Dual proteomics of infected macrophages reveal bacterial and host players involved in the Francisella intracellular life cycle and cell to cell dissemination by merocytophagy.

Bacterial pathogens adapt and replicate within host cells, while host cells develop mechanisms to eliminate them. Using a dual proteomic approach, we characterized the intra-macrophage proteome of the facultative intracellular pathogen, Francisella novicida. More than 900 Francisella proteins were identified in infected macrophages after a 10-h infection. Biotin biosynthesis-related proteins were upregulated, emphasizing the role of biotin-associated genes in Francisella replication. Conversely, proteins encoded by the Francisella pathogenicity island (FPI) were downregulated, supporting the importance of the F. tularensis Type VI Secretion System for vacuole escape, not cytosolic replication. In the host cell, over 300 proteins showed differential expression among the 6200 identified during infection. The most upregulated host protein was cis-aconitate decarboxylase IRG1, known for itaconate production with antimicrobial properties in Francisella. Surprisingly, disrupting IRG1 expression did not impact Francisella's intracellular life cycle, suggesting redundancy with other immune proteins or inclusion in larger complexes. Over-representation analysis highlighted cell-cell contact and actin polymerization in macrophage deregulated proteins. Using flow cytometry and live cell imaging, we demonstrated that merocytophagy involves diverse cell-to-cell contacts and actin polymerization-dependent processes. These findings lay the groundwork for further exploration of merocytophagy and its molecular mechanisms in future research.Data are available via ProteomeXchange with identifier PXD035145.

Functional diversity of NLRP3 gain-of-function mutants associated with CAPS autoinflammation.

NLRP3-associated autoinflammatory disease is a heterogenous group of monogenic conditions caused by NLRP3 gain-of-function mutations. The poor functional characterization of most NLRP3 variants hinders diagnosis despite efficient anti-IL-1 treatments. Additionally, while NLRP3 is controlled by priming and activation signals, gain-of-functions have only been investigated in response to priming. Here, we characterize 34 NLRP3 variants in vitro, evaluating their activity upon induction, priming, and/or activation signals, and their sensitivity to four inhibitors. We highlight the functional diversity of the gain-of-function mutants and describe four groups based on the signals governing their activation, correlating partly with the symptom severity. We identify a new group of NLRP3 mutants responding to the activation signal without priming, associated with frequent misdiagnoses. Our results identify key NLRP3 residues controlling inflammasome activity and sensitivity to inhibitors, and antagonistic mechanisms with broader efficacy for therapeutic strategies. They provide new insights into NLRP3 activation, an explanatory mechanism for NLRP3-AID heterogeneity, and original tools for NLRP3-AID diagnosis and drug development.

Clinical practice guideline recommendations to improve the mental health of adult trauma patients: protocol for a systematic review.

INTRODUCTION: Mental disorders are common in adult patients with traumatic injuries. To limit the burden of poor psychological well-being in this population, recognised authorities have issued recommendations through clinical practice guidelines (CPGs). However, the uptake of evidence-based recommendations to improve the mental health of trauma patients has been low until recently. This may be explained by the complexity of optimising mental health practices and interpretating CGPs scope and quality. Our aim is to systematically review CPG mental health recommendations in the context of trauma care and appraise their quality. METHODS AND ANALYSIS: We will identify CPG through a search strategy applied to Medline, Embase, CINAHL, PsycINFO and Web of Science databases, as well as guidelines repositories and websites of trauma associations. We will target CPGs on adult and acute trauma populations including at least one recommendation on any prevention, screening, assessment, intervention, patient and family engagement, referral or follow-up procedure related to mental health endorsed by recognised organisations in high-income countries. No language limitations will be applied, and we will limit the search to the last 15 years. Pairs of reviewers will independently screen titles, abstracts, full texts, and carry out data extraction and quality assessment of CPGs using the Appraisal of Guidelines Research and Evaluation (AGREE) II. We will synthesise the evidence on recommendations for CPGs rated as moderate or high quality using a matrix based on the Grading of Recommendations Assessment, Development and Evaluation quality of evidence, strength of recommendation, health and social determinants and whether recommendations were made using a population-based approach. ETHICS AND DISSEMINATION: Ethics approval is not required, as we will conduct secondary analysis of published data. The results will be disseminated in a peer-reviewed journal, at international and national scientific meetings. Accessible summary will be distributed to interested parties through professional, healthcare quality and persons with lived experience associations. PROSPERO REGISTRATION NUMBER: (ID454728).

