ratio relative to control diet, as well as higher relative abundances of Blautia, and decreases in Prevotella. We found that tph2, htr1a and slc6a4 mRNA expression were increased in subregions of the dorsal raphe nucleus in the HFD, relative to control diet. Specific bacterial taxa were associated with increased serotonergic gene expression in the cDRD. Thus, we propose that HFD-induced obesity is associated with altered microbiome-gut-serotonergic brain axis signaling, leading to increased anxiety-related defensive behavioral responses in rats.

The Effect of Sport Specialization on Injury Risk in NCAA Athletes: Results From the SAFE Consortium.

OBJECTIVE: Youth athletes are beginning to specialize in a single sport more often. Previous studies in sports medicine and orthopedics have shown an association between intensity of sport specialization and incidence of injuries. This study is the first of its kind to explore the effects of early sport specialization on injury risk through a multicenter framework with a concentration on NCAA athletics. DESIGN: Retrospective cohort study. SETTING: SAFE Consortium. PARTICIPANTS: A total of 211 collegiate athletes from the NCAA's 3 levels of competition: Division I, II, and III. Data were collected by the SAFE investigators. INTERVENTION: N/A. MAIN OUTCOME MEASURES: Participants completed a questionnaire about their demographics, sport participation, specialization status, physical injuries, recovery period, and treatment method. Specialization status was calculated with a previously published 3-point scale: low, moderate, and high. Injuries were categorized as upper extremity injuries (UEIs) and lower extremity injuries (LEIs). RESULTS: Highly specialized athletes were more likely to report UEIs and LEIs than low specialized athletes (P < 0.0001). Moderate specialization, in contrast to low specialization, was associated with a higher likelihood of LEIs (P = 0.03) but not UEIs (P = 0.052). Highly specialized athletes were more likely to report an injury of any kind. CONCLUSIONS: The SAFE investigators found high specialization was associated with a history of UEIs and LEIs. Return to play was longer for highly specialized athletes versus low specialized athletes (112 days and 85 days, respectively). Highly specialized athletes were more likely to be from Division I and to require surgery.

A Collaborative Classroom Investigation of the Evolution of SABATH Methyltransferase Substrate Preference Shifts over 120 My of Flowering Plant History.

Next-generation sequencing has resulted in an explosion of available data, much of which remains unstudied in terms of biochemical function; yet, experimental characterization of these sequences has the potential to provide unprecedented insight into the evolution of enzyme activity. One way to make inroads into the experimental study of the voluminous data available is to engage students by integrating teaching and research in a college classroom such that eventually hundreds or thousands of enzymes may be characterized. In this study, we capitalize on this potential to focus on SABATH methyltransferase enzymes that have been shown to methylate the important plant hormone, salicylic acid (SA), to form methyl salicylate. We analyze data from 76 enzymes of flowering plant species in 23 orders and 41 families to investigate how widely conserved substrate preference is for SA methyltransferase orthologs. We find a high degree of conservation of substrate preference for SA over the structurally similar metabolite, benzoic acid, with recent switches that appear to be associated with gene duplication and at least three cases of functional compensation by paralogous enzymes. The presence of Met in active site position 150 is a useful predictor of SA methylation preference in SABATH methyltransferases but enzymes with other residues in the homologous position show the same substrate preference. Although our dense and systematic sampling of SABATH enzymes across angiosperms has revealed novel insights, this is merely the "tip of the iceberg" since thousands of sequences remain uncharacterized in this enzyme family alone.

Five-Year Outcomes of Transcatheter or Surgical Aortic-Valve Replacement.

