John P. Reilly

Red Blood Cells Induce Necroptosis of Lung Endothelial Cells and Increase Susceptibility to Lung Inflammation

RATIONALE Red blood cell (RBC) transfusions are associated with increased risk of acute respiratory distress syndrome (ARDS) in the critically ill, yet the mechanisms for enhanced susceptibility to ARDS conferred by RBC transfusions remain unknown. OBJECTIVES To determine the mechanisms of lung endothelial cell (EC) High Mobility Group Box 1 (HMGB1) release following exposure to RBCs and to determine whether RBC transfusion increases susceptibility to lung inflammation in vivo through release of the danger signal HMGB1. METHODS In vitro studies examining human lung EC viability and HMGB1 release following exposure to allogenic RBCs were conducted under static conditions and using a microengineered model of RBC perfusion. The plasma from transfused and nontransfused patients with severe sepsis was examined for markers of cellular injury. A murine model of RBC transfusion followed by LPS administration was used to determine the effects of RBC transfusion and HMGB1 release on LPS-induced lung inflammation. MEASUREMENTS AND MAIN RESULTS After incubation with RBCs, lung ECs underwent regulated necrotic cell death (necroptosis) and released the essential mediator of necroptosis, receptor-interacting serine/threonine-protein kinase 3 (RIP3), and HMGB1. RIP3 was detectable in the plasma of patients with severe sepsis, and was increased with blood transfusion and among nonsurvivors of sepsis. RBC transfusion sensitized mice to LPS-induced lung inflammation through release of the danger signal HMGB1. CONCLUSIONS RBC transfusion enhances susceptibility to lung inflammation through release of HMGB1 and induces necroptosis of lung EC. Necroptosis and subsequent danger signal release is a novel mechanism of injury following transfusion that may account for the increased risk of ARDS in critically ill transfused patients.

Heterogeneous phenotypes of acute respiratory distress syndrome after major trauma.

RATIONALE Acute respiratory distress syndrome (ARDS) is a heterogeneous syndrome that can develop at various times after major trauma. OBJECTIVES To identify and characterize distinct phenotypes of ARDS after trauma, based on timing of syndrome onset. METHODS Latent class analyses were used to identify patterns of ARDS onset in a cohort of critically ill trauma patients. Identified patterns were tested for associations with known ARDS risk factors and associations were externally validated at a separate institution. Eleven plasma biomarkers representing pathophysiologic domains were compared between identified patterns in the validation cohort. MEASUREMENTS AND MAIN RESULTS Three patterns of ARDS were identified; class I (52%) early onset on Day 1 or 2, class II (40%) onset on Days 3 and 4, and class III (8%) later onset on Days 4 and 5. Early-onset ARDS was associated with higher Abbreviated Injury Scale scores for the thorax (P < 0.001), lower lowest systolic blood pressure before intensive care unit admission (P = 0.003), and a greater red blood cell transfusion requirement during resuscitation (P = 0.030). In the external validation cohort, early-onset ARDS was also associated with a higher Abbreviated Injury Scale score for the thorax (P = 0.001) and a lower lowest systolic blood pressure before intensive care unit enrollment (P = 0.006). In addition, the early-onset phenotype demonstrated higher plasma levels of soluble receptor for advanced glycation end-products and angiopoietin-2. CONCLUSIONS Degree of hemorrhagic shock and severity of thoracic trauma are associated with an early-onset phenotype of ARDS after major trauma. Lung injury biomarkers suggest a dominant alveolar-capillary barrier injury pattern in this phenotype.

ABO Blood Type A Is Associated With Increased Risk of ARDS in Whites Following Both Major Trauma and Severe Sepsis