Neural signal data collection and analysis of Percept™ PC BrainSense recordings for thalamic stimulation in epilepsy.

Deep brain stimulation (DBS) using Medtronic's Percept™ PC implantable pulse generator is FDA-approved for treating Parkinson's disease (PD), essential tremor, dystonia, obsessive compulsive disorder, and epilepsy. Percept™ PC enables simultaneous recording of neural signals from the same lead used for stimulation. Many Percept™ PC sensing features were built with PD patients in mind, but these features are potentially useful to refine therapies for many different disease processes. When starting our ongoing epilepsy research study, we found it difficult to find detailed descriptions about these features and have compiled information from multiple sources to understand it as a tool, particularly for use in patients other than those with PD. Here we provide a tutorial for scientists and physicians interested in using Percept™ PC's features and provide examples of how neural time series data is often represented and saved. We address characteristics of the recorded signals and discuss Percept™ PC hardware and software capabilities in data pre-processing, signal filtering, and DBS lead performance. We explain the power spectrum of the data and how it is shaped by the filter response of Percept™ PC as well as the aliasing of the stimulation due to digitally sampling the data. We present Percept™ PC's ability to extract biomarkers that may be used to optimize stimulation therapy. We show how differences in lead type affects noise characteristics of the implanted leads from seven epilepsy patients enrolled in our clinical trial. Percept™ PC has sufficient signal-to-noise ratio, sampling capabilities, and stimulus artifact rejection for neural activity recording. Limitations in sampling rate, potential artifacts during stimulation, and shortening of battery life when monitoring neural activity at home were observed. Despite these limitations, Percept™ PC demonstrates potential as a useful tool for recording neural activity in order to optimize stimulation therapies to personalize treatment.

Viruses traverse the human proteome through peptide interfaces that can be biomimetically leveraged for drug discovery.

We present a drug design strategy based on structural knowledge of protein-protein interfaces selected through virus-host coevolution and translated into highly potential small molecules. This approach is grounded on Vinland, the most comprehensive atlas of virus-human protein-protein interactions with annotation of interacting domains. From this inspiration, we identified small viral protein domains responsible for interaction with human proteins. These peptides form a library of new chemical entities used to screen for replication modulators of several pathogens. As a proof of concept, a peptide from a KSHV protein, identified as an inhibitor of influenza virus replication, was translated into a small molecule series with low nanomolar antiviral activity. By targeting the NEET proteins, these molecules turn out to be of therapeutic interest in a nonalcoholic steatohepatitis mouse model with kidney lesions. This study provides a biomimetic framework to design original chemistries targeting cellular proteins, with indications going far beyond infectious diseases.

Pathogenicity and virulence of Francisella tularensis.

Tularaemia is a zoonotic disease caused by the Gram-negative bacterium, Francisella tularensis. Depending on its entry route into the organism, F. tularensis causes different diseases, ranging from life-threatening pneumonia to less severe ulceroglandular tularaemia. Various strains with different geographical distributions exhibit different levels of virulence. F. tularensis is an intracellular bacterium that replicates primarily in the cytosol of the phagocytes. The main virulence attribute of F. tularensis is the type 6 secretion system (T6SS) and its effectors that promote escape from the phagosome. In addition, F. tularensis has evolved a peculiar envelope that allows it to escape detection by the immune system. In this review, we cover tularaemia, different Francisella strains, and their pathogenicity. We particularly emphasize the intracellular life cycle, associated virulence factors, and metabolic adaptations. Finally, we present how F. tularensis largely escapes immune detection to be one of the most infectious and lethal bacterial pathogens.

