Peripheral blood mononuclear cell tissue factor (F3 gene) transcript levels and circulating extracellular vesicles are elevated in severe coronavirus 2019 (COVID-19) disease.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with excessive coagulation, thrombosis, and mortality. OBJECTIVE: To provide insight into mechanisms that contribute to excessive coagulation in coronavirus 2019 (COVID-19) disease. PATIENTS/METHODS: Blood from COVID-19 patients was investigated for coagulation-related gene expression and functional activities. RESULTS: Single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells from severe COVID-19 patients revealed a 5.2-fold increase in tissue factor (TF [F3 gene]) transcript expression levels (P < .05), the trigger of extrinsic coagulation; a 7.7-fold increase in C1-inhibitor (SERPING1 gene; P < .01) transcript expression levels, an inhibitor of intrinsic coagulation; and a 4.4-fold increase in anticoagulant thrombomodulin (TM [THBD gene]) transcript expression levels (P < .001). Bulk RNA-seq analysis of sorted CD14+ monocytes on an independent cohort of COVID-19 patients confirmed these findings (P < .05). Indicative of excessive coagulation, 41% of COVID-19 patients' plasma samples contained high D-dimer levels (P < .0001); of these, 19% demonstrated extracellular vesicle TF activity (P = .109). COVID-19 patients' ex vivo plasma-based thrombin generation correlated positively with D-dimer levels (P < .01). Plasma procoagulant extracellular vesicles were elevated ∼9-fold in COVID-19 patients (P < .01). Public scRNA-seq data sets from bronchoalveolar lung fluid and our peripheral blood mononuclear cell scRNA-seq data show CD14+ monocytes/macrophages TF transcript expression levels are elevated in severe but not mild or moderate COVID-19 patients. CONCLUSIONS: Beyond local lung injury, SARS-CoV-2 infection increases systemic TF (F3) transcript levels and elevates circulating extracellular vesicles that likely contribute to disease-associated coagulation, thrombosis, and related mortality.

COVID-19 disease and immune dysregulation.

The SARS-CoV-2 virus has complex and divergent immune alterations in differing hosts and over disease evolution. Much of the nuanced COVID-19 disease immune dysregulation was originally dominated by innate cytokine changes, which has since been replaced with a more complex picture of innate and adaptive changes characterized by simultaneous hyperinflammatory and immunosuppressive phenomena in effector cells. These intricacies are summarized in this review as well as potential relevance from acute infection to a multisystem inflammatory syndrome commonly seen in children. Additional consideration is made for the influence of variant to variant host cellular changes and the impact of potential vaccination upon these phenotypes. Finally, therapeutic benefit for immune alterations are discussed.

Characteristics and Outcomes of Critically Ill Children With Multisystem Inflammatory Syndrome.

OBJECTIVES: To characterize the prevalence of pediatric critical illness from multisystem inflammatory syndrome in children (MIS-C) and to assess the influence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strain on outcomes. DESIGN: Retrospective cohort study. SETTING: Database evaluation using the Virtual Pediatric Systems Database. PATIENTS: All children with MIS-C admitted to the PICU in 115 contributing hospitals between January 1, 2020, and June 30, 2021. MEASUREMENTS AND MAIN RESULTS: Of the 145,580 children admitted to the PICU during the study period, 1,338 children (0.9%) were admitted with MIS-C with the largest numbers of children admitted in quarter 1 (Q1) of 2021 ( n = 626). The original SARS-CoV-2 viral strain and the D614G Strain were the predominant strains through 2020, with Alpha B.1.1.7 predominating in Q1 and quarter 2 (Q2) of 2021. Overall, the median PICU length of stay (LOS) was 2.7 days (25-75% interquartile range [IQR], 1.6-4.7 d) with a median hospital LOS of 6.6 days (25-75% IQR, 4.7-9.3 d); 15.2% received mechanical ventilation with a median duration of mechanical ventilation of 3.1 days (25-75% IQR, 1.9-5.8 d), and there were 11 hospital deaths. During the study period, there was a significant decrease in the median PICU and hospital LOS and a decrease in the frequency of mechanical ventilation, with the most significant decrease occurring between quarter 3 and quarter 4 (Q4) of 2020. Children admitted to a PICU from the general care floor or from another ICU/step-down unit had longer PICU LOS than those admitted directly from an emergency department. CONCLUSIONS: Overall mortality from MIS-C was low, but the disease burden was high. There was a peak in MIS-C cases during Q1 of 2021, following a shift in viral strains in Q1 of 2021. However, an improvement in MIS-C outcomes starting in Q4 of 2020 suggests that viral strain was not the driving factor for outcomes in this population.

