Therapeutic Avenues to Modulate B-Cell Function in Patients With Cardiovascular Disease.

The adaptive immune system plays an important role in the development and progression of atherosclerotic cardiovascular disease. B cells can have both proatherogenic and atheroprotective roles, making treatments aimed at modulating B cells important therapeutic targets. The innate-like B-cell response is generally considered atheroprotective, while the adaptive response is associated with mixed consequences for atherosclerosis. Additionally, interactions of B cells with components of the adaptive and innate immune system, including T cells and complement, also represent key points for therapeutic regulation. In this review, we discuss therapeutic approaches based on B-cell depletion, modulation of B-cell survival, manipulation of both the antibody-dependent and antibody-independent B-cell response, and emerging immunization techniques.

Early Evidence of Chronic Obstructive Pulmonary Disease Obscured by Race-Specific Prediction Equations.

Rationale: The identification of early chronic obstructive pulmonary disease (COPD) is essential to appropriately counsel patients regarding smoking cessation, provide symptomatic treatment, and eventually develop disease-modifying treatments. Disease severity in COPD is defined using race-specific spirometry equations. These may disadvantage non-White individuals in diagnosis and care. Objectives: Determine the impact of race-specific equations on African American (AA) versus non-Hispanic White individuals. Methods: Cross-sectional analyses of the COPDGene (Genetic Epidemiology of Chronic Obstructive Pulmonary Disease) cohort were conducted, comparing non-Hispanic White (n = 6,766) and AA (n = 3,366) participants for COPD manifestations. Measurements and Main Results: Spirometric classifications using race-specific, multiethnic, and "race-reversed" prediction equations (NHANES [National Health and Nutrition Examination Survey] and Global Lung Function Initiative "Other" and "Global") were compared, as were respiratory symptoms, 6-minute-walk distance, computed tomography imaging, respiratory exacerbations, and St. George's Respiratory Questionnaire. Application of different prediction equations to the cohort resulted in different classifications by stage, with NHANES and Global Lung Function Initiative race-specific equations being minimally different, but race-reversed equations moving AA participants to more severe stages and especially between the Global Initiative for Chronic Obstructive Lung Disease (GOLD) stage 0 and preserved ratio impaired spirometry groups. Classification using the established NHANES race-specific equations demonstrated that for each of GOLD stages 1-4, AA participants were younger, had fewer pack-years and more current smoking, but had more exacerbations, shorter 6-minute-walk distance, greater dyspnea, and worse BODE (body mass index, airway obstruction, dyspnea, and exercise capacity) scores and St. George's Respiratory Questionnaire scores. Differences were greatest in GOLD stages 1 and 2. Race-reversed equations reclassified 774 AA participants (43%) from GOLD stage 0 to preserved ratio impaired spirometry. Conclusions: Race-specific equations underestimated disease severity among AA participants. These effects were particularly evident in early disease and may result in late detection of COPD.

Computational Design of Bidentate Hypervalent Iodine Catalysts in Halogen Bond-Mediated Organocatalysis.

In recent years, halogen bond-based organocatalysis has garnered significant attention as an alternative to hydrogen-based catalysis, capturing considerable interest within the scientific community. This transition has witnessed the evolution of catalytic scaffolds from monodentate to bidentate architectures, and from monovalent to hypervalent species. In this DFT-based study, we explored a bidentate hypervalent iodine(III)-based system that has already undergone experimental validation. Additionally, we explore various functionalisations (-CF$_3$, -CH$_3$, -tBu, -OH, -OMe, -NO$_2$, -CN) and scaffold modifications, such as sulfur oxidation, theoretically proposed for an indole-based Michael addition. The investigated systems favour bidentate O-type binding, underlining the importance of ligand coordination in catalytic activity. Electron-deficient scaffolds exhibited stronger binding and lower activation energies, indicating the pivotal role of electronic properties for $\sigma$-hole-based catalysis. Of these groups, Lewis-base-like moieties formed stabilising intramolecular interactions with hypervalent iodines when in the ortho-position. Furthermore, inductive electron withdrawal was deemed more effective than mesomeric withdrawal in enhancing catalytic efficacy for these systems. Lastly, increasing sulfur oxidation was theoretically proven to improve catalytic activity significantly.