BACKGROUND: There are scant data on long-term clinical outcomes and bioprosthetic-valve function after transcatheter aortic-valve replacement (TAVR) as compared with surgical aortic-valve replacement in patients with severe aortic stenosis and intermediate surgical risk. METHODS: We enrolled 2032 intermediate-risk patients with severe, symptomatic aortic stenosis at 57 centers. Patients were stratified according to intended transfemoral or transthoracic access (76.3% and 23.7%, respectively) and were randomly assigned to undergo either TAVR or surgical replacement. Clinical, echocardiographic, and health-status outcomes were followed for 5 years. The primary end point was death from any cause or disabling stroke. RESULTS: At 5 years, there was no significant difference in the incidence of death from any cause or disabling stroke between the TAVR group and the surgery group (47.9% and 43.4%, respectively; hazard ratio, 1.09; 95% confidence interval [CI], 0.95 to 1.25; P = 0.21). Results were similar for the transfemoral-access cohort (44.5% and 42.0%, respectively; hazard ratio, 1.02; 95% CI, 0.87 to 1.20), but the incidence of death or disabling stroke was higher after TAVR than after surgery in the transthoracic-access cohort (59.3% vs. 48.3%; hazard ratio, 1.32; 95% CI, 1.02 to 1.71). At 5 years, more patients in the TAVR group than in the surgery group had at least mild paravalvular aortic regurgitation (33.3% vs. 6.3%). Repeat hospitalizations were more frequent after TAVR than after surgery (33.3% vs. 25.2%), as were aortic-valve reinterventions (3.2% vs. 0.8%). Improvement in health status at 5 years was similar for TAVR and surgery. CONCLUSIONS: Among patients with aortic stenosis who were at intermediate surgical risk, there was no significant difference in the incidence of death or disabling stroke at 5 years after TAVR as compared with surgical aortic-valve replacement. (Funded by Edwards Lifesciences; PARTNER 2 ClinicalTrials.gov number, NCT01314313.).

Changes in transmural mass transport correlate with ascending thoracic aortic aneurysm diameter in a fibulin-4 E57K knockin mouse model.

Thoracic aortic aneurysm is characterized by dilation of the aortic diameter by greater than 50%, which can lead to dissection or rupture. Common histopathology includes extracellular matrix remodeling that may affect transmural mass transport, defined as the movement of fluids and solutes across the wall. We measured in vitro ascending thoracic aorta mass transport in a mouse model with partial aneurysm phenotype penetration due to a mutation in the extracellular matrix protein fibulin-4 [Fbln4E57K/E57K, referred to as MU-A (aneurysm) or MU-NA (no aneurysm)]. To push the aneurysm phenotype, we also included MU mice with reduced levels of lysyl oxidase [Fbln4E57K/E57K;Lox+/-, referred to as MU-XA (extreme aneurysm)] and compared all groups to wild-type (WT) littermates. The phenotype variation allows investigation of how aneurysm severity correlates with mass transport parameters and extracellular matrix organization. We found that MU-NA ascending thoracic aortae have similar hydraulic conductance (Lp) to WT, but 397% higher solute permeability (ω) for 4 kDa FITC-dextran. In contrast, MU-A and MU-XA ascending thoracic aortae have 44-68% lower Lp and similar ω to WT. The results suggest that ascending thoracic aortic aneurysm progression involves an initial increase in ω, followed by a decrease in Lp after the aneurysm has formed. All MU ascending thoracic aortae are longer and have increased elastic fiber fragmentation in the extracellular matrix. There is a negative correlation between diameter and Lp or ω in MU ascending thoracic aortae. Changes in mass transport due to elastic fiber fragmentation could contribute to aneurysm progression or be leveraged for treatment.NEW & NOTEWORTHY Transmural mass transport is quantified in the ascending thoracic aorta of mice with a mutation in fibulin-4 that is associated with thoracic aortic aneurysms. Fluid and solute transport depend on aneurysm severity, correlate with elastic fiber fragmentation, and may be affected by proteoglycan deposition. Transport properties of the ascending thoracic aorta are provided and can be used in computational models. The changes in mass transport may contribute to aneurysm progression or be leveraged for aneurysm treatment.

Supplemental findings of the 2021 National Blood Collection and Utilization Survey.