BACKGROUND ABO glycosyltransferases catalyze antigen modifications on various glycans and glycoproteins and determine the ABO blood types. Blood type A has been associated with increased risk of vascular diseases and differential circulating levels of proteins related to inflammation and endothelial function. The objective of this study was to determine the association of ABO blood types with ARDS risk in patients with major trauma and severe sepsis. METHODS We conducted prospective cohort studies in two populations at an urban tertiary referral, level I trauma center. Critically ill patients (n 5 732) presenting after major trauma were followed for 5 days for ARDS development. Additionally, 976 medical patients with severe sepsis were followed for 5 days for ARDS. Multivariable logistic regression was used to adjust for confounders. RESULTS ARDS developed in 197 of the 732 trauma patients (27%). Blood type A was associated with increased ARDS risk among whites (37% vs 24%; adjusted OR, 1.88; 95% CI, 1.14-3.12; P 5 .014), but not blacks (adjusted OR, 0.61; 95% CI, 0.33-1.13; P=.114). ARDS developed in 222 of the 976 patients with severe sepsis (23%). Blood type A was also associated with an increased ARDS risk among whites (31% vs 21%; adjusted OR, 1.67; 95% CI, 1.08-2.59; P=.021) but, again, not among blacks (adjusted OR, 1.17; 95% CI, 0.59-2.33; P=.652). CONCLUSIONS Blood type A is associated with an increased risk of ARDS in white patients with major trauma and severe sepsis. These results suggest a role for ABO glycans and glycosyltransferases in ARDS susceptibility.

Computed tomography-defined abdominal adiposity is associated with acute kidney injury in critically ill trauma patients*.

Objectives:Higher body mass index is associated with increased risk of acute kidney injury after major trauma. Since body mass index is nonspecific, reflecting lean, fluid, and adipose mass, we evaluated the use of CT to determine if abdominal adiposity underlies the body mass index-acute kidney injury association. Design:Prospective cohort study. Setting:Level I Trauma Center of a university hospital. Patients:Patients older than 13 years with an Injury Severity Score greater than or equal to 16 admitted to the trauma ICU were followed for development of acute kidney injury over 5 days. Those with isolated severe head injury or on chronic dialysis were excluded. Interventions:None. Measurements and Main Results:Clinical, anthropometric, and demographic variables were collected prospectively. CT images at the level of the L4–5 intervertebral disc space were extracted from the medical record and used by two operators to quantitate visceral adipose tissue and subcutaneous adipose tissue areas. Acute kidney injury was defined by Acute Kidney Injury Network creatinine and dialysis criteria. Of 400 subjects, 327 (81.8%) had CT scans suitable for analysis: 264 of 285 (92.6%) blunt trauma subjects and 63 of 115 (54.8%) penetrating trauma subjects. Visceral adipose tissue and subcutaneous adipose tissue areas were highly correlated between operators (intraclass correlation > 0.99, p < 0.001 for each) and within operator (intraclass correlation > 0.99, p < 0.001 for each). In multivariable analysis, the standardized risk of acute kidney injury was 15.1% (95% CI, 10.6–19.6%), 18.1% (14–22.2%), and 23.1% (18.3–27.9%) at the 25th, 50th, and 75th percentiles of visceral adipose tissue area, respectively (p = 0.001), with similar findings when using subcutaneous adipose tissue area as the adiposity measure. Conclusions:Quantitation of abdominal adiposity using CT scans obtained for clinical reasons is feasible and highly reliable in critically ill trauma patients. Abdominal adiposity is independently associated with acute kidney injury in this population, confirming that excess adipose tissue contributes to the body mass index-acute kidney injury association. Further studies of the potential mechanisms linking adiposity with acute kidney injury are warranted.

Is It Possible to Prevent ARDS

Since its initial description, the acute respiratory distress syndrome (ARDS) has been the target of many therapeutic interventions, but with only modest success.1 Improved understanding of the complex pathogenesis responsible for ARDS has led to several promising pharmacologic therapies aimed at reducing the morbidity and mortality associated with ARDS in critically ill patients. However, the majority of clinical trials of pharmacologic interventions have shown no beneficial effects, leaving ARDS mortality and morbidity unacceptably high. One possible explanation for the inefficacy of ARDS therapies is timing of delivery. Conventional ARDS clinical trials have been designed to enroll patients within 48 hours of ARDS onset, sometimes well into the critical illness phase. In rodent models, several potential therapies are effective at preventing lung injury if administered prior to insult but are less effective at rescuing the animals once the injury is substantial.2,3 These findings suggest that some therapies may be better at interrupting the initial cascade of events leading to ARDS (eg, leukocyte recruitment, platelet activation, and microvascular coagulation) than reversing lung injury once the alveolar-capillary barrier has broken down.

