The NLRP3 inflammasome is essential for IL-18 production in a model of macrophage activation syndrome.

Hyperinflammatory disease is associated with an aberrant immune response resulting in cytokine storm. One such instance of hyperinflammatory disease is known as macrophage activation syndrome (MAS). The pathology of MAS can be characterised by significantly elevated serum levels of interleukin (IL)-18 and interferon (IFN)-γ. Given the role for IL-18 in MAS, we sought to establish the role of inflammasomes in the disease process. Using a murine model of CpG-DNA induced MAS, we discovered that the expression of the NLRP3 inflammasome was increased and correlated with IL-18 production. Inhibition of the NLRP3 inflammasome, or downstream caspase-1, prevented MAS-mediated upregulation of plasma IL-18 but interestingly did not alleviate key features of hyperinflammatory disease including hyperferritinaemia and splenomegaly. Furthermore IL-1 receptor blockade with IL-1Ra did not prevent the development of CpG-induced MAS, despite being clinically effective in the treatment of MAS. These data demonstrate that in the development of MAS, the NLRP3 inflammasome was essential for the elevation in plasma IL-18, a key cytokine in clinical cases of MAS, but was not a driving factor in the pathogenesis of CpG-induced MAS.

Lymphatic network drainage resolves cerebral edema and facilitates recovery from experimental cerebral malaria.

While brain swelling, associated with fluid accumulation, is a known feature of pediatric cerebral malaria (CM), how fluid and macromolecules are drained from the brain during recovery from CM is unknown. Using the experimental CM (ECM) model, we show that fluid accumulation in the brain during CM is driven by vasogenic edema and not by perivascular cerebrospinal fluid (CSF) influx. We identify that fluid and molecules are removed from the brain extremely quickly in mice with ECM to the deep cervical lymph nodes (dcLNs), predominantly through basal routes and across the cribriform plate and the nasal lymphatics. In agreement, we demonstrate that ligation of the afferent lymphatic vessels draining to the dcLNs significantly impairs fluid drainage from the brain and lowers anti-malarial drug recovery from the ECM syndrome. Collectively, our results provide insight into the pathways that coordinate recovery from CM.

Isn't it time for health professionals to shift their focus from preaching politics to promoting peace?

The magnitude of suffering on both the Israeli and Palestinian sides of the current war is beyond comprehension. Political agendas, misinformation and bias related to the conflict are being seen far too frequently in healthcare and medical academia. We believe it is time for healthcare professionals to redirect our attention away from politics and use our medical training to advocate for peace, care, and the welfare of all people, regardless of which side of the conflict they fall into. Politics in the workplace, particularly when disseminated information is divisive and, at times, based on opinion rather than fact, risks significant harm to patients, their families, and healthcare staff, as well as to institutional reputation. If we genuinely care for the well-being of patients and staff, we must lead by example and prevent healthcare systems and medical journals from being hijacked by politics.

The NLRP3 inflammasome is essential for IL-18 production in a murine model of macrophage activation syndrome.

Hyperinflammatory disease is associated with an aberrant immune response resulting in cytokine storm. One such instance of hyperinflammatory disease is known as macrophage activation syndrome (MAS). The pathology of MAS can be characterised by significantly elevated serum levels of interleukin (IL)-18 and interferon (IFN)-γ. Given the role for IL-18 in MAS, we sought to establish the role of inflammasomes in the disease process. Using a murine model of CpG-DNA induced MAS, we discovered that the expression of the NLRP3 inflammasome was increased and correlated with IL-18 production. Inhibition of the NLRP3 inflammasome, or downstream caspase-1, prevented MAS-mediated upregulation of plasma IL-18 but interestingly did not alleviate key features of hyperinflammatory disease including hyperferritinaemia and splenomegaly. Furthermore IL-1 receptor blockade with IL-1Ra did not prevent the development of CpG-induced MAS, despite being clinically effective in the treatment of MAS. These data demonstrate that in the development of MAS, the NLRP3 inflammasome was essential for the elevation in plasma IL-18, a key cytokine in clinical cases of MAS, but was not a driving factor in the pathogenesis of CpG-induced MAS.

The Alopecia Areata Severity and Morbidity Index (ASAMI) Study: Results From a Global Expert Consensus Exercise on Determinants of Alopecia Areata Severity.