Ultrasound-guided paramedian approach: A novel technique for lumbar puncture in the Emergency Department.

Lumbar puncture is performed to evaluate for multiple neurologic conditions, including meningitis and subarachnoid hemorrhage. However, success rates with the landmark-based technique are limited. Ultrasound is most commonly used for pre-marking without dynamic guidance, which presents several limitations, including absence of real-time guidance and lack of reliability if any patient movement occurs after skin marking. We describe a novel, ultrasound-guided paramedian approach which was successfully performed in the Emergency Department setting for lumbar puncture. Physicians should consider this technique as an alternate model using real-time guidance to reduce needle passes in those with difficult anatomy.

Artificial intelligence and point-of-care ultrasound: Benefits, limitations, and implications for the future.

The utilization of artificial intelligence (AI) in medical imaging has become a rapidly growing field as a means to address contemporary demands and challenges of healthcare. Among the emerging applications of AI is point-of-care ultrasound (POCUS), in which the combination of these two technologies has garnered recent attention in research and clinical settings. In this Controversies paper, we will discuss the benefits, limitations, and future considerations of AI in POCUS for patients, clinicians, and healthcare systems.

Impact of tertiary lymphoid structures on prognosis and therapeutic response in pancreatic ductal adenocarcinoma.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is known to have a heterogeneous desmoplastic tumour microenvironment (TME) with a large number of immunosuppressive cells. Recently, high B-cell infiltration in PDAC has received growing interest as a potential therapeutic target. METHODS: Our literature review summarises the characteristics of tumour-associated tertiary lymphoid structures (TLSs) and highlight the key studies exploring the clinical outcomes of TLSs in PDAC patients and the direct effect on the TME. RESULTS: The location, density and maturity stages of TLSs within tumours play a key role in determining the prognosis and is a new emerging target in cancer immunotherapy. DISCUSSION: TLS development is imperative to improve the prognosis of PDAC patients. In the future, studying the genetics and immune characteristics of tumour infiltrating B cells and TLSs may lead towards enhancing adaptive immunity in PDAC and designing personalised therapies.

Use of the Spirometric "Fixed-Ratio" Underdiagnoses COPD in African-Americans in a Longitudinal Cohort Study.

BACKGROUND: COPD diagnosis is tightly linked to the fixed-ratio spirometry criteria of FEV1/FVC < 0.7. African-Americans are less often diagnosed with COPD. OBJECTIVE: Compare COPD diagnosis by fixed-ratio with findings and outcomes by race. DESIGN: Genetic Epidemiology of COPD (COPDGene) (2007-present), cross-sectional comparing non-Hispanic white (NHW) and African-American (AA) participants for COPD diagnosis, manifestations, and outcomes. SETTING: Multicenter, longitudinal US cohort study. PARTICIPANTS: Current or former smokers with ≥ 10-pack-year smoking history enrolled at 21 clinical centers including over-sampling of participants with known COPD and AA. Exclusions were pre-existing non-COPD lung disease, except for a history of asthma. MEASUREMENTS: Subject diagnosis by conventional criteria. Mortality, imaging, respiratory symptoms, function, and socioeconomic characteristics, including area deprivation index (ADI). Matched analysis (age, sex, and smoking status) of AA vs. NHW within participants without diagnosed COPD (GOLD 0; FEV1 ≥ 80% predicted and FEV1/FVC ≥ 0.7). RESULTS: Using the fixed ratio, 70% of AA (n = 3366) were classified as non-COPD, versus 49% of NHW (n = 6766). AA smokers were younger (55 vs. 62 years), more often current smoking (80% vs. 39%), with fewer pack-years but similar 12-year mortality. Density distribution plots for FEV1 and FVC raw spirometry values showed disproportionate reductions in FVC relative to FEV1 in AA that systematically led to higher ratios. The matched analysis demonstrated GOLD 0 AA had greater symptoms, worse DLCO, spirometry, BODE scores (1.03 vs 0.54, p < 0.0001), and greater deprivation than NHW. LIMITATIONS: Lack of an alternative diagnostic metric for comparison. CONCLUSIONS: The fixed-ratio spirometric criteria for COPD underdiagnosed potential COPD in AA participants when compared to broader diagnostic criteria. Disproportionate reductions in FVC relative to FEV1 leading to higher FEV1/FVC were identified in these participants and associated with deprivation. Broader diagnostic criteria for COPD are needed to identify the disease across all populations.