BACKGROUND: The Department of Health and Human Services' National Blood Collection and Utilization Survey (NBCUS) has been conducted biennially since 1997. Data are used to estimate national blood collection and use. Supplemental data from the 2021 NBCUS not presented elsewhere are presented here. METHODS: Data on survey participation, donor characteristics, blood component cost, transfusion-associated adverse reactions, and implementation of blood safety measures, including pathogen-reduction of platelets, during 2021, were analyzed. Comparisons are made to 2019 survey data where available (2013-2019 for survey participation). RESULTS: During 2021, there were 11,507,000 successful blood donations in the United States, a 4.8% increase from 2019. Persons aged 45-64 years accounted for 42% of all successful blood donations. Donations by persons aged 65 years and older increased by 40.7%, while donations among minorities and donors aged <25 years decreased. From 2019 to 2021, the median price hospitals paid per unit of leukoreduced red blood cells, leukoreduced and pathogen-reduced apheresis platelets, and fresh frozen plasma increased. The largest increase in price per unit of blood component in 2021 was for leukoreduced apheresis platelets, which increased by ~$51. Between 2019 and 2021, the proportion of transfusing facilities reporting use of pathogen-reduced platelets increased, from 13% to 60%. Transfusion-related adverse reactions declined slightly between 2019 and 2021, although the rate of transfusion-transmitted bacterial infections remained unchanged. CONCLUSION: During 2021, blood donations increased nationally, although donations from those aged <25 years and minorities declined. The prices hospitals paid for most blood products increased, as did the use of pathogen-reduced platelets.

Continued stabilization of blood collections and transfusions in the United States: Findings from the 2021 National Blood Collection and Utilization Survey.

BACKGROUND: National Blood Collection and Utilization Surveys (NBCUS) have reported decreases in U.S. blood collections and transfusions since 2008. The declines began to stabilize in 2015-2017, with a subsequent increase in transfusions in 2019. Data from the 2021 NBCUS were analyzed to understand the current dynamics of blood collection and use in the United States. METHODS: In March 2022, all community-based (53) and hospital-based (83) blood collection centers, a randomly selected 40% of transfusing hospitals performing 100-999 annual inpatient surgeries, and all transfusing hospitals performing ≥1000 annual inpatient surgeries were sent a 2021 NBCUS survey to ascertain blood collection and transfusion data. Responses were compiled, and national estimates were calculated for the number of units of blood and blood components collected, distributed, transfused, and outdated in 2021. Weighting and imputation were applied to account for non-responses and missing data, respectively. RESULTS: Survey response rates were 92.5% (49/53) for community-based blood centers, 74.7% (62/83) for hospital-based blood centers, and 76.3% (2102/2754) for transfusing hospitals. Overall, 11,784,000 (95% confidence interval [CI], 11,392,000-12,177,000) whole blood and apheresis red blood cell (RBC) units were collected in 2021, a 1.7% increase from 2019; 10,764,000 (95% CI, 10,357,000-11,171,000) whole blood-derived and apheresis RBC units were transfused, a 0.8% decrease. Total platelet units distributed increased by 0.8%; platelet units transfused decreased by 3.0%; plasma units distributed increased by 16.2%; and plasma units transfused increased by 1.4%. DISCUSSION: The 2021 NBCUS findings demonstrate a stabilization in U.S. blood collections and transfusions, suggesting a plateau has been reached for both.

Impact of the COVID-19 pandemic on blood donation and transfusions in the United States in 2020.

INTRODUCTION: Reports have suggested the COVID-19 pandemic resulted in blood donation shortages and adverse impacts on the blood supply. Using data from the National Blood Collection and Utilization Survey (NBCUS), we quantified the pandemic's impact on red blood cell (RBC) and apheresis platelet collections and transfusions in the United States during year 2020. METHODS: The 2021 NBCUS survey instrument was modified to include certain blood collection and utilization variables for 2020. The survey was distributed to all US blood collection centers, all US hospitals performing ≥1000 surgeries annually, and a 40% random sample of hospitals performing 100-999 surgeries annually. Weighting and imputation were used to generate national estimates for whole blood and apheresis platelet donation; RBC and platelet transfusion; and convalescent plasma distribution. RESULTS: Whole blood collections were stable from 2019 (9,790,000 units; 95% CI: 9,320,000-10,261,000) to 2020 (9,738,000 units; 95% CI: 9,365,000-10,110,000). RBC transfusions decreased by 6.0%, from 10,852,000 units (95% CI: 10,444,000-11,259,000) in 2019 to 10,202,000 units (95% CI: 9,811,000-10,593,000) in 2020. Declines were steepest during March-April 2020, with transfusions subsequently rebounding. Apheresis platelet collections increased from 2,359,000 units (95% CI: 2,240,000-2,477,000) in 2019 to 2,408,000 units (95% CI: 2,288,000-2,528,000) in 2020. Apheresis platelet transfusions increased from 1,996,000 units (95% CI: 1,846,000-2,147,000) in 2019 to 2,057,000 units (95% CI: 1,902,000-2,211,000) in 2020. CONCLUSION: The COVID-19 pandemic resulted in reduced blood donations and transfusions in some months during 2020 but only a minimal annualized decline compared with 2019.