Importance: Current measures of alopecia areata (AA) severity, such as the Severity of Alopecia Tool score, do not adequately capture overall disease impact. Objective: To explore factors associated with AA severity beyond scalp hair loss, and to support the development of the Alopecia Areata Severity and Morbidity Index (ASAMI). Evidence Review: A total of 74 hair and scalp disorder specialists from multiple continents were invited to participate in an eDelphi project consisting of 3 survey rounds. The first 2 sessions took place via a text-based web application following the Delphi study design. The final round took place virtually among participants via video conferencing software on April 30, 2022. Findings: Of all invited experts, 64 completed the first survey round (global representation: Africa [4.7%], Asia [9.4%], Australia [14.1%], Europe [43.8%], North America [23.4%], and South America [4.7%]; health care setting: public [20.3%], private [28.1%], and both [51.6%]). A total of 58 specialists completed the second round, and 42 participated in the final video conference meeting. Overall, consensus was achieved in 96 of 107 questions. Several factors, independent of the Severity of Alopecia Tool score, were identified as potentially worsening AA severity outcomes. These factors included a disease duration of 12 months or more, 3 or more relapses, inadequate response to topical or systemic treatments, rapid disease progression, difficulty in cosmetically concealing hair loss, facial hair involvement (eyebrows, eyelashes, and/or beard), nail involvement, impaired quality of life, and a history of anxiety, depression, or suicidal ideation due to or exacerbated by AA. Consensus was reached that the Alopecia Areata Investigator Global Assessment scale adequately classified the severity of scalp hair loss. Conclusions and Relevance: This eDelphi survey study, with consensus among global experts, identified various determinants of AA severity, encompassing not only scalp hair loss but also other outcomes. These findings are expected to facilitate the development of a multicomponent severity tool that endeavors to competently measure disease impact. The findings are also anticipated to aid in identifying candidates for current and emerging systemic treatments. Future research must incorporate the perspectives of patients and the public to assign weight to the domains recognized in this project as associated with AA severity.

Brazilin is a natural product inhibitor of the NLRP3 inflammasome.

Excessive or aberrant NLRP3 inflammasome activation has been implicated in the progression and initiation of many inflammatory conditions; however, currently no NLRP3 inflammasome inhibitors have been approved for therapeutic use in the clinic. Here we have identified that the natural product brazilin effectively inhibits both priming and activation of the NLRP3 inflammasome in cultured murine macrophages, a human iPSC microglial cell line and in a mouse model of acute peritoneal inflammation. Through computational modeling, we predict that brazilin can adopt a favorable binding pose within a site of the NLRP3 protein which is essential for its conformational activation. Our results not only encourage further evaluation of brazilin as a therapeutic agent for NLRP3-related inflammatory diseases, but also introduce this small-molecule as a promising scaffold structure for the development of derivative NLRP3 inhibitor compounds.

Discovery of an inhibitor of DNA-driven inflammation that preferentially targets the AIM2 inflammasome.

Inflammation driven by DNA sensors is now understood to be important to disease pathogenesis. Here, we describe new inhibitors of DNA sensing, primarily of the inflammasome forming sensor AIM2. Biochemistry and molecular modeling has revealed 4-sulfonic calixarenes as potent inhibitors of AIM2 that likely work by binding competitively to the DNA-binding HIN domain. Although less potent, these AIM2 inhibitors also inhibit DNA sensors cGAS and TLR9 demonstrating a broad utility against DNA-driven inflammatory responses. The 4-sulfonic calixarenes inhibited AIM2-dependent post-stroke T cell death, highlighting a proof of concept that the 4-sulfonic calixarenes could be effective at combating post-stroke immunosuppression. By extension, we propose a broad utility against DNA-driven inflammation in disease. Finally, we reveal that the drug suramin, by virtue of its structural similarities, is an inhibitor of DNA-dependent inflammation and propose that suramin could be rapidly repurposed to meet an increasing clinical need.

Primary Human Herpes Virus-6 Causing Recalcitrant Pyrexia after Pilocytic Astrocytoma Resection.

INTRODUCTION: Human herpes virus-6 (HHV-6) is a ubiquitous virus but can lead to deleterious clinical manifestations due to its predilection for the pediatric central nervous system. Despite significant literature describing its common clinical course, it is rarely considered as a causative agent in CSF pleocytosis in the setting of craniotomy and external ventricular drainage device. Identification of a primary HHV-6 infection allowed for timely treatment with an antiviral agent along with earlier discontinuation of antibiotic regimen and expedited placement of a ventriculoperitoneal shunt. CASE PRESENTATION: A two-year-old girl presented with 3 months of progressive gait disturbance and intranuclear ophthalmoplegia. Following craniotomy for removal of 4th ventricular pilocytic astrocytoma and decompression of hydrocephalus, she suffered a prolonged clinical course due to persistent fevers and worsening CSF leukocytosis despite multiple antibiotic regimens. The patient was admitted to the hospital during the COVID-19 pandemic and isolated with her parents in the intensive care unit with strict infection control measures. FilmArray Meningitis/Encephalitis (FAME) panel ultimately detected HHV-6. Clinical confirmation of HHV-6-induced meningitis was proposed given improvement in CSF leukocytosis and fever reduction following the initiation of antiviral medications. Pathologic analysis of brain tumor tissue failed to show HHV-6 genome positivity, suggesting a primary peripheral etiology of infection. CONCLUSION: Here, we present the first known case of HHV-6 infection detected by FAME following intracranial tumor resection. We propose a modified algorithm for persistent fever of unknown origin which may decrease symptomatic sequelae, minimize additional procedures, and shorten length of ICU stay.