Metabolic Adaptation of Macrophages as Mechanism of Defense against Crystalline Silica.

Silicosis is a lethal pneumoconiosis for which no therapy is available. Silicosis is a global threat, and more than 2.2 million people per year are exposed to silica in the United States. The initial response to silica is mediated by innate immunity. Phagocytosis of silica particles by macrophages is followed by recruitment of mitochondria to phagosomes, generation of mitochondrial reactive oxygen species, and cytokine (IL-1ß, TNF-α, IFN-ß) release. In contrast with LPS, the metabolic remodeling of silica-exposed macrophages is unclear. This study contrasts mitochondrial and metabolic alterations induced by LPS and silica on macrophages and correlates them with macrophage viability and cytokine production, which are central to the pathogenesis of silicosis. Using high-resolution respirometer and liquid chromatography-high-resolution mass spectrometry, we determined the effects of silica and LPS on mitochondrial respiration and determined changes in central carbon metabolism of murine macrophage cell lines RAW 264.7 and IC-21. We show that silica induces metabolic reprogramming of macrophages. Silica, as well as LPS, enhances glucose uptake and increases aerobic glycolysis in macrophages. In contrast with LPS, silica affects mitochondria respiration, reducing complex I and enhancing complex II activity, to sustain cell viability. These mitochondrial alterations are associated in silica, but not in LPS-exposed macrophages, with reductions of tricarboxylic acid cycle intermediates, including succinate, itaconate, glutamate, and glutamine. Furthermore, in contrast with LPS, these silica-induced metabolic adaptations do not correlate with IL-1ß or TNF-α production, but with the suppressed release of IFN-ß. Our data highlight the importance of complex II activity and tricarboxylic acid cycle remodeling to macrophage survival and cytokine-mediated inflammation in silicosis.

Comparison of dynamic versus static ultrasound to confirm endotracheal tube depth.

INTRODUCTION: After endotracheal intubation is performed, the location of the endotracheal tube (ETT) is confirmed followed by assessment of ETT depth. Physical examination can be unreliable and chest radiographs can lead to delayed recognition. Ultrasound may facilitate rapid determination of ETT depth at the bedside; however, the ideal technique is unknown. METHODS: This was a randomized trial comparing the static versus dynamic technique for ETT depth assessment using a cadaver model. The ETT was randomized to correct versus deep placement. Seven physicians blinded to ETT location assessed the location using static (direct visualization of an inflated cuff) versus dynamic (active inflation of the ETT cuff) visualization. Outcomes included diagnostic accuracy, time to identification, and operator confidence with subgroup analyses by physician ultrasound experience. RESULTS: 420 total assessments were performed. The static technique was 99.1% (95% CI 94.8%-100%) sensitive and 97.1% (95% CI 91.9%-99.4%) specific. The dynamic technique was 100% (95% CI 96.7%-100%) sensitive and 100% (95% CI 96.7%-100%) specific. Time to identification was faster for the static technique (6.6 s; 95% CI 5.9-7.4 s) versus the dynamic technique (8.7 s; 95% CI 8.0-9.5 s). Operator confidence was lower for the static technique (4.4/5.0; 95% CI 4.3-4.5) versus the dynamic technique (4.7/5.0; 95% CI 4.6-4.8). There were no differences in the findings when assessed among expert or non-expert sonographers. CONCLUSION: There was no statistically significant difference in the accuracy of ETT depth identification between the static or dynamic technique. However, utilizing the dynamic technique showed a statistically significant improvement in sonographer confidence and a concomitant increase in time to identification.

Sustained radiation reduction following initial quality improvement intervention in a paediatric cardiac catheterisation laboratory.

