Conformational surveillance of Orai1 by a rhomboid intramembrane protease prevents inappropriate CRAC channel activation.

Calcium influx through plasma membrane calcium release-activated calcium (CRAC) channels, which are formed of hexamers of Orai1, is a potent trigger for many important biological processes, most notably in T cell-mediated immunity. Through a bioinformatics-led cell biological screen, we have identified Orai1 as a substrate for the rhomboid intramembrane protease RHBDL2. We show that RHBDL2 prevents stochastic calcium signaling in unstimulated cells through conformational surveillance and cleavage of inappropriately activated Orai1. A conserved disease-linked proline residue is responsible for RHBDL2's recognizing the active conformation of Orai1, which is required to sharpen switch-like signaling triggered by store-operated calcium entry. Loss of RHBDL2 control of CRAC channel activity causes severe dysregulation of downstream CRAC channel effectors, including transcription factor activation, inflammatory cytokine expression, and T cell activation. We propose that this surveillance function may represent an ancient activity of rhomboid proteases in degrading unwanted signaling proteins.

Investigating the potential of single-cell DNA methylation data to detect allele-specific methylation and imprinting.

Allele-specific methylation (ASM) is an epigenetic modification whereby one parental allele becomes methylated and the other unmethylated at a specific locus. ASM is most often driven by the presence of nearby heterozygous variants that influence methylation, but also occurs somatically in the context of genomic imprinting. In this study, we investigate ASM using publicly available single-cell reduced representation bisulfite sequencing (scRRBS) data on 608 B cells sampled from six healthy B cell samples and 1,230 cells from 11 chronic lymphocytic leukemia (CLL) samples. We developed a likelihood-based criterion to test whether a CpG exhibited ASM, based on the distributions of methylated and unmethylated reads both within and across cells. Applying our likelihood ratio test, 65,998 CpG sites exhibited ASM in healthy B cell samples according to a Bonferroni criterion (p < 8.4 × 10-9), and 32,862 CpG sites exhibited ASM in CLL samples (p < 8.5 × 10-9). We also called ASM at the sample level. To evaluate the accuracy of our method, we called heterozygous variants from the scRRBS data, which enabled variant-based calls of ASM within each cell. Comparing sample-level ASM calls to the variant-based measures of ASM, we observed a positive predictive value of 76%-100% across samples. We observed high concordance of ASM across samples and an overrepresentation of ASM in previously reported imprinted genes and genes with imprinting binding motifs. Our study demonstrates that single-cell bisulfite sequencing is a potentially powerful tool to investigate ASM, especially as studies expand to increase the number of samples and cells sequenced.

Cardiac Catheterization Laboratory Management of the Comatose Adult Patient With an Out-of-Hospital Cardiac Arrest: A Scientific Statement From the American Heart Association.

Out-of-hospital cardiac arrest is a leading cause of death, accounting for ≈50% of all cardiovascular deaths. The prognosis of such individuals is poor, with <10% surviving to hospital discharge. Survival with a favorable neurologic outcome is highest among individuals who present with a witnessed shockable rhythm, received bystander cardiopulmonary resuscitation, achieve return of spontaneous circulation within 15 minutes of arrest, and have evidence of ST-segment elevation on initial ECG after return of spontaneous circulation. The cardiac catheterization laboratory plays an important role in the coordinated Chain of Survival for patients with out-of-hospital cardiac arrest. The catheterization laboratory can be used to provide diagnostic, therapeutic, and resuscitative support after sudden cardiac arrest from many different cardiac causes, but it has a unique importance in the treatment of cardiac arrest resulting from underlying coronary artery disease. Over the past few years, numerous trials have clarified the role of the cardiac catheterization laboratory in the management of resuscitated patients or those with ongoing cardiac arrest. This scientific statement provides an update on the contemporary approach to managing resuscitated patients or those with ongoing cardiac arrest.

Dynamics and variability of transcriptomic dysregulation in congenital myotonic dystrophy during pediatric development.