School Closures in the United States and Severe Respiratory Illnesses in Children: A Normalized Nationwide Sample.

OBJECTIVES: To determine the association between nationwide school closures and prevalence of common admission diagnoses in the pediatric critical care unit. DESIGN: Retrospective cohort study. SETTING: National database evaluation using the Virtual Pediatric Systems LLC database. PATIENTS: All patients admitted to the PICU in 81 contributing hospitals in the United States. MEASUREMENTS AND MAIN RESULTS: Diagnosis categories were determined for all 110,418 patients admitted during the 20-week study period in each year (2018, 2019, and 2020). Admission data were normalized relative to statewide school closure dates for each patient using geographic data. The "before school closure" epoch was defined as 8 weeks prior to school closure, and the "after school closure" epoch was defined as 12 weeks following school closure. For each diagnosis, admission ratios for each study day were calculated by dividing 2020 admissions by 2018-2019 admissions. The 10 most common diagnosis categories were examined. Significant changes in admission ratios were identified for bronchiolitis, pneumonia, and asthma. These changes occurred at 2, 8, and 35 days following school closure, respectively. PICU admissions decreased by 82% for bronchiolitis, 76% for pneumonia, and 76% for asthma. Nonrespiratory diseases such as diabetic ketoacidosis, status epilepticus, traumatic injury, and poisoning/ingestion did not show significant changes following school closure. CONCLUSIONS: School closures are associated with a dramatic reduction in the prevalence of severe respiratory disease requiring PICU admission. School closure may be an effective tool to mitigate future pandemics but should be balanced with potential academic, economic, mental health, and social consequences.

Dysregulation of the leukocyte signaling landscape during acute COVID-19.

The global COVID-19 pandemic has claimed the lives of more than 750,000 US citizens. Dysregulation of the immune system underlies the pathogenesis of COVID-19, with inflammation mediated tissue injury to the lung in the setting of suppressed systemic immune function. To define the molecular mechanisms of immune dysfunction in COVID-19 we utilized a systems immunology approach centered on the circulating leukocyte phosphoproteome measured by mass cytometry. We find that although COVID-19 is associated with wholesale activation of a broad set of signaling pathways across myeloid and lymphoid cell populations, STAT3 phosphorylation predominated in both monocytes and T cells. STAT3 phosphorylation was tightly correlated with circulating IL-6 levels and high levels of phospho-STAT3 was associated with decreased markers of myeloid cell maturation/activation and decreased ex-vivo T cell IFN-γ production, demonstrating that during COVID-19 dysregulated cellular activation is associated with suppression of immune effector cell function. Collectively, these data reconcile the systemic inflammatory response and functional immunosuppression induced by COVID-19 and suggest STAT3 signaling may be the central pathophysiologic mechanism driving immune dysfunction in COVID-19.

The Temporal Relationship Between Local School Closure and Increased Incidence of Pediatric Diabetic Ketoacidosis.