BACKGROUND: As part of a quality improvement project beginning in October 2011, our centre introduced changes to reduce radiation exposure during paediatric cardiac catheterisations. This led to significant initial decreases in radiation to patients. Starting in April 2016, we sought to determine whether these initial reductions were sustained. METHODS: After a 30-day trial period, we implemented (1) weight-based reductions in preset frame rates for fluoroscopy and angiography, (2) increased use of collimators and safety shields, (3) utilisation of stored fluoroscopy and virtual magnification, and (4) hiring of a devoted radiation technician. We collected patient weight (kg), total fluoroscopy time (min), and procedure radiation dosage (cGy-cm2) for cardiac catheterisations between October, 2011 and September, 2019. RESULTS: A total of 1889 procedures were evaluated (196 pre-intervention, 303 in the post-intervention time period, and 1400 in the long-term group). Fluoroscopy times (18.3 ± 13.6 pre; 19.8 ± 14.1 post; 17.11 ± 15.06 long-term, p = 0.782) were not significantly different between the three groups. Patient mean radiation dose per kilogram decreased significantly after the initial quality improvement intervention (39.7% reduction, p = 0.039) and was sustained over the long term (p = 0.043). Provider radiation exposure was also significantly decreased from the onset of this project through the long-term period (overall decrease of 73%, p < 0.01) despite several changes in the interventional cardiologists who made up the team over this time period. CONCLUSION: Introduction of technical and clinical practice changes can result in a significant reduction in radiation exposure for patients and providers in a paediatric cardiac catheterisation laboratory. These reductions can be maintained over the long term.

Predicting Injury and Illness with Machine Learning in Elite Youth Soccer: A Comprehensive Monitoring Approach over 3 Months.

The search for monitoring tools that provide early indication of injury and illness could contribute to better player protection. The aim of the present study was to i) determine the feasibility of and adherence to our monitoring approach, and ii) identify variables associated with up-coming illness and injury. We incorporated a comprehensive set of monitoring tools consisting of external load and physical fitness data, questionnaires, blood, neuromuscular-, hamstring, hip abductor and hip adductor performance tests performed over a three-month period in elite under-18 academy soccer players. Twenty-five players (age: 16.6 ± 0.9 years, height: 178 ± 7 cm, weight: 74 ± 7 kg, VO2max: 59 ± 4 ml/min/kg) took part in the study. In addition to evaluating adherence to the monitoring approach, data were analyzed using a linear support vector machine (SVM) to predict illness and injuries. The approach was feasible, with no injuries or dropouts due to the monitoring process. Questionnaire adherence was high at the beginning and decreased steadily towards the end of the study. An SVM resulted in the best classification results for three classification tasks, i.e., illness prediction, illness determination and injury prediction. For injury prediction, one of four injuries present in the test data set was detected, with 96.3% of all data points (i.e., injuries and non-injuries) correctly detected. For both illness prediction and determination, there was only one illness in the test data set that was detected by the linear SVM. However, the model showed low precision for injury and illness prediction with a considerable number of false-positives. The results demonstrate the feasibility of a holistic monitoring approach with the possibility of predicting illness and injury. Additional data points are needed to improve the prediction models. In practical application, this may lead to overcautious recommendations on when players should be protected from injury and illness.

A Dengue Virus Serotype 1 mRNA-LNP Vaccine Elicits Protective Immune Responses.

Dengue virus (DENV) is the most common vector-borne viral disease, with nearly 400 million worldwide infections each year concentrated in the tropical and subtropical regions of the world. Severe dengue complications are often associated with a secondary heterotypic infection of one of the four circulating serotypes. In this scenario, humoral immune responses targeting cross-reactive, poorly neutralizing epitopes can lead to increased infectivity of susceptible cells via antibody-dependent enhancement (ADE). In this way, antibodies produced in response to infection or vaccination are capable of contributing to enhanced disease in subsequent infections. Currently, there are no available therapeutics to combat DENV disease, and there is an urgent need for a safe and efficacious vaccine. Here, we developed a nucleotide-modified mRNA vaccine encoding the membrane and envelope structural proteins from DENV serotype 1 encapsulated in lipid nanoparticles (prM/E mRNA-LNP). Vaccination of mice elicited robust antiviral immune responses comparable to viral infection, with high levels of neutralizing antibody titers and antiviral CD4+ and CD8+ T cells. Immunocompromised AG129 mice vaccinated with the prM/E mRNA-LNP vaccine were protected from a lethal DENV challenge. Vaccination with either a wild-type vaccine or a vaccine with mutations in the immunodominant fusion loop epitope elicited equivalent humoral and cell-mediated immune responses. Neutralizing antibodies elicited by the vaccine were sufficient to protect against a lethal challenge. Both vaccine constructs demonstrated serotype-specific immunity with minimal serum cross-reactivity and reduced ADE in comparison to a live DENV1 viral infection.IMPORTANCE With 400 million worldwide infections each year, dengue is the most common vector-borne viral disease. Forty percent of the world's population is at risk, with dengue experiencing consistent geographic spread over the years. With no therapeutics available and vaccines performing suboptimally, the need for an effective dengue vaccine is urgent. Here, we develop and characterize a novel mRNA vaccine encoding the dengue serotype 1 envelope and premembrane structural proteins that is delivered via a lipid nanoparticle. Our DENV1 prM/E mRNA-LNP vaccine induces neutralizing antibody and cellular immune responses in immunocompetent mice and protects an immunocompromised mouse from a lethal DENV challenge. Existing antibodies against dengue can enhance subsequent infections via antibody-dependent enhancement (ADE). Importantly our vaccine induced only serotype-specific immune responses and did not induce ADE.