Myotonic dystrophy type 1 (DM1) is a multi-systemic disorder caused by expansion of CTG microsatellite repeats within DMPK. The most severe form, congenital myotonic dystrophy (CDM), has symptom onset at birth due to large intergenerational repeat expansions. Despite a common mutation, CDM individuals present with a distinct clinical phenotype and absence of common DM1 symptoms. Given the clinical divergence, it is unknown if the hallmark of DM1 pathology, dysregulation of alternative splicing (AS) due to sequestration of MBNL proteins within toxic CUG repeat RNAs, contributes to disease throughout pediatric development. To evaluate global transcriptomic dysregulation, RNA-seq was performed on 36 CDM skeletal muscle biopsies ages 2 weeks to 16 years, including two longitudinal samples. Fifty DM1 and adult/pediatric controls were also sequenced as comparative groups. Despite a large CTG expansion and shared age of onset, CDM individuals presented with a heterogenous, MBNL-dependent mis-splicing signature. Estimation of intracellular MBNL concentrations from splicing responses of select events correlated with total spliceopathy and revealed a distinct, triphasic pattern of AS dysregulation across pediatric development. CDM infants (< 2 years) possess severe mis-splicing that significantly improves in early childhood (2-8 years) independent of sex or CTG repeat load. Adolescent individuals (8-16 years) stratified into two populations with a full range of global splicing dysregulation. DMPK expression changes correlated with alterations in splicing severity during development. This study reveals the complex dynamics of the CDM muscle transcriptome and provides insights into new therapeutic strategies, timing of therapeutic intervention, and biomarker development.

Vaccine Effectiveness Against Influenza A-Associated Hospitalization, Organ Failure, and Death: United States, 2022-2023.

BACKGROUND: Influenza circulation during the 2022-2023 season in the United States largely returned to pre-coronavirus disease 2019 (COVID-19)-pandemic patterns and levels. Influenza A(H3N2) viruses were detected most frequently this season, predominately clade 3C.2a1b.2a, a close antigenic match to the vaccine strain. METHODS: To understand effectiveness of the 2022-2023 influenza vaccine against influenza-associated hospitalization, organ failure, and death, a multicenter sentinel surveillance network in the United States prospectively enrolled adults hospitalized with acute respiratory illness between 1 October 2022, and 28 February 2023. Using the test-negative design, vaccine effectiveness (VE) estimates against influenza-associated hospitalization, organ failures, and death were measured by comparing the odds of current-season influenza vaccination in influenza-positive case-patients and influenza-negative, SARS-CoV-2-negative control-patients. RESULTS: A total of 3707 patients, including 714 influenza cases (33% vaccinated) and 2993 influenza- and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-negative controls (49% vaccinated) were analyzed. VE against influenza-associated hospitalization was 37% (95% confidence interval [CI]: 27%-46%) and varied by age (18-64 years: 47% [30%-60%]; ≥65 years: 28% [10%-43%]), and virus (A[H3N2]: 29% [6%-46%], A[H1N1]: 47% [23%-64%]). VE against more severe influenza-associated outcomes included: 41% (29%-50%) against influenza with hypoxemia treated with supplemental oxygen; 65% (56%-72%) against influenza with respiratory, cardiovascular, or renal failure treated with organ support; and 66% (40%-81%) against influenza with respiratory failure treated with invasive mechanical ventilation. CONCLUSIONS: During an early 2022-2023 influenza season with a well-matched influenza vaccine, vaccination was associated with reduced risk of influenza-associated hospitalization and organ failure.

Effectiveness of Updated 2023-2024 (Monovalent XBB.1.5) COVID-19 Vaccination Against SARS-CoV-2 Omicron XBB and BA.2.86/JN.1 Lineage Hospitalization and a Comparison of Clinical Severity-IVY Network, 26 Hospitals, October 18, 2023-March 9, 2024.

BACKGROUND: Assessing variant-specific COVID-19 vaccine effectiveness (VE) and severity can inform public health risk assessments and decisions about vaccine composition. BA.2.86 and its descendants, including JN.1 (referred to collectively as "JN lineages"), emerged in late 2023 and exhibited substantial divergence from co-circulating XBB lineages. METHODS: We analyzed patients hospitalized with COVID-19-like illness at 26 hospitals in 20 U.S. states admitted October 18, 2023-March 9, 2024. Using a test-negative, case-control design, we estimated effectiveness of an updated 2023-2024 (Monovalent XBB.1.5) COVID-19 vaccine dose against sequence-confirmed XBB and JN lineage hospitalization using logistic regression. Odds of severe outcomes, including intensive care unit (ICU) admission and invasive mechanical ventilation (IMV) or death, were compared for JN versus XBB lineage hospitalizations using logistic regression. RESULTS: 585 case-patients with XBB lineages, 397 case-patients with JN lineages, and 4,580 control-patients were included. VE in the first 7-89 days after receipt of an updated dose was 54.2% (95% CI = 36.1%-67.1%) against XBB lineage hospitalization and 32.7% (95% CI = 1.9%-53.8%) against JN lineage hospitalization. Odds of ICU admission (adjusted odds ratio [aOR] 0.80; 95% CI = 0.46-1.38) and IMV or death (aOR 0.69; 95% CI = 0.34-1.40) were not significantly different among JN compared to XBB lineage hospitalizations. CONCLUSIONS: Updated 2023-2024 COVID-19 vaccination provided protection against both XBB and JN lineage hospitalization, but protection against the latter may be attenuated by immune escape. Clinical severity of JN lineage hospitalizations was not higher relative to XBB.