Importance: The incidence of pediatric diabetic ketoacidosis (DKA) increased early in the COVID-19 pandemic, but the relative contribution of behavioral changes and viral-related pathophysiology are unknown. Objective: To evaluate the relationship between school closure date and onset of increased DKA to help clarify the etiology of the increased incidence. Design: A multi-center, quality-controlled Pediatric Intensive Care Unit (PICU) database was used to identify the number of admissions to a participating PICU with DKA on each calendar day from 60 days before local school closure to 90 days after, and compared to baseline data from the same periods in 2018-2019. Interrupted time series and multiple linear regression analyses were used to identify admission rates that differed significantly between 2020 and baseline. Setting: Eighty-one PICUs in the United StatesParticipants: Children ages 29 days to 17 years admitted to a PICU with DKAExposures: Statewide school closureMain outcome/measure: Rate of admission to the PICU for DKA. Results: There were 1936 admissions for children with DKA in 2020 and 1795 admissions/year to those same PICUs in 2018-2019. Demographics and clinical outcomes did not differ before school closure, but pandemic-era patients were less often white and had longer hospital length of stay in the post-school closure period. The difference between 2020 admissions and 2018-2019 admissions was not different than zero before school closure, and significantly higher than zero after school closure, but was significantly increased in 2020 at >30 days after school closure (p = 0.039). Conclusions/Relevance: An increase in pediatric DKA admissions began one month after school closures. Given that behavioral changes started near school closure dates and viral activity peaked weeks after, this suggests that behavioral factors may not be the primary etiology and it is possible that SARS-CoV-2 infection may have direct effects on pediatric DKA.

Overlapping but Disparate Inflammatory and Immunosuppressive Responses to SARS-CoV-2 and Bacterial Sepsis: An Immunological Time Course Analysis.

Both severe SARS-CoV-2 infections and bacterial sepsis exhibit an immunological dyscrasia and propensity for secondary infections. The nature of the immunological dyscrasias for these differing etiologies and their time course remain unclear. In this study, thirty hospitalized patients with SARS-CoV-2 infection were compared with ten critically ill patients with bacterial sepsis over 21 days, as well as ten healthy control subjects. Blood was sampled between days 1 and 21 after admission for targeted plasma biomarker analysis, cellular phenotyping, and leukocyte functional analysis via enzyme-linked immunospot assay. We found that circulating inflammatory markers were significantly higher early after bacterial sepsis compared with SARS-CoV-2. Both cohorts exhibited profound immune suppression through 21 days (suppressed HLA-DR expression, reduced mononuclear cell IFN-gamma production), and expanded numbers of myeloid-derived suppressor cells (MDSCs). In addition, MDSC expansion and ex vivo production of IFN-gamma and TNF-alpha were resolving over time in bacterial sepsis, whereas in SARS-CoV-2, immunosuppression and inflammation were accelerating. Despite less severe initial physiologic derangement, SARS-CoV-2 patients had similar incidence of secondary infections (23% vs 30%) as bacterial sepsis patients. Finally, COVID patients who developed secondary bacterial infections exhibited profound immunosuppression evident by elevated sPD-L1 and depressed HLA-DR. Although both bacterial sepsis and SARS-CoV-2 are associated with inflammation and immune suppression, their immune dyscrasia temporal patterns and clinical outcomes are different. SARS-CoV-2 patients had less severe early inflammation and organ dysfunction but had persistent inflammation and immunosuppression and suffered worse clinical outcomes, especially when SARS-CoV-2 infection was followed by secondary bacterial infection.

Interleukin-7 Reverses Lymphopenia and Improves T-Cell Function in Coronavirus Disease 2019 Patient With Inborn Error of Toll-Like Receptor 3: A Case Report.

BACKGROUND: Immunotherapy treatment for coronavirus disease 2019 combined with antiviral therapy and supportive care remains under intense investigation. However, the capacity to distinguish patients who would benefit from immunosuppressive or immune stimulatory therapies remains insufficient. Here, we present a patient with severe coronavirus disease 2019 with a defective immune response, treated successfully with interleukin-7 on compassionate basis with resultant improved adaptive immune function. CASE SUMMARY: A previously healthy 43-year-old male developed severe acute respiratory distress syndrome due to the severe acute respiratory syndrome coronavirus 2 virus with acute hypoxemic respiratory failure and persistent, profound lymphopenia. Functional analysis demonstrated depressed lymphocyte function and few antigen-specific T cells. Interleukin-7 administration resulted in reversal of lymphopenia and improved T-cell function. Respiratory function and clinical status rapidly improved, and he was discharged home. Whole exome sequencing identified a deleterious autosomal dominant mutation in TICAM1, associated with a dysfunctional type I interferon antiviral response with increased severity of coronavirus disease 2019 disease. CONCLUSIONS: Immunoadjuvant therapies to boost host immunity may be efficacious in life-threatening severe coronavirus disease 2019 infections, particularly by applying a precision medicine approach in selecting patients expressing an immunosuppressive phenotype.