Prior Heterologous Flavivirus Exposure Results in Reduced Pathogenesis in a Mouse Model of Zika Virus Infection.

The 2015/2016 Zika virus epidemic in South and Central America left the scientific community urgently trying to understand the factors that contribute to Zika virus pathogenesis. Because multiple other flaviviruses are endemic in areas where Zika virus emerged, it is hypothesized that a key to understanding Zika virus disease severity is to study Zika virus infection in the context of prior flavivirus exposure. Human and animal studies have highlighted the idea that having been previously exposed to a different flavivirus may modulate the immune response to Zika virus. However, it is still unclear how prior flavivirus exposure impacts Zika viral burden and disease. In this murine study, we longitudinally examine multiple factors involved in Zika disease, linking viral burden with increased neurological disease severity, weight loss, and inflammation. We show that prior heterologous flavivirus exposure with dengue virus type 2 or 3 or the vaccine strain of yellow fever provides protection from mortality in a lethal Zika virus challenge. However, reduction in viral burden and Zika disease varies depending on the infecting primary flavivirus; with primary Zika virus infection being most protective from Zika virus challenge, followed by dengue virus 2, with yellow fever and dengue virus 3 protecting against mortality but showing more severe disease. This study demonstrates the variation in protective effects of prior flavivirus exposure on Zika virus pathogenesis and identifies distinct relationships between primary flavivirus infection and the potential for Zika virus disease. IMPORTANCE The emergence and reemergence of various vector-borne diseases in recent years highlights the need to understand the mechanisms of protection for each pathogen. In this study, we investigated the impact of prior exposure to Zika virus, dengue virus serotypes 2 or 3, or the vaccine strain of yellow fever on pathogenesis and disease outcomes in a mouse model of Zika virus infection. We found that prior exposure to a heterologous flavivirus was protective from mortality, and to varying degrees, prior flavivirus exposure was protective against neurological disease, weight loss, and severe viral burden during a lethal Zika challenge. Using a longitudinal and cross-sectional study design, we were able to link multiple disease parameters, including viral burden, with neurological disease severity, weight loss, and inflammatory response in the context of flavivirus infection. This study demonstrates a measurable but varied impact of prior flavivirus exposure in modulating flavivirus pathophysiology. Given the cyclic nature of most flavivirus outbreaks, this work will contribute to the forecasting of disease severity for future outbreaks.

Pre-existing T Cell Memory against Zika Virus.