A mathematical model of Familial Mediterranean Fever predicts mechanisms controlling inflammation.

BACKGROUND: Familial Mediterranean Fever (FMF) is a monogenic disease caused by gain-of-function mutations in the MEditerranean FeVer (MEFV) gene. The molecular dysregulations induced by these mutations and the associated causal mechanisms are complex and intricate. OBJECTIVE: We sought to provide a computational model capturing the mechanistic details of biological pathways involved in FMF physiopathology and enabling the study of the patient's immune cell dynamics. METHODS: We carried out a literature survey to identify experimental studies published from January 2000 to December 2020, and integrated its results into a molecular map and a mathematical model. Then, we studied the network of molecular interactions and the dynamic of monocytes to identify key players for inflammation phenotype in FMF patients. RESULTS: We built a molecular map of FMF integrating in a structured manner the current knowledge regarding pathophysiological processes participating in the triggering and perpetuation of the disease flares. The mathematical model derived from the map reproduced patient's monocyte behavior, in particular its proinflammatory role via the Pyrin inflammasome activation. Network analysis and in silico experiments identified NF-κB and JAK1/TYK2 as critical to modulate IL-1ß- and IL-18-mediated inflammation. CONCLUSION: The in silico model of FMF monocyte proved its ability to reproduce in vitro observations. Considering the difficulties related to experimental settings and financial investments to test combinations of stimuli/perturbation in vitro, this model could be used to test complex hypotheses in silico, thus narrowing down the number of in vitro and ex vivo experiments to perform.

Overnight staffing in Canadian neonatal and pediatric intensive care units.

Aim: Infants and children who require specialized medical attention are admitted to neonatal and pediatric intensive care units (ICUs) for continuous and closely supervised care. Overnight in-house physician coverage is frequently considered the ideal staffing model. It remains unclear how often this is achieved in both pediatric and neonatal ICUs in Canada. The aim of this study is to describe overnight in-house physician staffing in Canadian pediatric and level-3 neonatal ICUs (NICUs) in the pre-COVID-19 era. Methods: A national cross-sectional survey was conducted in 34 NICUs and 19 pediatric ICUs (PICUs). ICU directors or their delegates completed a 29-question survey describing overnight staffing by resident physicians, fellow physicians, nurse practitioners, and attending physicians. A comparative analysis was conducted between ICUs with and without in-house physicians. Results: We obtained responses from all 34 NICUs and 19 PICUs included in this study. A total of 44 ICUs (83%) with in-house overnight physician coverage provided advanced technologies, such as extracorporeal life support, and included all ICUs that catered to patients with cardiac, transplant, or trauma conditions. Residents provided the majority of overnight coverage, followed by the Critical Care Medicine fellows. An attending physician was in-house overnight in eight (15%) out of the 53 ICUs, seven of which were NICUs. Residents participating in rotations in the ICU would often have rotation durations of less than 6 weeks and were often responsible for providing care during shifts lasting 20-24 h. Conclusion: Most PICUs and level-3 NICUs in Canada have a dedicated in-house physician overnight. These physicians are mainly residents or fellows, but a notable variation exists in this arrangement. The potential effects on patient outcomes, resident learning, and physician satisfaction remain unclear and warrant further investigation.

Phosphoprotein phosphatase activity positively regulates oligomeric pyrin to trigger inflammasome assembly in phagocytes.