Changes in Pediatric ICU Utilization and Clinical Trends During the Coronavirus Pandemic.

BACKGROUND: Children have been less affected by the COVID-19 pandemic, but its repercussions on pediatric illnesses may have been significant. This study examines the indirect impact of the pandemic on a population of critically ill children in the United States. RESEARCH QUESTION: Were there significantly fewer critically ill children admitted to PICUs during the second quarter of 2020, and were there significant changes in the types of diseases admitted? STUDY DESIGN AND METHODS: This retrospective observational cohort study used the Virtual Pediatric Systems database. Participants were 160,295 children admitted to the PICU at 77 sites in the United States during quarters 1 (Q1) and 2 (Q2) of 2017 to 2019 (pre-COVID-19) and 2020 (COVID-19). RESULTS: The average number of admissions was similar between pre-COVID-19 Q1 and COVID-19 Q1 but decreased by 32% from pre-COVID-19 Q2 to COVID-19 Q2 (20,157 to 13,627 admissions per quarter). The largest decreases were in respiratory conditions, including asthma (1,327 subjects in pre-COVID-19 Q2 (6.6% of patients) vs 241 subjects in COVID-19 Q2 (1.8%; P < .001) and bronchiolitis (1,299 [6.5%] vs 121 [0.9%]; P < .001). The percentage of trauma admissions increased, although the raw number of trauma admissions decreased. Admissions for diabetes mellitus and poisoning/ingestion also increased. In the multivariable model, illness severity-adjusted odds of ICU mortality for PICU patients during COVID-19 Q2 increased compared with pre-COVID-19 Q2 (OR, 1.165; 95% CI, 1.00-1.357; P = .049). INTERPRETATION: Pediatric critical illness admissions decreased substantially during the second quarter of 2020, with significant changes in the types of diseases seen in PICUs in the United States. There was an increase in mortality in children admitted to the PICU during this period.

In Vitro-Administered Dexamethasone Suppresses T Cell Function With Reversal by Interleukin-7 in Coronavirus Disease 2019.

OBJECTIVES: Corticosteroid therapy has become standard of care therapy for hospitalized patients infected with the severe acute respiratory syndrome coronavirus-2 global pandemic-causing virus. Whereas systemic inflammation is a notably important feature in coronavirus disease 2019 pathogenesis, adaptive immune suppression and the inability to eradicate effectively the virus remain significant factors as well. We sought to evaluate the in vitro effects of dexamethasone phosphate on T cell function in peripheral blood mononuclear cells derived from patients with acute, severe, and moderate coronavirus disease 2019. DESIGN: Prospective in vitro laboratory study. SETTING: Coronavirus disease 2019-specific medical wards and ICUs at a single-center, quaternary-care academic hospital between October 1, 2020, and November 15, 2020. PATIENTS: Eleven patients diagnosed with coronavirus disease 2019 admitted to either the ICU or hospital coronavirus disease 2019 unit. Three patients had received at least one dose of dexamethasone prior to enrollment. INTERVENTIONS: Fresh whole blood was collected, and peripheral blood mononuclear cells were immediately isolated and plated onto precoated enzyme-linked immunospot plates for detection of interferon-γ production. Samples were incubated with CD3/CD28 antibodies alone and with three concentrations of dexamethasone. These conditions were also stimulated with recombinant human interleukin-7. Following overnight incubation, the plates were washed and stained for analysis using Cellular Technology Limited ImmunoSpot S6 universal analyzer (ImmunoSpot by Cellular Technology Limited, Cleveland, OH). MEASUREMENTS AND MAIN RESULTS: Functional cytokine production was assessed by quantitation of cell spot number and total well intensity after calculation for each enzyme-linked immunospot well using the Cellular Technology Limited ImmunoSpot Version 7.0 professional software (CTL Analyzers, Shaker Heights, OH). Comparisons were made using t test and using a nonparametric analysis of variance Friedman test. The number of functional T cells producing interferon-γ and the intensity of the response decrease significantly with exposure to 1.2-µg/mL dexamethasone. About 0.12 µg/mL does not significantly affect the functional immune response on enzyme-linked immunospot. Interleukin-7 increases the overall number of activated T cells, including those exposed to dexamethasone. CONCLUSIONS: Further evaluation of the effect of immunomodulatory therapies is warranted in coronavirus disease 2019. A refined functional, precision medicine approach that evaluates the cellular immune function of individual patients with coronavirus disease 2019 is needed to better define which therapies could have benefit or cause harm for specific patients.