Plasma Levels of Receptor Interacting Protein Kinase-3 (RIP3), an Essential Mediator of Necroptosis, are Associated with Acute Kidney Injury in Critically Ill Trauma Patients.

Background: Receptor interacting protein kinase-3 (RIP3) is a key mediator of necroptosis, a form of regulated cell death recently implicated in murine models of renal ischemia-reperfusion injury and transfusion-associated endothelial injury. The importance of necroptosis in human AKI is unknown. We hypothesized that plasma RIP3 concentrations would be associated with acute kidney injury (AKI) after severe trauma. Methods: We performed a case-control study nested in a prospective cohort of critically ill trauma patients. AKI was defined by AKI Network creatinine criteria within 6 days of presentation. Of 158 cohort subjects, we selected 13 who developed AKI stage 2 or 3, 27 with AKI stage 1, and 40 without AKI. We compared plasma RIP3 concentrations across these groups at presentation and 48 h. Since red blood cell (RBC) transfusion is an AKI risk factor, we also tested the association of RBCs transfused during resuscitation with RIP3 levels. Results: Median plasma RIP3 concentration rose more than 10-fold from presentation (15.6 (interquartile range 15.6–41.3) pg/mL) to 48 h (164.7 (66.9–300.6) pg/mL; P <0.001). RIP3 concentrations at 48 h were associated with AKI stage (no AKI: 144.8 (58.6–234.9) pg/mL; AKI stage 1: 165.8 (43.0–310.9) pg/mL; AKI stage 2–3: 365.5 (155.1–727.5) pg/mL; P = 0.010) whereas this association was not seen at presentation (P = 0.324). RBC transfusions were also associated with 48-h plasma RIP3 (no RBCs: 99.4 (15.6–166.1) pg/mL; 1–5 units: 182.6 (98.5–274.1) pg/mL; >5 units: 341.8 (150.1–423.8) pg/mL; P <0.001). Conclusions: In critically ill trauma patients, plasma levels of the necroptosis mediator RIP3 at 48 h were associated with AKI stage and RBC transfusions.

The ABO Histo-Blood Group and AKI in Critically Ill Patients with Trauma or Sepsis

BACKGROUND AND OBJECTIVE ABO blood types are determined by antigen modifications on glycoproteins and glycolipids and associated with altered plasma levels of inflammatory and endothelial injury markers implicated in AKI pathogenesis. We sought to determine the association of ABO blood types with AKI risk in critically ill patients with trauma or sepsis. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS We conducted two prospective cohort studies at an urban, academic, level I trauma center and tertiary referral center; 497 patients with trauma admitted to the surgical intensive care unit between 2005 and 2010 with an injury severity score >15 and 759 patients with severe sepsis admitted to the medical intensive care unit between 2008 and 2013 were followed for 6 days for the development of incident AKI. AKI was defined by Acute Kidney Injury Network creatinine and dialysis criteria. RESULTS Of 497 patients with trauma, 134 developed AKI (27%). In multivariable analysis, blood type A was associated with higher AKI risk relative to type O among patients of European descent (n=229; adjusted risk, 0.28 versus 0.14; risk difference, 0.14; 95% confidence interval, 0.03 to 0.24; P=0.02). Of 759 patients with sepsis, AKI developed in 326 (43%). Blood type A again conferred higher AKI risk relative to type O among patients of European descent (n=437; adjusted risk, 0.53 versus 0.40; risk difference, 0.14; 95% confidence interval, 0.04 to 0.23; P=0.01). Findings were similar when analysis was restricted to those patients who did not develop acute respiratory distress syndrome or were not transfused. We did not detect a significant association between blood type and AKI risk among individuals of African descent in either cohort. CONCLUSIONS Blood type A is independently associated with AKI risk in critically ill patients with trauma or severe sepsis of European descent, suggesting a role for ABO glycans in AKI susceptibility.