IMPORTANCE: Pyrin, a unique cytosolic receptor, initiates inflammatory responses against RhoA-inactivating bacterial toxins and effectors like Yersinia's YopE and YopT. Understanding pyrin regulation is crucial due to its association with dysregulated inflammatory responses, including Familial Mediterranean Fever (FMF), linked to pyrin gene mutations. FMF mutations historically acted as a defense mechanism against plague. Negative regulation of pyrin through PKN phosphorylation is well established, with Yersinia using the YopM effector to promote pyrin phosphorylation and counteract its activity. This study highlights the importance of phosphoprotein phosphatase activity in positively regulating pyrin inflammasome assembly in phagocytic cells of humans and mice. Oligomeric murine pyrin has S205 phosphorylated before inflammasome assembly, and this study implicates the dephosphorylation of murine pyrin S205 by two catalytic subunits of PP2A in macrophages. These findings offer insights for investigating the regulation of oligomeric pyrin and the balance of kinase and phosphatase activity in pyrin-associated infectious and autoinflammatory diseases.

Evaluating the effectiveness of a multifaceted intervention to reduce low-value care in adults hospitalized following trauma: a protocol for a pragmatic cluster randomized controlled trial.

BACKGROUND: While simple Audit & Feedback (A&F) has shown modest effectiveness in reducing low-value care, there is a knowledge gap on the effectiveness of multifaceted interventions to support de-implementation efforts. Given the need to make rapid decisions in a context of multiple diagnostic and therapeutic options, trauma is a high-risk setting for low-value care. Furthermore, trauma systems are a favorable setting for de-implementation interventions as they have quality improvement teams with medical leadership, routinely collected clinical data, and performance-linked to accreditation. We aim to evaluate the effectiveness of a multifaceted intervention for reducing low-value clinical practices in acute adult trauma care. METHODS: We will conduct a pragmatic cluster randomized controlled trial (cRCT) embedded in a Canadian provincial quality assurance program. Level I-III trauma centers (n = 30) will be randomized (1:1) to receive simple A&F (control) or a multifaceted intervention (intervention). The intervention, developed using extensive background work and UK Medical Research Council guidelines, includes an A&F report, educational meetings, and facilitation visits. The primary outcome will be the use of low-value initial diagnostic imaging, assessed at the patient level using routinely collected trauma registry data. Secondary outcomes will be low-value specialist consultation, low-value repeat imaging after a patient transfer, unintended consequences, determinants for successful implementation, and incremental cost-effectiveness ratios. DISCUSSION: On completion of the cRCT, if the intervention is effective and cost-effective, the multifaceted intervention will be integrated into trauma systems across Canada. Medium and long-term benefits may include a reduction in adverse events for patients and an increase in resource availability. The proposed intervention targets a problem identified by stakeholders, is based on extensive background work, was developed using a partnership approach, is low-cost, and is linked to accreditation. There will be no attrition, identification, or recruitment bias as the intervention is mandatory in line with trauma center designation requirements, and all outcomes will be assessed with routinely collected data. However, investigators cannot be blinded to group allocation and there is a possibility of contamination bias that will be minimized by conducting intervention refinement only with participants in the intervention arm. TRIAL REGISTRATION: This protocol has been registered on ClinicalTrials.gov (February 24, 2023, # NCT05744154 ).

Gut microbiota alterations are associated with phenotype and genotype in familial Mediterranean fever.

OBJECTIVES: Familial Mediterranean fever (FMF) is the most common monogenic autoinflammatory disease associated with MEFV mutations. Disease phenotype and response to treatment vary from one patient to another despite similar genotype suggesting the role of environmental factors. We analyze the gut microbiota of a large cohort of FMF patients in relation to disease characteristics. METHODS: The gut microbiota of 119 FMF patients and 61 healthy controls was analyzed using 16 s rRNA gene sequencing. Associations between bacterial taxa, clinical characteristics and genotypes were evaluated using multivariable association with linear models (MaAslin2) adjusting on age, sex, genotype, presence of AA amyloidosis (n = 17), hepatopathy (n = 5), colchicine intake, colchicine resistance (n = 27), use of biotherapy (n = 10), CRP levels, and number of daily feces. Bacterial network structures were also analysed. RESULTS: The gut microbiota of FMF patients differs from controls, with an increase in pro-inflammatory bacteria, such as Enterobacter, Klebsiella and Ruminococcus gnavus group. Disease characteristics and resistance to colchicine correlated with homozygous mutations and were associated with specific microbiota alteration. Colchicine treatment was associated with the expansion of anti-inflammatory taxa such as Faecalibacterium and Roseburia, while FMF severity was associated with the expansion of Ruminococcus gnavus group and Paracoccus. Colchicine-resistant patients exhibited an alteration of the bacterial network structure with decreased inter-taxa connectivity. CONCLUSION: The gut microbiota of FMF patients correlates with disease characteristics and severity, with an increase in pro-inflammatory taxa in the most severe patients. This suggests a specific role for the gut microbiota in shaping FMF outcomes and response to treatment.