Fatal West Nile Virus Infection in Horse Returning to United Kingdom from Spain, 2022.

We report fatal West Nile virus (WNV) infection in a 7-year-old mare returning to the United Kingdom from Spain. Case timeline and clustering of virus sequence with recent WNV isolates suggest that transmission occurred in Andalusía, Spain. Our findings highlight the importance of vaccination for horses traveling to WNV-endemic regions.

SARS-CoV-2 Spike Protein-Derived Cyclic Peptides as Modulators of Spike Interaction with GRP78.

The human glucose-regulated protein GRP78 is a human chaperone that translocactes to the cell surface when cells are under stress. Theoretical studies suggested it could be involved in SARS-CoV-2 virus entry to cells. In this work, we used in vitro surface plasmon resonance-based assays to show that human GRP78 indeed binds to SARS-CoV-2 spike protein. We have designed and synthesised cyclic peptides based on the loop structure of amino acids 480-488 of the SARS-CoV-2 spike protein S1 domain from the Wuhan and Omicron variants and showed that both peptides bind to GRP78. Consistent with the greater infectiousness of the Omicron variant, the Omicron-derived peptide displays slower dissociation from the target protein. Both peptides significantly inhibit the binding of wild-type S1 protein to the human protein GRP78 suggesting that further development of these cyclic peptide motifs may provide a viable route to novel anti-SARS-CoV-2 agents.

ROptimus: a parallel general-purpose adaptive optimization engine.

MOTIVATION: Various computational biology calculations require a probabilistic optimization protocol to determine the parameters that capture the system at a desired state in the configurational space. Many existing methods excel at certain scenarios, but fail in others due, in part, to an inefficient exploration of the parameter space and easy trapping into local minima. Here, we developed a general-purpose optimization engine in R that can be plugged to any, simple or complex, modelling initiative through a few lucid interfacing functions, to perform a seamless optimization with rigorous parameter sampling. RESULTS: ROptimus features simulated annealing and replica exchange implementations equipped with adaptive thermoregulation to drive Monte Carlo optimization process in a flexible manner, through constrained acceptance frequency but unconstrained adaptive pseudo temperature regimens. We exemplify the applicability of our R optimizer to a diverse set of problems spanning data analyses and computational biology tasks. AVAILABILITY AND IMPLEMENTATION: ROptimus is written and implemented in R, and is freely available from CRAN (http://cran.r-project.org/web/packages/ROptimus/index.html) and GitHub (http://github.com/SahakyanLab/ROptimus).

Male lake char release taurocholic acid as part of a mating pheromone.

The evolutionary origins of sexual preferences for chemical signals remain poorly understood, due, in part, to scant information on the molecules involved. In the current study, we identified a male pheromone in lake char (Salvelinus namaycush) to evaluate the hypothesis that it exploits a non-sexual preference for juvenile odour. In anadromous char species, the odour of stream-resident juveniles guides migratory adults into spawning streams. Lake char are also attracted to juvenile odour but have lost the anadromous phenotype and spawn on nearshore reefs, where juvenile odour does not persist long enough to act as a cue for spawning site selection by adults. Previous behavioural data raised the possibility that males release a pheromone that includes components of juvenile odour. Using metabolomics, we found that the most abundant molecule released by males was also released by juveniles but not females. Tandem mass spectrometry and nuclear magnetic resonance were used to identify the molecule as taurocholic acid (TCA), which was previously implicated as a component of juvenile odour. Additional chemical analyses revealed that males release TCA at high rates via their urine during the spawning season. Finally, picomolar concentrations of TCA attracted pre-spawning and spawning females but not males. Taken together, our results indicate that male lake char release TCA as a mating pheromone and support the hypothesis that the pheromone is a partial match of juvenile odour.