Brain tissue oxygen monitoring in traumatic brain injury-part II: isolated and combined insults in relation to outcome.

BACKGROUND: The primary aim was to explore the concept of isolated and combined threshold-insults for brain tissue oxygenation (pbtO2) in relation to outcome in traumatic brain injury (TBI). METHODS: A total of 239 TBI patients with data on clinical outcome (GOS) and intracranial pressure (ICP) and pbtO2 monitoring for at least 12 h, who had been treated at the neurocritical care unit, Addenbrooke's Hospital, Cambridge, UK, between 2002 and 2022 were included. Outcome was dichotomised into favourable/unfavourable (GOS 4-5/1-3) and survival/mortality (GOS 2-5/1). PbtO2 was studied over the entire monitoring period. Thresholds were analysed in relation to outcome based on median and mean values, percentage of time and dose per hour below critical values and visualised as the combined insult intensity and duration. RESULTS: Median pbtO2 was slightly, but not significantly, associated with outcome. A pbtO2 threshold at 25 and 20 mmHg, respectively, yielded the highest x2 when dichotomised for favourable/unfavourable outcome and mortality/survival in chi-square analyses. A higher dose and higher percentage of time spent with pbtO2 below 25 mmHg as well as lower thresholds were associated with unfavourable outcome, but not mortality. In a combined insult intensity and duration analysis, there was a transition from favourable towards unfavourable outcome when pbtO2 went below 25-30 mmHg for 30 min and similar transitions occurred for shorter durations when the intensity was higher. Although these insults were rare, pbtO2 under 15 mmHg was more strongly associated with unfavourable outcome if, concurrently, ICP was above 20 mmHg, cerebral perfusion pressure below 60 mmHg, or pressure reactivity index above 0.30 than if these variables were not deranged. In a multiple logistic regression, a higher percentage of monitoring time with pbtO2 < 15 mmHg was associated with a higher rate of unfavourable outcome. CONCLUSIONS: Low pbtO2, under 25 mmHg and particularly below 15 mmHg, for longer durations and in combination with disturbances in global cerebral physiological variables were associated with poor outcome and may indicate detrimental ischaemic hypoxia. Prospective trials are needed to determine if pbtO2-directed therapy is beneficial, at what individualised pbtO2 threshold therapies are warranted, and how this may depend on the presence/absence of concurrent cerebral physiological disturbances.

Brain tissue oxygen monitoring in traumatic brain injury: part I-To what extent does PbtO2 reflect global cerebral physiology?