Effect of a Continuous Bedside Pressure Mapping System for Reducing Interface Pressures: A Randomized Clinical Trial.

Importance: Continuous bedside pressure mapping (CBPM) technology can assist in detecting skin areas with excessive interface pressure and inform efficient patient repositioning to prevent the development of pressure injuries (PI). Objective: To evaluate the efficacy of CBPM technology in reducing interface pressure and the incidence of PIs. Design, Setting, and Participants: This parallel, 2-group randomized clinical trial was performed at a tertiary acute care center. The study started to enroll participants in December 2014 and was completed in May 2018. Participants included adults partially or completely dependent for bed mobility. Statistical analysis was performed from September 2018 to December 2022. Intervention: Nursing staff using visual feedback from CBPM technology for 72 hours. Main Outcomes and Measures: Absolute number of sensing points with pressure readings greater than 40 mm Hg, mean interface pressure across all sensing points under a patient's body, proportion of participants who had pressure readings greater than 40 mm Hg, and pressure-related skin and soft tissue changes. Results: There were 678 patients recruited. After attrition, 260 allocated to the control group (151 [58.1%] male; mean [SD] age, 61.9 [18.5] years) and 247 in the intervention group (147 [59.5%] male; mean [SD] age, 63.6 [18.1] years) were included in analyses. The absolute number of sensing points with pressures greater than 40 mm Hg were 11 033 in the control group vs 9314 in the intervention group (P = .16). The mean (SD) interface pressure was 6.80 (1.63) mm Hg in the control group vs 6.62 (1.51) mm Hg in the intervention group (P = .18). The proportion of participants who had pressure readings greater than 40 mm Hg was 99.6% in both the control and intervention groups. Conclusions and Relevance: In this randomized clinical trial to evaluate the efficacy of CBPM technology in the reduction of interface pressure and the incidence of PIs in a tertiary acute care center, no statistically significant benefit was seen for any of the primary outcomes. These results suggest that longer duration of monitoring and adequately powered studies where CBPM feedback is integrated into a multifaceted intervention to prevent PI are needed. Trial Registration: ClinicalTrials.gov Identifier: NCT02325388.

Codesigning a user-centred digital psychoeducational tool for youth mental well-being with families in Canada: study protocol for a sequential exploratory mixed methods study.

INTRODUCTION: On 11 March 2020, WHO declared the novel coronavirus (COVID-19) disease a global pandemic. Governments globally implemented physical distancing measures and closure of public institutions that resulted in varying implications to youth mental well-being (eg, social isolation, reduced extracurricular activities). These impacts may have detrimental short-term and long-term effects on youth mental well-being; care for youth with mental health disorders was already overstretched, underfunded and fragmented before the pandemic and youth are not often considered in mental health initiatives. There is a pressing need to partner with youth and families to target and improve youth mental well-being prior to the onset of a mental health disorder, as well as to conduct research on youth mental well-being needs related to pandemic recovery. Here we present a protocol for partnering with youth and families to codesign a user-centred digital tool for youth mental well-being. METHODS AND ANALYSIS: We will conduct a national research study to develop a catalogue of recommendations specific to supporting youth mental well-being, and a digital tool to support youth mental well-being through three phases of work: (1) expert consultation on data related to supporting youth mental well-being existing within our Pandemic Preparedness Research Program; (2) codesign of an innovative digital tool for youth mental well-being; and (3) assessment of the tool's usability and acceptability. ETHICS AND DISSEMINATION: This study has been approved by the Dalhousie Research Ethics Board (2023-6538) and the Conjoint Health Research Ethics Board (23-0039). This study will complement ongoing foundational research in youth conducted by our team that involves partnering with youth and families to understand the unique implications of the pandemic on this population.