Disruptions in endocytic traffic contribute to the activation of the NLRP3 inflammasome.

Inflammation driven by the NLRP3 inflammasome is coordinated through multiple signaling pathways and is regulated by subcellular organelles. Here, we tested the hypothesis that NLRP3 senses disrupted endosome trafficking to trigger inflammasome formation and inflammatory cytokine secretion. NLRP3-activating stimuli disrupted endosome trafficking and triggered localization of NLRP3 to vesicles positive for endolysosomal markers and for the inositol lipid PI4P. Chemical disruption of endosome trafficking sensitized macrophages to the NLRP3 activator imiquimod, driving enhanced inflammasome activation and cytokine secretion. Together, these data suggest that NLRP3 can sense disruptions in the trafficking of endosomal cargoes, which may explain in part the spatial activation of the NLRP3 inflammasome. These data highlight mechanisms that could be exploited in the therapeutic targeting of NLRP3.

Phenolic and quinone methide nor-triterpenes as selective NLRP3 inflammasome inhibitors.

Dysregulated inflammasome activity, particularly of the NLRP3 inflammasome, is associated with the development of several inflammatory diseases. The study of molecules directly targeting NLRP3 is an emerging field in the discovery of new therapeutic compounds for the treatment of inflammatory disorders. Friedelane triterpenes are biologically active phytochemicals having a wide range of activities including anti-inflammatory effects. In this work, we evaluated the potential anti-inflammatory activity of phenolic and quinonemethide nor-triterpenes (1-11) isolated from Maytenus retusa and some semisynthetic derivatives (12-16) through inhibition of the NLRP3 inflammasome in macrophages. Among them, we found that triterpenes 6 and 14 were the most potent, showing markedly reduced caspase-1 activity, IL-1ß secretion (IC50 = 1.15 µM and 0.19 µM, respectively), and pyroptosis (IC50 = 2.21 µM and 0.13 µM, respectively). Further characterization confirmed their selective inhibition of NLRP3 inflammasome in both canonical and non-canonical activation pathways with no effects on AIM2 or NLRC4 inflammasome activation.

The impact of positive reinforcement on teamwork climate, resiliency, and burnout during the COVID-19 pandemic: The TEAM-ICU (Transforming Employee Attitudes via Messaging strengthens Interconnection, Communication, and Unity) pilot study.

Burnout is an internationally recognized occupational phenomenon that negatively impacts the healthcare workforce and its recipients. The aim of this pilot study was to test whether positive reinforcement and integrating a language of support among co-workers can enhance resiliency, facilitate psychological wellness, and encourage hope. This embedded mixed methods prospective, behavioral, interventional study evaluated the effects of positive feedback on wellness among intensive care unit clinicians during the COVID-19 pandemic in a single center, quaternary care medical center. The deliberate positive feedback paradigm has the potential to augment resiliency and improve attitudes toward a teamwork climate. The routine use of deliberate positivity may represent a scalable, low-cost initiative to enhance wellness in a healthcare organization.

A Near Miss of a Retropharyngeal Abscess with MRSA in a 5-Week-Old Boy Due to an Unusual Presentation.

A retropharyngeal abscess (RPA) in early childhood is not uncommon due to at-risk lymph nodes in this deep neck space and is typified by fever, odynophagia, and a constellation of respiratory manifestations. However, RPA is exceedingly rare in the neonatal subpopulation and not part of the usual differential diagnosis algorithm in this age range. Herein, we present a unique case of a previously healthy 5-week-old male infant with protracted "congestion" and difficulty in oral feeding, whose clinical course is confounded by intermittent, positional bradycardia and subsequent apnea. He was eventually diagnosed with a methicillin-resistant Staphylococcus aureus (MRSA) RPA, leading to concurrent vascular and airways compromise in the form of baroreceptor-mediated bradycardia from mass-effect carotid body compression. This clinical case is an important reminder that any infant with positional vital sign changes should prompt urgent and thorough investigation for extraordinary and otherwise uncommon pathophysiologic states. The case also highlights the power of multidisciplinary collaboration across multiple specialties and parental advocacy in unifying a diagnosis for rare pediatric illnesses.