Linking Genetics to ARDS Pathogenesis : The Role of the Platelet

ARDS is a complex syndrome, characterized by damage to the alveolar-capillary barrier resulting in increased permeability in the setting of an amplified inflammatory response. The role of the activation of clotting pathways in this process has been increasingly recognized, including the significant impact of platelets on ARDS pathogenesis.1-3 In a steady state, inactive platelets circulate in blood and promote endothelial barrier integrity.4 However, in the setting of critical illnesses (eg, sepsis), several overlapping mechanisms activate platelets, resulting in platelet aggregation, platelet-leukocyte complex formation, and release of the granule contents of the platelet cell, including molecules that enhance inflammation and cell adhesion. While platelet-mediated thrombosis in the lungs has been implicated in ARDS pathogenesis, the activation of circulating platelets results in a host of other inflammatory effects contributing to ARDS development, including augmented neutrophil migration and increased endothelial barrier permeability.3 Given the accumulating evidence for the role of platelets in ARDS, significant interest has recently developed in therapeutically targeting platelet activation and function. In mouse models of ARDS, inhibiting P-selectin-mediated platelet-neutrophil interactions protect from lung injury.2 In humans, preexisting antiplatelet drugs, such as aspirin or clopidogrel, are associated with decreased risk of ARDS as well as decreased mortality among the critically ill.5,6 Several ongoing studies are investigating the role of antiplatelet therapy in ARDS, including the Lung Injury Prevention Study with Aspirin (LIPS-A),7 a multicentered randomized controlled clinical trial evaluating aspirin for the prevention of ARDS in patients at risk. While current antiplatelet agents have the potential to have therapeutic benefit in the critically ill, further enhancing our understanding of the mechanisms by which platelets contribute to ARDS may identify novel therapeutic targets or select patient subgroups most likely to respond to antiplatelet therapy. In this issue of CHEST (see page 607), Wei and colleagues8 use an innovative approach to identify a novel genetic variant linked to platelets, which is associated with ARDS risk. Several investigators have identified genetic variants in genes implicated in alveolar-capillary barrier integrity,9 inflammation,10 and coagulation as risk factors for ARDS11; however, variants resulting in platelet heterogeneity have not previously been linked to ARDS. Working with the knowledge that thrombocytopenia during critical illness is associated with poor outcomes,12 and that genome-wide association studies have identified several genetic variants linked to platelet count in the noncritically ill,13 Wei and colleagues8 set out to answer several questions. First, they determined the association of single nucleotide polymorphisms (SNPs) in candidate genes with platelet count in a cohort of critically ill patients at risk for ARDS. They selected candidate genes based on previous associations with platelet count in relatively healthy populations and identified an intronic SNP in LRRC16A as also associated with platelet count among the critically ill. Next, the same SNP in LRRC16A was determined to be associated with altered risk of ARDS. Lastly, they used sophisticated statistical methods to determine whether platelet count was a causal mediator linking the LRRC16A genotype to ARDS risk. Decreased platelet count, measured at ICU admission, was associated with increased ARDS risk in their cohort and functioned as a partial mediator of the association between LRRC16A genotype and ARDS. While not proving causation, this approach not only identified a novel genetic risk factor for ARDS, but also provided evidence for a potential mechanism resulting in the altered risk. The most significant limitation to the study conducted by Wei and colleagues8 is that the only causal mediator evaluated was a single measure of platelet count at ICU admission. Based on these data, it is difficult to determine whether the effects of platelet count on ARDS risk are a result of decreased platelet production or increased platelet activation, lung sequestration, and consumption. Additionally, platelet function may be equally as important in mediating ARDS risk as platelet count and may explain why count was only a partial causal mediator of the LRRC16A and ARDS association. In addition to validating their findings in an independent population, future research should evaluate the effects of LRRC16A on measures of platelet consumption and production, such as serial platelet counts or immature platelet fractions, as well as measures of platelet function, aggregation, and neutrophil adhesion. Despite these limitations, Wei and colleagues8 have successfully identified a variant in LRRC16A associated with altered ARDS risk, at least partially mediated via effects on platelet count. LRRC16A encodes the protein, capping protein ARP2/3 and myosin-I linker (CARMIL), important in actin-based cellular processes.14 This protein has not previously been implicated in ARDS, and the mechanisms underlying its effects on platelet count are not completely understood. Furthering our understanding of the role of LRRC16A and CARMIL protein in platelet development and/or function as well as ARDS pathogenesis may identify novel therapeutic targets within platelet cellular processes with relevance to ARDS. Likewise, LRRC16A variation may represent a key focus for pharmacogenetic interaction in ongoing trials of aspirin therapy. Therefore, the study by Wei and colleagues8 provides key data to focus our near-term research efforts on the role of platelets in ARDS.