Comparative study of GBP recruitment on two cytosol-dwelling pathogens, Francisella novicida and Shigella flexneri highlights differences in GBP repertoire and in GBP1 motif requirements.

Guanylate-Binding Proteins are interferon-inducible GTPases that play a key role in cell autonomous responses against intracellular pathogens. Despite sharing high sequence similarity, subtle differences among GBPs translate into functional divergences that are still largely not understood. A key GBP feature is the formation of supramolecular GBP complexes on the bacterial surface. Such complexes are observed when GBP1 binds lipopolysaccharide (LPS) from Shigella and Salmonella and further recruits GBP2-4. Here, we compared GBP recruitment on two cytosol-dwelling pathogens, Francisella novicida and S. flexneri. Francisella novicida was coated by GBP1 and GBP2 and to a lower extent by GBP4 in human macrophages. Contrary to S. flexneri, F. novicida was not targeted by GBP3, a feature independent of T6SS effectors. Multiple GBP1 features were required to promote targeting to F. novicida while GBP1 targeting to S. flexneri was much more permissive to GBP1 mutagenesis suggesting that GBP1 has multiple domains that cooperate to recognize F. novicida atypical LPS. Altogether our results indicate that the repertoire of GBPs recruited onto specific bacteria is dictated by GBP-specific features and by specific bacterial factors that remain to be identified.

Inducing Pyroptosis Via the Pyrin Inflammasome.

The pyrin inflammasome detects bacterial toxins and effectors that inhibit RhoA GTPases and triggers inflammatory cytokine release and a fast cell death termed pyroptosis. In addition, various endogenous molecules, drugs, synthetic molecules, or mutations can trigger pyrin inflammasome activation. The pyrin protein differs between humans and mice, and the repertoire of pyrin activators is also species-specific. Here, we present the various pyrin inflammasome activators, inhibitors, the kinetics of pyrin activation in response to the various activators, and their species specificity. In addition, we present different methods to monitor pyrin-mediated pyroptosis.

Investigating survival, quality of life and cognition in PROton versus photon therapy for IDH-mutated diffuse grade 2 and 3 GLIOmas (PRO-GLIO): a randomised controlled trial in Norway and Sweden.

INTRODUCTION: The use of proton therapy increases globally despite a lack of randomised controlled trials demonstrating its efficacy and safety. Proton therapy enables sparing of non-neoplastic tissue from radiation. This is principally beneficial and holds promise of reduced long-term side effects. However, the sparing of seemingly non-cancerous tissue is not necessarily positive for isocitrate dehydrogenase (IDH)-mutated diffuse gliomas grade 2-3, which have a diffuse growth pattern. With their relatively good prognosis, yet incurable nature, therapy needs to be delicately balanced to achieve a maximal survival benefit combined with an optimised quality of life. METHODS AND ANALYSIS: PRO-GLIO (PROton versus photon therapy in IDH-mutated diffuse grade 2 and 3 GLIOmas) is an open-label, multicentre, randomised phase III non-inferiority study. 224 patients aged 18-65 years with IDH-mutated diffuse gliomas grade 2-3 from Norway and Sweden will be randomised 1:1 to radiotherapy delivered with protons (experimental arm) or photons (standard arm). First intervention-free survival at 2 years is the primary endpoint. Key secondary endpoints are fatigue and cognitive impairment, both at 2 years. Additional secondary outcomes include several survival measures, health-related quality of life parameters and health economy endpoints. ETHICS AND DISSEMINATION: To implement proton therapy as part of standard of care for patients with IDH-mutated diffuse gliomas grade 2-3, it should be deemed safe. With its randomised controlled design testing proton versus photon therapy, PRO-GLIO will provide important information for this patient population concerning safety, cognition, fatigue and other quality of life parameters. As proton therapy is considerably more costly than its photon counterpart, cost-effectiveness will also be evaluated. PRO-GLIO is approved by ethical committees in Norway (Regional Committee for Medical & Health Research Ethics) and Sweden (The Swedish Ethical Review Authority) and patient inclusion has commenced. Trial results will be published in international peer-reviewed journals, relevant conferences, national and international meetings and expert forums. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov Registry (NCT05190172).