The mosquito-borne Zika virus (ZIKV) has spread rapidly into regions where dengue virus (DENV) is endemic, and flavivirus cross-reactive T cell responses have been observed repeatedly in animal models and in humans. Preexisting cellular immunity to DENV is thought to contribute to protection in subsequent ZIKV infection, but the epitope targets of cross-reactive T cell responses have not been comprehensively identified. Using human blood samples from the regions of Nicaragua and Sri Lanka where DENV is endemic that were collected before the global spread of ZIKV in 2016, we employed an in vitro expansion strategy to map ZIKV T cell epitopes in ZIKV-unexposed, DENV-seropositive donors. We identified 93 epitopes across the ZIKV proteome, and we observed patterns of immunodominance that were dependent on antigen size and sequence identity to DENV. We confirmed the immunogenicity of these epitopes through a computational HLA binding analysis, and we showed that cross-reactive T cells specifically recognize ZIKV peptides homologous to DENV sequences. We also found that these CD4 responses were derived from the memory T cell compartment. These data have implications for understanding the dynamics of flavivirus-specific T cell immunity in areas of endemicity.IMPORTANCE Multiple flaviviruses, including Zika virus (ZIKV) and the four serotypes of dengue virus (DENV), are prevalent in the same large tropical and equatorial areas, which are inhabited by hundreds of millions of people. The interplay of DENV and ZIKV infection is especially relevant, as these two viruses are endemic in largely overlapping regions, have significant sequence similarity, and share the same arthropod vector. Here, we define the targets of preexisting immunity to ZIKV in unexposed subjects in areas where dengue is endemic. We demonstrate that preexisting immunity to DENV could shape ZIKV-specific responses, and DENV-ZIKV cross-reactive T cell populations can be expanded by stimulation with ZIKV peptides. The issue of potential ZIKV and DENV cross-reactivity is of relevance for understanding patterns of natural immunity, as well as for the development of diagnostic tests and vaccines.

SARS-CoV-2 spike protein promotes IL-6 trans-signaling by activation of angiotensin II receptor signaling in epithelial cells.

Cytokine storm is suggested as one of the major pathological characteristics of SARS-CoV-2 infection, although the mechanism for initiation of a hyper-inflammatory response, and multi-organ damage from viral infection is poorly understood. In this virus-cell interaction study, we observed that SARS-CoV-2 infection or viral spike protein expression alone inhibited angiotensin converting enzyme-2 (ACE2) receptor protein expression. The spike protein promoted an angiotensin II type 1 receptor (AT1) mediated signaling cascade, induced the transcriptional regulatory molecules NF-κB and AP-1/c-Fos via MAPK activation, and increased IL-6 release. SARS-CoV-2 infected patient sera contained elevated levels of IL-6 and soluble IL-6R. Up-regulated AT1 receptor signaling also influenced the release of extracellular soluble IL-6R by the induction of the ADAM-17 protease. Use of the AT1 receptor antagonist, Candesartan cilexetil, resulted in down-regulation of IL-6/soluble IL-6R release in spike expressing cells. Phosphorylation of STAT3 at the Tyr705 residue plays an important role as a transcriptional inducer for SOCS3 and MCP-1 expression. Further study indicated that inhibition of STAT3 Tyr705 phosphorylation in SARS-CoV-2 infected and viral spike protein expressing epithelial cells did not induce SOCS3 and MCP-1 expression. Introduction of culture supernatant from SARS-CoV-2 spike expressing cells on a model human liver endothelial Cell line (TMNK-1), where transmembrane IL-6R is poorly expressed, resulted in the induction of STAT3 Tyr705 phosphorylation as well as MCP-1 expression. In conclusion, our results indicated that the presence of SARS-CoV-2 spike protein in epithelial cells promotes IL-6 trans-signaling by activation of the AT1 axis to initiate coordination of a hyper-inflammatory response.

mRNA induced expression of human angiotensin-converting enzyme 2 in mice for the study of the adaptive immune response to severe acute respiratory syndrome coronavirus 2.

The novel human coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a pandemic. Critical to the rapid evaluation of vaccines and antivirals against SARS-CoV-2 is the development of tractable animal models to understand the adaptive immune response to the virus. To this end, the use of common laboratory strains of mice is hindered by significant divergence of the angiotensin-converting enzyme 2 (ACE2), which is the receptor required for entry of SARS-CoV-2. In the current study, we designed and utilized an mRNA-based transfection system to induce expression of the hACE2 receptor in order to confer entry of SARS-CoV-2 in otherwise non-permissive cells. By employing this expression system in an in vivo setting, we were able to interrogate the adaptive immune response to SARS-CoV-2 in type 1 interferon receptor deficient mice. In doing so, we showed that the T cell response to SARS-CoV-2 is enhanced when hACE2 is expressed during infection. Moreover, we demonstrated that these responses are preserved in memory and are boosted upon secondary infection. Importantly, using this system, we functionally identified the CD4+ and CD8+ structural peptide epitopes targeted during SARS-CoV-2 infection in H2b restricted mice and confirmed their existence in an established model of SARS-CoV-2 pathogenesis. We demonstrated that, identical to what has been seen in humans, the antigen-specific CD8+ T cells in mice primarily target peptides of the spike and membrane proteins, while the antigen-specific CD4+ T cells target peptides of the nucleocapsid, membrane, and spike proteins. As the focus of the immune response in mice is highly similar to that of the humans, the identification of functional murine SARS-CoV-2-specific T cell epitopes provided in this study will be critical for evaluation of vaccine efficacy in murine models of SARS-CoV-2 infection.