Temperature Control in Acute Brain Injury: An Update.

Temperature control in severe acute brain injury (SABI) is a key component of acute management. This manuscript delves into the complex role of temperature management in SABI, encompassing conditions like traumatic brain injury (TBI), acute ischemic stroke (AIS), intracerebral hemorrhage (ICH), aneurysmal subarachnoid hemorrhage (aSAH), and hypoxemic/ischemic brain injury following cardiac arrest. Fever is a common complication in SABI and is linked to worse neurological outcomes due to increased inflammatory responses and intracranial pressure (ICP). Temperature management, particularly hypothermic temperature control (HTC), appears to mitigate these adverse effects primarily by reducing cerebral metabolic demand and dampening inflammatory pathways. However, the effectiveness of HTC varies across different SABI conditions. In the context of post-cardiac arrest, the impact of HTC on neurological outcomes has shown inconsistent results. In cases of TBI, HTC seems promising for reducing ICP, but its influence on long-term outcomes remains uncertain. For AIS, clinical trials have yet to conclusively demonstrate the benefits of HTC, despite encouraging preclinical evidence. This variability in efficacy is also observed in ICH, aSAH, bacterial meningitis, and status epilepticus. In pediatric and neonatal populations, while HTC shows significant benefits in hypoxic-ischemic encephalopathy, its effectiveness in other brain injuries is mixed. Although the theoretical basis for employing temperature control, especially HTC, is strong, the clinical outcomes differ among various SABI subtypes. The current consensus indicates that fever prevention is beneficial across the board, but the application and effectiveness of HTC are more nuanced, underscoring the need for further research to establish optimal temperature management strategies. Here we provide an overview of the clinical evidence surrounding the use of temperature control in various types of SABI.

Defining clinical endpoints in limb girdle muscular dystrophy: a GRASP-LGMD study.

BACKGROUND: The Limb Girdle Muscular Dystrophies (LGMDs) are characterized by progressive weakness of the shoulder and hip girdle muscles as a result of over 30 different genetic mutations. This study is designed to develop clinical outcome assessments across the group of disorders. METHODS/DESIGN: The primary goal of this study is to evaluate the utility of a set of outcome measures on a wide range of LGMD phenotypes and ability levels to determine if it would be possible to use similar outcomes between individuals with different phenotypes. We will perform a multi-center, 12-month study of 188 LGMD patients within the established Genetic Resolution and Assessments Solving Phenotypes in LGMD (GRASP-LGMD) Research Consortium, which is comprised of 11 sites in the United States and 2 sites in Europe. Enrolled patients will be clinically affected and have mutations in CAPN3 (LGMDR1), ANO5 (LGMDR12), DYSF (LGMDR2), DNAJB6 (LGMDD1), SGCA (LGMDR3), SGCB (LGMDR4), SGCD (LGMDR6), or SGCG (LGMDR5, or FKRP-related (LGMDR9). DISCUSSION: To the best of our knowledge, this will be the largest consortium organized to prospectively validate clinical outcome assessments (COAs) in LGMD at its completion. These assessments will help clinical trial readiness by identifying reliable, valid, and responsive outcome measures as well as providing data driven clinical trial decision making for future clinical trials on therapeutic agents for LGMD. The results of this study will permit more efficient clinical trial design. All relevant data will be made available for investigators or companies involved in LGMD therapeutic development upon conclusion of this study as applicable. TRIAL REGISTRATION: Clinicaltrials.gov NCT03981289; Date of registration: 6/10/2019.

Implementation of Extracorporeal CPR Programs for Out-of-Hospital Cardiac Arrest: Another Tale of Two County Hospitals.