LRRC8A is dispensable for a variety of microglial functions and response to acute stroke.

Microglia, resident brain immune cells, are critical in orchestrating responses to central nervous system (CNS) injury. Many microglial functions, such as phagocytosis, motility and chemotaxis, are suggested to rely on chloride channels, including the volume-regulated anion channel (VRAC), but studies to date have relied on the use of pharmacological tools with limited specificity. VRAC has also been proposed as a drug target for acute CNS injury, and its role in microglial function is of considerable interest for developing CNS therapeutics. This study aimed to definitively confirm the contribution of VRAC in microglia function by using conditional LRRC8A-knockout mice, which lacked the essential VRAC subunit LRRC8A in microglia. We demonstrated that while VRAC contributed to cell volume regulation, it had no effect on phagocytic activity, cell migration or P2YR12-dependent chemotaxis. Moreover, loss of microglial VRAC did not affect microglial morphology or the extent of ischemic damage following stroke. We conclude that VRAC does not critically regulate microglial responses to brain injury and could be targetable in other CNS cell types (e.g., astrocytes) without impeding microglial function. Our results also demonstrate a role for VRAC in cell volume regulation but show that VRAC is not involved in several major cellular functions that it was previously thought to regulate, and point to other, alternative mechanisms of chloride transport in innate immunity.

Itaconate and fumarate derivatives inhibit priming and activation of the canonical NLRP3 inflammasome in macrophages.

The NLRP3 inflammasome is a multiprotein complex that regulates caspase-1 activation and subsequent interleukin (IL)-1ß and IL-18 release from innate immune cells in response to infection or injury. Derivatives of the metabolites itaconate and fumarate, dimethyl itaconate (DMI), 4-octyl itaconate (4OI) and dimethyl fumarate (DMF) limit both expression and release of IL-1ß following NLRP3 inflammasome activation. However, the direct effects of these metabolite derivatives on NLRP3 inflammasome responses require further investigation. Using murine bone marrow-derived macrophages, mixed glia and organotypic hippocampal slice cultures (OHSCs), we demonstrate that DMI, 4OI and DMF pretreatments inhibit pro-inflammatory cytokine production in response to lipopolysaccharide (LPS), as well as inhibit subsequent NLRP3 inflammasome activation induced by nigericin. DMI, 4OI, DMF and monomethyl fumarate (MMF), another fumarate derivative, also directly inhibited biochemical markers of NLRP3 activation in LPS-primed macrophages, mixed glia, OHSCs and human macrophages in response to nigericin and imiquimod, including ASC speck formation, caspase-1 activation, gasdermin D cleavage and IL-1ß release. DMF, an approved treatment of multiple sclerosis, as well as DMI, 4OI and MMF, inhibited NLRP3 activation in macrophages in response to lysophosphatidylcholine, which is used to induce demyelination, suggesting a possible mechanism for DMF in multiple sclerosis through NLRP3 inhibition. The derivatives also reduced pro-IL-1α cleavage in response to the calcium ionophore ionomycin. Together, these findings reveal the immunometabolic regulation of both the priming and activation steps of NLRP3 activation in macrophages. Furthermore, we highlight itaconate and fumarate derivatives as potential therapeutic options in NLRP3- and IL-1α-driven diseases, including in the brain.

Creation of a dedicated line team for critically ill patients with COVID-19: A multidisciplinary approach to maximize resource utilization during the COVID-19 pandemic.

BACKGROUND: Pandemics create challenges for medical centers, which call for innovative adaptations to care for patients during the unusually high census, to distribute stress and work hours among providers, to reduce the likelihood of transmission to health care workers, and to maximize resource utilization. METHODS: We describe a multidisciplinary vascular access team's development to improve frontline providers' workflow by placing central venous and arterial catheters. Herein we describe the development, organization, and processes resulting in the rapid formation and deployment of this team, reporting on notable clinical issues encountered, which might serve as a basis for future quality improvement and investigation. We describe a retrospective, single-center descriptive study in a large, quaternary academic medical center in a major city. The COVID-19 vascular access team included physicians with specialized experience in placing invasive catheters and whose usual clinical schedule had been lessened through deferment of elective cases. The target population included patients with confirmed or suspected COVID-19 in the medical ICU (MICU) needing invasive catheter placement. The line team placed all invasive catheters on patients in the MICU with suspected or confirmed COVID-19. RESULTS AND CONCLUSIONS: Primary data collected were the number and type of catheters placed, time of team member exposure to potentially infected patients, and any complications over the first three weeks. Secondary outcomes pertained to workflow enhancement and quality improvement. 145 invasive catheters were placed on 67 patients. Of these 67 patients, 90% received arterial catheters, 64% central venous catheters, and 25% hemodialysis catheters. None of the central venous catheterizations or hemodialysis catheters were associated with early complications. Arterial line malfunction due to thrombosis was the most frequent complication. Division of labor through specialized expert procedural teams is feasible during a pandemic and offloads frontline providers while potentially conferring safety benefits.