T Cell Responses Induced by Attenuated Flavivirus Vaccination Are Specific and Show Limited Cross-Reactivity with Other Flavivirus Species.

Members of the flavivirus genus share a high level of sequence similarity and often circulate in the same geographical regions. However, whether T cells induced by one viral species cross-react with other related flaviviruses has not been globally addressed. In this study, we tested pools of epitopes derived from dengue (DENV), Zika (ZIKV), Japanese encephalitis (JEV), West Nile (WNV), and yellow fever (YFV) viruses by intracellular cytokine staining (ICS) using peripheral blood mononuclear cells (PBMCs) of individuals naturally exposed to DENV or immunized with DENV (TV005) or YF17D vaccine. CD8 T cell responses recognized epitopes from multiple flaviviruses; however, the magnitude of cross-reactive responses was consistently severalfold lower than those to the autologous epitope pools and was associated with lower expression of activation markers such as CD40L, CD69, and CD137. Next, we characterized the antigen sensitivity of short-term T cell lines (TCL) representing 29 different individual epitope/donor combinations. TCL derived from DENV monovalent vaccinees induced CD8 and CD4 T cells that cross-reacted within the DENV serocomplex but were consistently associated with >100-fold-lower antigen sensitivity for most other flaviviruses, with no cross-recognition of YFV-derived peptides. CD8 and CD4 TCL from YF17D vaccinees were associated with very limited cross-reactivity with any other flaviviruses and in five out of eight cases >1,000-fold-lower antigen sensitivity. Overall, our data suggest limited cross-reactivity for both CD4 and CD8 T cell responses between flaviviruses and have implications for understanding immunity elicited by natural infection and strategies to develop live attenuated vaccines against flaviviral species.IMPORTANCE The envelope (E) protein is the dominant target of neutralizing antibodies for dengue virus (DENV) and yellow fever virus (YFV). Accordingly, several DENV vaccine constructs use the E protein in a live attenuated vaccine format, utilizing a backbone derived from a heterologous flavivirus (such as YF) as a delivery vector. This backbone comprises the nonstructural (NS) and capsid (C) antigens, which are dominant targets of T cell responses. Here, we demonstrate that cross-reactivity at the level of T cell responses among different flaviviruses is very limited, despite high levels of sequence homology. Thus, the use of heterologous flavivirus species as a live attenuated vaccine vector is not likely to generate optimal T cell responses and might thus impair vaccine performance.

Selective estrogen receptor modulator, tamoxifen, inhibits Zika virus infection.

Zika virus (ZIKV) is an arbovirus belonging to the flaviviridae family with a risk assessment that has been increasing in recent years and was labeled a global health emergency by the World Health Organization in 2016. There are currently no Food and Drug Administration-approved treatment options available for ZIKV, so expeditious development of treatment options is urgent. To expedite this process, an on-market drug, tamoxifen (TAM), was selected as a promising candidate for repurposing due to its wide range of biological activities and because it has already been shown to possess activity against hepatitis C virus, a flavivirus in a separate genus. Anti-ZIKV activity of TAM was assessed by compound screens using an infectious virus and mechanistic details were gleaned from time of addition and virucidal studies. TAM and an active metabolite, 4-hydroxytamoxifen (TAM-OH), both showed promising antiviral activity (EC50 ≈9 and 5 µM, respectively) in initial compound screening and up to 8-h postinfection, though the virucidal assay indicated that they do not possess any direct virucidal activity. Additionally, TAM was assessed for its activity against ZIKV in the human male germ cell line, SEM-1, due to the sexually transmitted nature of ZIKV owing to its extended survival times in germ cells. Virus titers show diminished replication of ZIKV over 7 days compared to controls. These data indicate that TAM has the potential to be repurposed as an anti-ZIKV therapeutic and warrants further investigation.