BACKGROUND: The primary aim was to explore the association of global cerebral physiological variables including intracranial pressure (ICP), cerebrovascular reactivity (PRx), cerebral perfusion pressure (CPP), and deviation from the PRx-based optimal CPP value (∆CPPopt; actual CPP-CPPopt) in relation to brain tissue oxygenation (pbtO2) in traumatic brain injury (TBI). METHODS: A total of 425 TBI patients with ICP- and pbtO2 monitoring for at least 12 h, who had been treated at the neurocritical care unit, Addenbrooke's Hospital, Cambridge, UK, between 2002 and 2022 were included. Generalized additive models (GAMs) and linear mixed effect models were used to explore the association of ICP, PRx, CPP, and CPPopt in relation to pbtO2. PbtO2 < 20 mmHg, ICP > 20 mmHg, PRx > 0.30, CPP < 60 mmHg, and ∆CPPopt < - 5 mmHg were considered as cerebral insults. RESULTS: PbtO2 < 20 mmHg occurred in median during 17% of the monitoring time and in less than 5% in combination with ICP > 20 mmHg, PRx > 0.30, CPP < 60 mmHg, or ∆CPPopt < - 5 mmHg. In GAM analyses, pbtO2 remained around 25 mmHg over a large range of ICP ([0;50] mmHg) and PRx [- 1;1], but deteriorated below 20 mmHg for extremely low CPP below 30 mmHg and ∆CPPopt below - 30 mmHg. In linear mixed effect models, ICP, CPP, PRx, and ∆CPPopt were significantly associated with pbtO2, but the fixed effects could only explain a very small extent of the pbtO2 variation. CONCLUSIONS: PbtO2 below 20 mmHg was relatively frequent and often occurred in the absence of disturbances in ICP, PRx, CPP, and ∆CPPopt. There were significant, but weak associations between the global cerebral physiological variables and pbtO2, suggesting that hypoxic pbtO2 is often a complex and independent pathophysiological event. Thus, other variables may be more crucial to explain pbtO2 and, likewise, pbtO2 may not be a suitable outcome measure to determine whether global cerebral blood flow optimization such as CPPopt therapy is successful.

Methylation Profiling Identifies Stability of Isocitrate Dehydrogenase Mutation Over Time.

OBJECTIVE: Isocitrate dehydrogenase (IDH) mutation status is a key diagnostic and prognostic feature of gliomas. It is thought to occur early in glioma tumorigenesis and remain stable over time. However, there are reports documenting a loss of IDH mutation status in a subset of patients with glioma recurrence. Here, we identified patients with a documented loss of IDH mutation status longitudinally and performed multi-platform analysis in order to determine if IDH mutations are stable throughout glioma evolution. METHODS: We retrospectively identified patients from our institution from 2009 to 2018 with immunohistochemistry (IHC)-recorded IDH mutation status changes longitudinally. Archived formalin-fixed paraffin-embedded and frozen tissue samples from these patients were collected from our institution's tumour bank. Samples were analysed using methylation profiling, copy number variation, Sanger sequencing, droplet digital PCR (ddPCR) and IHC. RESULTS: We reviewed 1491 archived glioma samples including 78 patients with multiple IDH mutant tumour samples collected longitudinally. In all instances of documented loss of IDH mutation status, multi-platform profiling identified a mixture of low tumour cell content and non-neoplastic tissue including perilesional, reactive or inflammatory cells. CONCLUSIONS: All patients with a documented loss of IDH mutation status longitudinally were resolved through multi-platform analysis. These findings support the hypothesis that IDH mutations occur early in gliomagenesis and in the absence of copy number changes at the IDH loci and are stable throughout tumour treatment and evolution. Our study highlights the importance of accurate surgical sampling and the role of DNA methylome profiling in diagnostically uncertain cases for integrated pathological and molecular diagnosis.

Phytophthora : taxonomic and phylogenetic revision of the genus.