Severe Acute Respiratory Syndrome-Associated Coronavirus 2 Infection and Organ Dysfunction in the ICU: Opportunities for Translational Research.

OBJECTIVES: Since the beginning of the coronavirus disease 2019 pandemic, hundreds of thousands of patients have been treated in ICUs across the globe. The severe acute respiratory syndrome-associated coronavirus 2 virus enters cells via the angiotensin-converting enzyme 2 receptor and activates several distinct inflammatory pathways, resulting in hematologic abnormalities and dysfunction in respiratory, cardiac, gastrointestinal renal, endocrine, dermatologic, and neurologic systems. This review summarizes the current state of research in coronavirus disease 2019 pathophysiology within the context of potential organ-based disease mechanisms and opportunities for translational research. DATA SOURCES: Investigators from the Research Section of the Society of Critical Care Medicine were selected based on expertise in specific organ systems and research focus. Data were obtained from searches conducted in Medline via the PubMed portal, Directory of Open Access Journals, Excerpta Medica database, Latin American and Caribbean Health Sciences Literature, and Web of Science from an initial search from December 2019 to October 15, 2020, with a revised search to February 3, 2021. The medRxiv, Research Square, and clinical trial registries preprint servers also were searched to limit publication bias. STUDY SELECTION: Content experts selected studies that included mechanism-based relevance to the severe acute respiratory syndrome-associated coronavirus 2 virus or coronavirus disease 2019 disease. DATA EXTRACTION: Not applicable. DATA SYNTHESIS: Not applicable. CONCLUSIONS: Efforts to improve the care of critically ill coronavirus disease 2019 patients should be centered on understanding how severe acute respiratory syndrome-associated coronavirus 2 infection affects organ function. This review articulates specific targets for further research.

Distinct inflammatory profiles distinguish COVID-19 from influenza with limited contributions from cytokine storm.

We pursued a study of immune responses in coronavirus disease 2019 (COVID-19) and influenza patients. Compared to patients with influenza, patients with COVID-19 exhibited largely equivalent lymphocyte counts, fewer monocytes, and lower surface human leukocyte antigen (HLA)-class II expression on selected monocyte populations. Furthermore, decreased HLA-DR on intermediate monocytes predicted severe COVID-19 disease. In contrast to prevailing assumptions, very few (7 of 168) patients with COVID-19 exhibited cytokine profiles indicative of cytokine storm syndrome. After controlling for multiple factors including age and sample time point, patients with COVID-19 exhibited lower cytokine levels than patients with influenza. Up-regulation of IL-6, G-CSF, IL-1RA, and MCP1 predicted death in patients with COVID-19 but were not statistically higher than patients with influenza. Single-cell transcriptional profiling revealed profound suppression of interferon signaling among patients with COVID-19. When considered across the spectrum of peripheral immune profiles, patients with COVID-19 are less inflamed than patients with influenza.