Enhanced Recovery after Surgery for Cesarean Delivery Decreases Length of Hospital Stay and Opioid Consumption: A Quality Improvement Initiative.

OBJECTIVE: The aim of this study is to assess the effect of a resident-led enhanced recovery after surgery (ERAS) protocol for scheduled prelabor cesarean deliveries on hospital length of stay and postpartum opioid consumption. STUDY DESIGN: This retrospective cohort study included patients who underwent scheduled prelabor cesarean deliveries before and after implementation of an ERAS protocol at a single academic tertiary care institution. The primary outcome was length of stay following cesarean delivery. Secondary outcomes included protocol adherence, inpatient opioid consumption, and patient-centered outcomes. The protocol included multimodal analgesia and antiemetic medications, expedited urinary catheter removal, early discontinuation of maintenance intravenous fluids, and early ambulation. RESULTS: A total of 250 patients were included in the study: 122 in the pre-ERAS cohort and 128 in the post-ERAS cohort. There were no differences in baseline demographics, medical comorbidities, or cesarean delivery characteristics between the two groups. Following protocol implementation, hospital length of stay decreased by an average of 7.9 hours (pre-ERAS 82.1 vs. post-ERAS 74.2, p < 0.001). There was 89.8% adherence to the entire protocol as written. Opioid consumption decreased by an average of 36.5 mg of oxycodone per patient, with no significant differences in pain scores from postoperative day 1 to postoperative day 4 (all p > 0.05). CONCLUSION: A resident-driven quality improvement project was associated with decreased length of hospital stay, decreased opioid consumption, and unchanged visual analog pain scores at the time of hospital discharge. Implementation of this ERAS protocol is feasible and effective. KEY POINTS: · Enhanced recovery after surgery (ERAS) principles can be effectively applied to cesarean delivery with excellent protocol adherence.. · Patients who participated in the ERAS pathway had significant decreases in hospital length of stay and opioid pain medication consumption with unchanged visual analog pain scores postoperative days 1 through 4.. · Resident-driven quality improvement projects can make a substantial impact in patient care for both process measures (e.g., protocol adherence) and outcome measures (e.g., opioid use)..

Post-operative arrhythmias in patients with hypoplastic left heart syndrome and anatomic variants: incidence, type, and course.

BACKGROUND: Arrhythmias are common in the post-operative course of patients with hypoplastic left heart syndrome. We sought to determine the types, incidence, risk factors, and impact of arrhythmias in patients with HLHS and anatomic variants. METHODS: We performed a retrospective chart review of 120 consecutive patients with HLHS and anatomical variants, who had single-ventricle palliation at our institution from January, 2006 to December, 2016. RESULTS: A total of thirty-one patients (26%) had 37 episodes of arrhythmias over a median follow-up period of 3.5 years. Of the 37 episodes, 12 (32.4%) were ectopic atrial tachycardia, 9 (24.3%) were paroxysmal supraventricular tachycardia, 4 (10.8%) were junctional ectopic tachycardia, 5 (13.6%) were sinus node dysfunction, 3 (8.1%) were heart block, 2 (5.4%) were atrial flutter, and 2 (5.4%) were ventricular tachycardia. Twenty-four (65%) of the arrhythmias occurred at post-stage 1 surgery. Most (64.8%) of the arrhythmias were resolved. Arrhythmias that occurred at post-stage 1 surgery were more likely to resolve compared to post-stages 2 or 3 (p = 0.006). No anatomical, surgical, or clinical variables were associated with arrhythmia except for age (OR per unit decrease in age at stage 1 palliation: 1.12 (95% CI 1.003, 1.250); p = 0.0439). Arrhythmias were not associated with length of hospital stay or mortality. CONCLUSION: Arrhythmias are common in patients with HLHS and anatomic variants, with EAT and PSVT being the most common types. Arrhythmias were associated with younger age at surgery, but did not affect mortality or length of hospital stay.