Many members of the Oomycota genus Phytophthora cause economic and environmental impact diseases in nurseries, horticulture, forest, and natural ecosystems and many are of regulatory concern around the world. At present, there are 223 described species, including eight unculturable and three lost species. Twenty-eight species need to be redescribed or validated. A lectotype, epitype or neotype was selected for 20 species, and a redescription based on the morphological/molecular characters and phylogenetic placement is provided. In addition, the names of five species are validated: P. cajani, P. honggalleglyana (Synonym: P. hydropathica), P. megakarya, P. pisi and P. pseudopolonica for which morphology and phylogeny are given. Two species, P. ×multiformis and P. uniformis are presented as new combinations. Phytophthora palmivora is treated with a representative strain as both lecto- and epitypification are pending. This manuscript provides the updated multigene phylogeny and molecular toolbox with seven genes (ITS rDNA, ß-tub, COI, EF1α, HSP90, L10, and YPT1) generated from the type specimens of 212 validly published, and culturable species (including nine hybrid taxa). The genome information of 23 types published to date is also included. Several aspects of the taxonomic revision and phylogenetic re-evaluation of the genus including species concepts, concept and position of the phylogenetic clades recognized within Phytophthora are discussed. Some of the contents of this manuscript, including factsheets for the 212 species, are associated with the "IDphy: molecular and morphological identification of Phytophthora based on the types" online resource (https://idtools.org/tools/1056/index.cfm). The first version of the IDphy online resource released to the public in September 2019 contained 161 species. In conjunction with this publication, we are updating the IDphy online resource to version 2 to include the 51 species recently described. The current status of the 223 described species is provided along with information on type specimens with details of the host (substrate), location, year of collection and publications. Additional information is provided regarding the ex-type culture(s) for the 212 valid culturable species and the diagnostic molecular toolbox with seven genes that includes the two metabarcoding genes (ITS and COI) that are important for Sanger sequencing and also very valuable Molecular Operational Taxonomic Units (MOTU) for second and third generation metabarcoding High-throughput sequencing (HTS) technologies. The IDphy online resource will continue to be updated annually to include new descriptions. This manuscript in conjunction with IDphy represents a monographic study and the most updated revision of the taxonomy and phylogeny of Phytophthora, widely considered one of the most important genera of plant pathogens. Taxonomic novelties: New species: Phytophthora cajani K.S. Amin, Baldev & F.J. Williams ex Abad, Phytophthora honggalleglyana Abad, Phytophthora megakarya Brasier & M.J. Griffin ex Abad, Phytophthora pisi Heyman ex Abad, Phytophthora pseudopolonica W.W. Li, W.X. Huai & W.X. Zhao ex Abad & Kasiborski; New combinations: Phytophthora ×multiformis (Brasier & S.A. Kirk) Abad, Phytophthora uniformis (Brasier & S.A. Kirk) Abad; Epitypifications (basionyms): Peronospora cactorum Lebert & Cohn, Pythiacystis citrophthora R.E. Sm. & E.H. Sm., Phytophthora colocasiae Racib., Phytophthora drechsleri Tucker, Phytophthora erythroseptica Pethybr., Phytophthora fragariae Hickman, Phytophthora hibernalis Carne, Phytophthora ilicis Buddenh. & Roy A. Young, Phytophthora inundata Brasier et al., Phytophthora megasperma Drechsler, Phytophthora mexicana Hotson & Hartge, Phytophthora nicotianae Breda de Haan, Phytophthora phaseoli Thaxt., Phytophthora porri Foister, Phytophthora primulae J.A. Toml., Phytophthora sojae Kaufm. & Gerd., Phytophthora vignae Purss, Pythiomorpha gonapodyides H.E. Petersen; Lectotypifications (basionym): Peronospora cactorum Lebert & Cohn, Pythiacystis citrophthora R.E. Sm. & E.H. Sm., Phytophthora colocasiae Racib., Phytophthora drechsleri Tucker, Phytophthora erythroseptica Pethybr., Phytophthora fragariae Hickman, Phytophthora hibernalis Carne, Phytophthora ilicis Buddenh. & Roy A. Young, Phytophthora megasperma Drechsler, Phytophthora mexicana Hotson & Hartge, Phytophthora nicotianae Breda de Haan, Phytophthora phaseoli Thaxt., Phytophthora porri Foister, Phytophthora primulae J.A. Toml., Phytophthora sojae Kaufm. & Gerd., Phytophthora vignae Purss, Pythiomorpha gonapodyides H.E. Petersen; Neotypifications (basionym): Phloeophthora syringae Kleb., Phytophthora meadii McRae Citation: Abad ZG, Burgess TI, Bourret T, Bensch K, Cacciola S, Scanu B, Mathew R, Kasiborski B, Srivastava S, Kageyama K, Bienapfl JC, Verkleij G, Broders K, Schena L, Redford AJ (2023). Phytophthora: taxonomic and phylogenetic revision of the genus. Studies in Mycology 106: 259-348. doi: 10.3114/sim.2023.106.05.