Mutations in the B30.2 and the central helical scaffold domains of pyrin differentially affect inflammasome activation.

Familial Mediterranean Fever (FMF) is the most common monogenic autoinflammatory disorder. FMF is caused by mutations in the MEFV gene, encoding pyrin, an inflammasome sensor. The best characterized pathogenic mutations associated with FMF cluster in exon 10. Yet, mutations have been described along the whole MEFV coding sequence. Exon 10 encodes the B30.2 domain of the pyrin protein, but the function of this human-specific domain remains unclear. Pyrin is an inflammasome sensor detecting RhoA GTPase inhibition following exposure to bacterial toxins such as TcdA. Here, we demonstrate that the B30.2 domain is dispensable for pyrin inflammasome activation in response to this toxin. Deletion of the B30.2 domain mimics the most typical FMF-associated mutation and confers spontaneous inflammasome activation in response to pyrin dephosphorylation. Our results indicate that the B30.2 domain is a negative regulator of the pyrin inflammasome that acts independently from and downstream of pyrin dephosphorylation. In addition, we identify the central helical scaffold (CHS) domain of pyrin, which lies immediately upstream of the B30.2 domain as a second regulatory domain. Mutations affecting the CHS domain mimic pathogenic mutations in the B30.2 domain and render the pyrin inflammasome activation under the sole control of the dephosphorylation. In addition, specific mutations in the CHS domain strongly increase the cell susceptibility to steroid catabolites, recently described to activate pyrin, in both a cell line model and in monocytes from genotype-selected FMF patients. Taken together, our work reveals the existence of two distinct regulatory regions at the C-terminus of the pyrin protein, that act in a distinct manner to regulate positively or negatively inflammasome activation. Furthermore, our results indicate that different mutations in pyrin regulatory domains have different functional impacts on the pyrin inflammasome which could contribute to the diversity of pyrin-associated autoinflammatory diseases.

Family coaching during Spontaneous Awakening Trials and Spontaneous Breathing Trials (FamCAB): pilot study protocol.

INTRODUCTION: Many patients in the intensive care unit (ICU) require weaning from deep sedation (Spontaneous Awakening Trials, SATs) and mechanical ventilation (Spontaneous Breathing Trials, SBTs) in their journey to recovery. These procedures can be distressing for patients and their families. The presence of family members as 'coaches' during SATs/SBTs could provide patients with reassurance, reduce stress for patients and families and potentially improve procedural success rates. METHODS AND ANALYSIS: This study will be executed in two phases:Development of a coaching module: a working group including patient partners (i.e., former ICU patients or family members of former ICU patients), researchers, and ICU clinicians will develop an educational module on family coaching during SATs/SBTs (FamCAB). This module will provide families of critically ill patients basic information about SATs/SBTs as well as coaching guidance.Pilot testing: family members of ICU patients will complete the FamCAB module and provide information on: (1) demographics, (2) anxiety and (3) satisfaction with care in the ICU. Family members will then coach the patient through the next clinically indicated SATs and/or SBTs. Information around duration of time and success rates of SATs and/or SBTs (ability to conduct a complete assessment) alongside feedback will be collected. ICU clinical staff (including physicians and nurses) will be asked for feedback on practicality and perceived benefits or drawbacks of family coaching during these procedures. Feasibility and acceptability of family coaching in SATs/SBTs will be determined. DISCUSSION: The results of this work will inform whether a larger study to explore family coaching during SATs/SBTs is warranted. ETHICS AND DISSEMINATION: This study has received ethical approval from the University of Calgary Conjoint Health Research Ethics Board. Results from this pilot study will be made available via peer-reviewed journals and presented at critical care conferences on completion.