Canine-to-Human Transmission of Mycoplasma canis in the Central Nervous System.

Dog bites remain a common occurrence in our society, particularly in toddlers and small children under the age of 2. Injuries to the head and face, more common in younger children, can often lead to significant morbidity. Additionally, there continues to be considerable clinical equipoise for standardized post-dog bite injury management. Here, we present the only reported pediatric case in the literature of Mycoplasma canis-associated central nervous system (CNS) infection in an 11-month-old infant who sustained a dog bite to the calvarium. The prevalence of dog bites during the SARS-CoV-2 pandemic had interestingly tripled in number after stay-at-home orders in 1 particular pediatric emergency department in Colorado. This observation paired with advances in microbiological identification like MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight mass spectrometer) may lead to the identification of future cases of uniquely canine pathogens that play a role in human infection.

Looking into the IL-1 of the storm: are inflammasomes the link between immunothrombosis and hyperinflammation in cytokine storm syndromes?

Inflammasomes and the interleukin (IL)-1 family of cytokines are key mediators of both inflammation and immunothrombosis. Inflammasomes are responsible for the release of the pro-inflammatory cytokines IL-1ß and IL-18, as well as releasing tissue factor (TF), a pivotal initiator of the extrinsic coagulation cascade. Uncontrolled production of inflammatory cytokines results in what is known as a "cytokine storm" leading to hyperinflammatory disease. Cytokine storms can complicate a variety of diseases and results in hypercytokinemia, coagulopathies, tissue damage, multiorgan failure, and death. Patients presenting with cytokine storm syndromes have a high mortality rate, driven in part by disseminated intravascular coagulation (DIC). While our knowledge on the factors propagating cytokine storms is increasing, how cytokine storm influences DIC remains unknown, and therefore treatments for diseases, where these aspects are a key feature are limited, with most targeting specific cytokines. Currently, no therapies target the immunothrombosis aspect of hyperinflammatory syndromes. Here we discuss how targeting the inflammasome and pyroptosis may be a novel therapeutic strategy for the treatment of hyperinflammation and its associated pathologies.

Subglottic Post-Extubation Upper Airway Obstruction Is Associated With Long-Term Airway Morbidity in Children.

OBJECTIVES: Post-extubation upper airway obstruction is the most common cause of extubation failure in children, but there are few data regarding long-term morbidity. We aim to describe the frequency of long-term airway sequelae in intubated children and determine the association with post-extubation upper airway obstruction. DESIGN: Retrospective, post hoc analysis of previously identified prospective cohort of children in the pediatric/cardiothoracic ICU at Children's Hospital Los Angeles from July 2012 to April 2015. A single provider blinded to the upper airway obstruction classification reviewed the electronic medical records of all patients in the parent study, before and after the index extubation (extubation during parent study), to identify pre-index and post-index upper airway disease. Primary outcomes were prevalence of newly diagnosed airway anomalies following index extubation. SETTING: Single center, tertiary, 391-bed children's hospital. PATIENTS: From the parent study, 327 children younger than 18 years (intubated for at least 12 hr) were included if they received subsequent care (regardless of specialty) after the index extubation. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: New airway anomalies were identified in 40 of 327 children (12.2%). Patients labeled with subglottic upper airway obstruction at the index extubation were more likely to be diagnosed with new airway anomalies on subsequent follow-up, receive long-term Otolaryngology follow-up, or receive airway surgery (all p ≤ 0.006). In multivariable modeling, upper airway obstruction as the primary reason for initial intubation (odds ratio, 3.71; CI, 1.50-9.19), reintubation during the index ICU admission (odds ratio, 4.44; CI, 1.67-11.80), pre-index airway anomaly (odds ratio, 3.31; CI, 1.36-8.01), and post-extubation subglottic upper airway obstruction (odds ratio, 3.50; CI, 1.46-8.34) remained independently associated with the diagnosis of new airway anomalies. CONCLUSIONS: Post-extubation subglottic upper airway obstruction is associated with a three-fold greater odds of long-term airway morbidity. These patients may represent an at-risk population that should be monitored closely after leaving the ICU.