Extracorporeal cardiopulmonary resuscitation (ECPR) is a form of intensive life support that has seen increasing use globally to improve outcomes for patients who experience out-of-hospital cardiac arrest (OHCA). Hospitals with advanced critical care capabilities may be interested in launching an ECPR program to offer this support to the patients they serve; however, to do so, they must first consider the significant investment of resources necessary to start and sustain the program. The existing literature describes many single-center ECPR programs and often focuses on inpatient care and patient outcomes in hospitals with cardiac surgery capabilities. However, building a successful ECPR program and using this technology to support an individual patient experiencing refractory cardiac arrest secondary to a shockable rhythm depends on efficient out-of-hospital and emergency department (ED) management. This article describes the process of implementing 2 intensivist-led ECPR programs with limited cardiac surgery capability. We focus on emergency medical services and ED clinician roles in identifying patients, mobilizing resources, initiation and management of veno-arterial extracorporeal membrane oxygenation (VA-ECMO) in the ED, and ongoing efforts to improve ECPR program quality. Each center experienced a significant learning curve to reach goals of arrest-to-flow times of cannulation for ECPR. Building consensus from multidisciplinary stakeholders, including out-of-hospital stakeholders; establishing shared expectations of ECPR outcomes; and ensuring adequate resource support for ECPR activation were all key lessons in improving our ECPR programs.

Protection from UV light is an evolutionarily conserved feature of the haematopoietic niche.

Haematopoietic stem and progenitor cells (HSPCs) require a specific microenvironment, the haematopoietic niche, which regulates HSPC behaviour1,2. The location of this niche varies across species, but the evolutionary pressures that drive HSPCs to different microenvironments remain unknown. The niche is located in the bone marrow in adult mammals, whereas it is found in other locations in non-mammalian vertebrates, for example, in the kidney marrow in teleost fish. Here we show that a melanocyte umbrella above the kidney marrow protects HSPCs against ultraviolet light in zebrafish. Because mutants that lack melanocytes have normal steady-state haematopoiesis under standard laboratory conditions, we hypothesized that melanocytes above the stem cell niche protect HSPCs against ultraviolet-light-induced DNA damage. Indeed, after ultraviolet-light irradiation, unpigmented larvae show higher levels of DNA damage in HSPCs, as indicated by staining of cyclobutane pyrimidine dimers and have reduced numbers of HSPCs, as shown by cmyb (also known as myb) expression. The umbrella of melanocytes associated with the haematopoietic niche is highly evolutionarily conserved in aquatic animals, including the sea lamprey, a basal vertebrate. During the transition from an aquatic to a terrestrial environment, HSPCs relocated into the bone marrow, which is protected from ultraviolet light by the cortical bone around the marrow. Our studies reveal that melanocytes above the haematopoietic niche protect HSPCs from ultraviolet-light-induced DNA damage in aquatic vertebrates and suggest that during the transition to terrestrial life, ultraviolet light was an evolutionary pressure affecting the location of the haematopoietic niche.

Prehospital Administration of Norepinephrine and Epinephrine for Shock after Resuscitation from Cardiac Arrest.

INTRODUCTION: Shock after resuscitation from out-of-hospital cardiac arrest (OHCA) is often treated with vasopressors. We examined whether infusion of epinephrine versus norepinephrine was associated with prehospital rearrest and neurologically favorable survival among OHCA patients. METHODS: This retrospective study included OHCA cases in Seattle, Washington from 2014-2021 who had return of spontaneous circulation (ROSC) followed by vasopressor infusion. Our primary exposure was infusion of epinephrine or norepinephrine. Our primary outcome was prehospital rearrest. Secondary outcomes included survival and neurologically favorable outcome (Cerebral Performance Category score of 1 or 2). We used multivariable logistic regression to test associations between exposures and outcomes adjusting for key covariates. RESULTS: Of 451 OHCA patients with ROSC followed by vasopressor infusion, 253 (56%) received norepinephrine and 198 (44%) received epinephrine infusions. Those who received epinephrine were older (median 66 [interquartile ranges {IQR} 53-79] vs 63 [IQR 47-75] years), but otherwise had similar baseline characteristics. Patients who received epinephrine were twice as likely to rearrest (55% vs 25%). After adjustment, the difference in rearrest rates between epinephrine and norepinephrine persisted (OR 3.28, 95%CI 2.25-5.08), and the odds of pulses at hospital arrival were lower in the epinephrine group (OR 0.52 95%CI 0.32-0.83). After adjustment, there was no difference in neurologically favorable survival, survival to hospital admission, or survival to discharge. CONCLUSION: Patients who received epinephrine infusions after ROSC suffered prehospital rearrest more frequently than those who received norepinephrine. Survival and neurological status at hospital discharge were similar. Future trials should examine the optimal approach to hemodynamic management for post-OHCA shock.

Prehospital Ground and Helicopter-Based Extracorporeal Cardiopulmonary Resuscitation (ECPR) Reduce Barriers to ECPR: A GIS Model.