Mortality Benefit of Recombinant Human Interleukin-1 Receptor Antagonist for Sepsis Varies by Initial Interleukin-1 Receptor Antagonist Plasma Concentration*

Objective: Plasma interleukin-1 beta may influence sepsis mortality, yet recombinant human interleukin-1 receptor antagonist did not reduce mortality in randomized trials. We tested for heterogeneity in the treatment effect of recombinant human interleukin-1 receptor antagonist by baseline plasma interleukin-1 beta or interleukin-1 receptor antagonist concentration. Design: Retrospective subgroup analysis of randomized controlled trial. Setting: Multicenter North American and European clinical trial. Patients: Five hundred twenty-nine subjects with sepsis and hypotension or hypoperfusion, representing 59% of the original trial population. Interventions: Random assignment of placebo or recombinant human interleukin-1 receptor antagonist × 72 hours. Measurements and Main Results: We measured prerandomization plasma interleukin-1 beta and interleukin-1 receptor antagonist and tested for statistical interaction between recombinant human interleukin-1 receptor antagonist treatment and baseline plasma interleukin-1 receptor antagonist or interleukin-1 beta concentration on 28-day mortality. There was significant heterogeneity in the effect of recombinant human interleukin-1 receptor antagonist treatment by plasma interleukin-1 receptor antagonist concentration whether plasma interleukin-1 receptor antagonist was divided into deciles (interaction p = 0.046) or dichotomized (interaction p = 0.028). Interaction remained present across different predicted mortality levels. Among subjects with baseline plasma interleukin-1 receptor antagonist above 2,071 pg/mL (n = 283), recombinant human interleukin-1 receptor antagonist therapy reduced adjusted mortality from 45.4% to 34.3% (adjusted risk difference, –0.12; 95% CI, –0.23 to –0.01), p = 0.044. Mortality in subjects with plasma interleukin-1 receptor antagonist below 2,071 pg/mL was not reduced by recombinant human interleukin-1 receptor antagonist (adjusted risk difference, +0.07; 95% CI, –0.04 to +0.17), p = 0.230. Interaction between plasma interleukin-1 beta concentration and recombinant human interleukin-1 receptor antagonist treatment was not statistically significant. Conclusions: We report a heterogeneous effect of recombinant human interleukin-1 receptor antagonist on 28-day sepsis mortality that is potentially predictable by plasma interleukin-1 receptor antagonist in one trial. A precision clinical trial of recombinant human interleukin-1 receptor antagonist targeted to septic patients with high plasma interleukin-1 receptor antagonist may be worthy of consideration.

Admission plasma levels of the neuronal injury marker neuron-specific enolase are associated with mortality and delirium in sepsis

PURPOSE Neuron-specific enolase (NSE) concentrations are prognostic following traumatic and anoxic brain injury and may provide a method to quantify neuronal injury in other populations. We determined the association of admission plasma NSE concentrations with mortality and delirium in critically ill septic patients. METHODS We performed a retrospective analysis of 124 patients from a larger sepsis cohort. Plasma NSE was measured in the earliest blood draw at intensive care unit admission. Primary outcomes were 30-day mortality and intensive care unit delirium determined by chart review. RESULTS Sixty-one patients (49.2%) died within 30 days, and delirium developed in 34 (31.5%) of the 108 patients who survived at least 24 hours and were not persistently comatose. Each doubling of the NSE concentration was associated with a 7.3% (95% confidence interval [CI] 2.5-12.0, P= .003) increased risk of 30-day mortality and a 5.2% (95% CI 3.2-7.2, P< .001) increased risk of delirium. An NSE concentration >12.5 μg/L was independently associated with a 23.3% (95% CI 6.7-39.9, P= .006) increased risk of 30-day mortality and a 29.3% (95% CI 8.8-49.8, P= .005) increased risk of delirium. CONCLUSIONS Higher plasma NSE concentrations were associated with mortality and delirium in critically ill septic patients, suggesting that NSE may have utility as a marker of neuronal injury in sepsis.