Time-Course of Changes in Multidimensional Fatigue and Functional Exercise Capacity and Their Associations during a Short Inpatient Pulmonary Rehabilitation Program.

This study aimed to assess the time-course of changes in multidimensional fatigue and functional exercise capacity and their associations during an inpatient pulmonary rehabilitation (PR) program. Seventy COPD patients from three centres were enrolled for a four-week PR program and were evaluated before (T0) and at the end of each week (T1, T2, T3, and T4). Weekly change in multidimensional fatigue was assessed by the multidimensional inventory questionnaire (MFI-20) and functional exercise capacity by the 6-minute walking distance (6MWD). Reaction time (RT) and heart rate variability (HRV) were also assessed as complementary markers of fatigue. HRV did not change during the study (all p > 0.05). MFI-20 score and RT decreased during the first part of the program (p < 0.001) and levelled off at T2 (all p > 0.05 compared with each preceding time). While 6MWD improved by almost 70% during the first part of the PR, it continued to increase, albeit at a greatly reduced pace, between T2 and T4 (p < 0.05). In parallel, a negative association was found between MFI-20 score and 6MWD at each evaluation time (r ranged from 0.43 to 0.71), with a significantly stronger T3 correlation compared with the other time periods (all p < 0.05). The strengthening of the association between fatigue and functional exercise capacity at T3, which occurred concomitantly with the slowdown of functional exercise capacity improvement, is consistent with a role for fatigue in the limitation of performance changes during PR. The limitation of fatigue during PR is thus an interesting aspect to improve the magnitude of performance changes.

Reliability of visual review of intracranial electroencephalogram in identifying the seizure onset zone: A systematic review and implications for the accuracy of automated methods.

Visual review of intracranial electroencephalography (iEEG) is often an essential component for defining the zone of resection for epilepsy surgery. Unsupervised approaches using machine and deep learning are being employed to identify seizure onset zones (SOZs). This prompts a more comprehensive understanding of the reliability of visual review as a reference standard. We sought to summarize existing evidence on the reliability of visual review of iEEG in defining the SOZ for patients undergoing surgical workup and understand its implications for algorithm accuracy for SOZ prediction. We performed a systematic literature review on the reliability of determining the SOZ by visual inspection of iEEG in accordance with best practices. Searches included MEDLINE, Embase, Cochrane Library, and Web of Science on May 8, 2022. We included studies with a quantitative reliability assessment within or between observers. Risk of bias assessment was performed with QUADAS-2. A model was developed to estimate the effect of Cohen kappa on the maximum possible accuracy for any algorithm detecting the SOZ. Two thousand three hundred thirty-eight articles were identified and evaluated, of which one met inclusion criteria. This study assessed reliability between two reviewers for 10 patients with temporal lobe epilepsy and found a kappa of .80. These limited data were used to model the maximum accuracy of automated methods. For a hypothetical algorithm that is 100% accurate to the ground truth, the maximum accuracy modeled with a Cohen kappa of .8 ranged from .60 to .85 (F-2). The reliability of reviewing iEEG to localize the SOZ has been evaluated only in a small sample of patients with methodologic limitations. The ability of any algorithm to estimate the SOZ is notably limited by the reliability of iEEG interpretation. We acknowledge practical limitations of rigorous reliability analysis, and we propose design characteristics and study questions to further investigate reliability.