INTRODUCTION: Evidence suggests that Extracorporeal Cardiopulmonary Resuscitation (ECPR) can improve survival rates for nontraumatic out-of-hospital cardiac arrest (OHCA). However, when ECPR is indicated over 50% of potential candidates are unable to qualify in the current hospital-based system due to geographic limitations. This study employs a Geographic Information System (GIS) model to estimate the number of ECPR eligible patients within the United States in the current hospital-based system, a prehospital ECPR ground-based system, and a prehospital ECPR Helicopter Emergency Medical Services (HEMS)-based system. METHODS: We constructed a GIS model to estimate ground and helicopter transport times. Time-dependent rates of ECPR eligibility were derived from the Resuscitation Outcome Consortium (ROC) database, while the Cardiac Arrest Registry to Enhance Survival (CARES) registry determined the number of OHCA patients meeting ECPR criteria within designated transportation times. Emergency Medical Services (EMS) response time, ECPR candidacy determination time, and on-scene time were modeled based on data from the EROCA trial. The combined model was used to estimate the total ECPR eligibility in each system. RESULTS: The CARES registry recorded 736,066 OHCA patients from 2013 to 2021. After applying clinical criteria, 24,661 (3.4%) ECPR-indicated OHCA were identified. When considering overall ECPR eligibility within 45 min from OHCA to initiation, only 11.76% of OHCA where ECPR was indicated were eligible in the current hospital-based system. The prehospital ECPR HEMS-based system exhibited a four-fold increase in ECPR eligibility (49.3%), while the prehospital ground-based system showed a more than two-fold increase (28.4%). CONCLUSIONS: The study demonstrates a two-fold increase in ECPR eligibility for a prehospital ECPR ground-based system and a four-fold increase for a prehospital ECPR HEMS-based system compared to the current hospital-based ECPR system. This novel GIS model can inform future ECPR implementation strategies, optimizing systems of care.

Characteristics of victims of trauma requiring invasive mechanical ventilation with a short stay in critical care.

BACKGROUND: Many patients who are admitted to the intensive care unit (ICU) have needs which rapidly resolve and are discharged alive within 24 h. We sought to characterize the outcomes of critically ill trauma victims at our institution with a short stay in the ICU. METHODS: We conducted a retrospective cohort study of all critically ill adult trauma victims presenting to our ED between January 1st, 2011 and December 31st, 2019. We included patients who were endotracheally intubated in either the prehospital setting or the ED and were admitted either to the operating room (OR), angiography suite, or ICU. Our primary outcome was the proportion of patients who were discharged alive from the ICU within 24 h. RESULTS: We included 3869 patients meeting the criteria above who were alive at 24 h. This population was 78% male with a median age of 40 and 76% of patients suffered from blunt trauma. The median injury severity score (ISS) of the group was 21 [inter-quartile range (IQR) 11-30]. In-hospital mortality amongst the group was 12%. 17% of the group were discharged alive from the ICU within 24 h. Thirty-four percent of the group had an ISS ≤ 15. Of the group which left the ICU alive within 24 h, six patients (0.9%) died in the hospital, 2 % of patients were re-admitted to an ICU, and 0.6% of patients required re-intubation. CONCLUSIONS: We found that 17% of patients who were intubated in the prehospital setting or emergency department and subsequently hospitalized were discharged alive from the ICU within 24 h.

Cerebrospinal Creatine Kinase BB Isoenzyme: A Biomarker for Predicting Outcome After Cardiac Arrest.

BACKGROUND: Cerebrospinal fluid creatine kinase BB isoenzyme (CSF CK-BB) after cardiac arrest (CA) has been shown to have a high positive predictive value for poor neurological outcome, but it has not been evaluated in the setting of targeted temperature management (TTM) and modern CA care. We aimed to evaluate CSF CK-BB as a prognostic biomarker after CA. METHODS: We performed a retrospective cohort study of patients with CA admitted between 2010 and 2020 to a three-hospital health system who remained comatose and had CSF CK-BB assayed between 36 and 84 h after CA. We examined the proportion of patients at hospital discharge who achieved favorable or intermediate neurological outcome, defined as Cerebral Performance Category score of 1-3, compared with those with poor outcome (Cerebral Performance Category score 4-5) for various CSF CK-BB thresholds. We also evaluated additive value of bilateral absence of somatosensory evoked potentials (SSEPs). RESULTS: Among 214 eligible patients, the mean age was 54.7 ± 4.8 years, 72% of patients were male, 33% were nonwhite, 17% had shockable rhythm, 90% were out-of-hospital CA, and 83% received TTM. A total of 19 (9%) awakened. CSF CK-BB ≥ 230 U/L predicted a poor outcome at hospital discharge, with a specificity of 100% (95% confidence interval [CI] 82-100%) and sensitivity of 69% (95% CI 62-76%). When combined with bilaterally absent N20 response on SSEP, specificity remained 100% while sensitivity increased to 80% (95% CI 73-85%). Discordant CK-BB and SSEP findings were seen in 13 (9%) patients. CONCLUSIONS: Cerebrospinal fluid creatine kinase BB isoenzyme levels accurately predicted poor neurological outcome among CA survivors treated with TTM. The CSF CK-BB cutoff of 230 U/L optimizes sensitivity to 69% while maintaining a specificity of 100%. CSF CK-BB could be a useful addition to multimodal neurological prognostication after CA.

An ANXA11 P93S variant dysregulates TDP-43 and causes corticobasal syndrome.

INTRODUCTION: Variants of uncertain significance (VUS) surged with affordable genetic testing, posing challenges for determining pathogenicity. We examine the pathogenicity of a novel VUS P93S in Annexin A11 (ANXA11) - an amyotrophic lateral sclerosis/frontotemporal dementia-associated gene - in a corticobasal syndrome kindred. Established ANXA11 mutations cause ANXA11 aggregation, altered lysosomal-RNA granule co-trafficking, and transactive response DNA binding protein of 43 kDa (TDP-43) mis-localization. METHODS: We described the clinical presentation and explored the phenotypic diversity of ANXA11 variants. P93S's effect on ANXA11 function and TDP-43 biology was characterized in induced pluripotent stem cell-derived neurons alongside multiomic neuronal and microglial profiling. RESULTS: ANXA11 mutations were linked to corticobasal syndrome cases. P93S led to decreased lysosome colocalization, neuritic RNA, and nuclear TDP-43 with cryptic exon expression. Multiomic microglial signatures implicated immune dysregulation and interferon signaling pathways. DISCUSSION: This study establishes ANXA11 P93S pathogenicity, broadens the phenotypic spectrum of ANXA11 mutations, underscores neuronal and microglial dysfunction in ANXA11 pathophysiology, and demonstrates the potential of cellular models to determine variant pathogenicity. HIGHLIGHTS: ANXA11 P93S is a pathogenic variant. Corticobasal syndrome is part of the ANXA11 phenotypic spectrum. Hybridization chain reaction fluorescence in situ hybridization (HCR FISH) is a new tool for the detection of cryptic exons due to TDP-43-related loss of splicing regulation. Microglial ANXA11 and related immune pathways are important drivers of disease. Cellular models are powerful tools for adjudicating variants of uncertain significance.

Total and Subgenomic RNA Viral Load in Patients Infected With SARS-CoV-2 Alpha, Delta, and Omicron Variants.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomic and subgenomic RNA levels are frequently used as a correlate of infectiousness. The impact of host factors and SARS-CoV-2 lineage on RNA viral load is unclear. METHODS: Total nucleocapsid (N) and subgenomic N (sgN) RNA levels were measured by quantitative reverse transcription polymerase chain reaction (RT-qPCR) in specimens from 3204 individuals hospitalized with coronavirus disease 2019 (COVID-19) at 21 hospitals. RT-qPCR cycle threshold (Ct) values were used to estimate RNA viral load. The impact of time of sampling, SARS-CoV-2 variant, age, comorbidities, vaccination, and immune status on N and sgN Ct values were evaluated using multiple linear regression. RESULTS: Mean Ct values at presentation for N were 24.14 (SD 4.53) for non-variants of concern, 25.15 (SD 4.33) for Alpha, 25.31 (SD 4.50) for Delta, and 26.26 (SD 4.42) for Omicron. N and sgN RNA levels varied with time since symptom onset and infecting variant but not with age, comorbidity, immune status, or vaccination. When normalized to total N RNA, sgN levels were similar across all variants. CONCLUSIONS: RNA viral loads were similar among hospitalized adults, irrespective of infecting variant and known risk factors for severe COVID-19. Total N and subgenomic RNA N viral loads were highly correlated, suggesting that subgenomic RNA measurements add little information for the purposes of estimating infectivity.