Mechanism matters: mortality and endothelial cell damage marker differences between blunt and penetrating traumatic injuries across three prehospital clinical trials.

Injury mechanism is an important consideration when conducting clinical trials in trauma. Mechanisms of injury may be associated with differences in mortality risk and immune response to injury, impacting the potential success of the trial. We sought to characterize clinical and endothelial cell damage marker differences across blunt and penetrating injured patients enrolled in three large, prehospital randomized trials which focused on hemorrhagic shock. In this secondary analysis, patients with systolic blood pressure < 70 or systolic blood pressure < 90 and heart rate > 108 were included. In addition, patients with both blunt and penetrating injuries were excluded. The primary outcome was 30-day mortality. Mortality was characterized using Kaplan-Meier and Cox proportional-hazards models. Generalized linear models were used to compare biomarkers. Chi squared tests and Wilcoxon rank-sum were used to compare secondary outcomes. We characterized data of 696 enrolled patients that met all secondary analysis inclusion criteria. Blunt injured patients had significantly greater 24-h (18.6% vs. 10.7%, log rank p = 0.048) and 30-day mortality rates (29.7% vs. 14.0%, log rank p = 0.001) relative to penetrating injured patients with a different time course. After adjusting for confounders, blunt mechanism of injury was independently predictive of mortality at 30-days (HR 1.84, 95% CI 1.06-3.20, p = 0.029), but not 24-h (HR 1.65, 95% CI 0.86-3.18, p = 0.133). Elevated admission levels of endothelial cell damage markers, VEGF, syndecan-1, TM, S100A10, suPAR and HcDNA were associated with blunt mechanism of injury. Although there was no difference in multiple organ failure (MOF) rates across injury mechanism (48.4% vs. 42.98%, p = 0.275), blunt injured patients had higher Denver MOF score (p < 0.01). The significant increase in 30-day mortality and endothelial cell damage markers in blunt injury relative to penetrating injured patients highlights the importance of considering mechanism of injury within the inclusion and exclusion criteria of future clinical trials.

Prehospital Time Following Traumatic Injury Is Independently Associated With the Need for In-Hospital Blood and Early Mortality for Specific Injury Types.

OBJECTIVE: Treating traumatic hemorrhage is time sensitive. Prehospital care and transport modes (eg, helicopter and ground) may influence in-hospital events. We hypothesized that prehospital time (on-scene time [OST] and total prehospital time [TPT]) and transport mode are associated with same-day transfusion and mortality. Furthermore, we sought to identify regions of anatomic injury that modify the relationship between prehospital time and outcomes in strata corresponding to transport types. METHODS: We obtained prehospital, in-hospital, and trauma registry data from an 8-center cohort of adult nonburn trauma patients from 2017 to 2022 directly transported from the scene to the hospital and having an Injury Severity Score (ISS) > 9 for the Task Order 1 project of the Linking Investigators in Trauma and Emergency Services research network. We excluded patients missing prehospital times, patients < 18 years of age, patients from interfacility transfers, and recipients of prehospital blood. Our same-day outcomes were in-hospital transfusions within 4 hours and 24-hour mortality. Each outcome was adjusted using multivariable logistic regression for covariates of prehospital phases (OST and TPT), mode of transport (helicopter and ground), age, sex, ISS, Glasgow Coma Scale motor subscale score < 6, and field hypotension (systolic blood pressure < 90 mm Hg). We evaluated the association of prehospital time on outcomes for scene missions by transport mode across severe injury patterns defined by Abbreviated Injury Scale > 2 body regions. RESULTS: Of 78,198 subjects, 34,504 were eligible for the study with a mean age of 47.6 ± 20.3 years, ISS of 18 ± 11, OST of 15.9 ± 9.5 minutes, and TPT of 48.7 ± 20.3 minutes. Adjusted for injury severity and demographic factors, transport type significantly modified the relationship between prehospital time and outcomes. The association of OST and TPT with the odds of 4-hour transfusion was absent for the ground emergency medical services (GEMS) cohort and present for the helicopter emergency medical services (HEMS) ambulance cohort, whereas these times were associated with decreased 24-hour mortality for both transport types. When stratifying by injury to most anatomic regions, OST and TPT were associated with a decreased need for 4-hour transfusions in the GEMS cohort. However, OST was associated with increased early transfusion only among patients with severe injuries of the thorax, and this association persisted after adjusting additionally for injury type (odds ratio [OR] = 1.03; 95% confidence interval [CI], 1.00-1.05; P = .02). The presence of polytrauma supported an association between prehospital time and decreased 24-hour mortality for the GEMS cohort (OST: OR = 0.97; 95% CI, 0.95-0.99; P < .01; TPT: OR = 0.99; 95% CI, 0.98-0.99; P = .02), whereas no injuries showed significant association of helicopter prehospital time on mortality after adjustment. CONCLUSION: We determined that transport type affects the relationship between prehospital time and hospital outcomes (4-hour transfusion: positive relationship for HEMS and negative for GEMS, 24-hour mortality: negative for both transport types). Furthermore, we identified regions of anatomic injury that modify the relationship between prehospital time and outcomes in strata corresponding to transport types. Of these regions, most notable were severe isolated injuries to the thorax that supported a positive relationship between HEMS OST and 4-hour transfusions and polytrauma that showed a negative relationship between GEMS OST or TPT and 24-hour mortality after adjustment.

Association between trauma triage and time-to-vaso-occlusive events in patients with sickle cell disease after traumatic injury: a retrospective study.

Introduction: Sickle cell disease (SCD) is associated with vaso-occlusive events (VOEs) that can lead to disease complications, including early mortality. Given that similar inflammatory responses characterize VOE and traumatic injury, injured patients with SCD may be vulnerable to acute complications. This study is the first to examine whether traumatic injury is associated with increased severity of future VOEs. Methods: This cohort study was conducted using electronic health record data from an SCD clinic in Western Pennsylvania; 356 patients with SCD from January 2000 to July 2021 were identified via retrospective chart review. 55 patients were eligible based on continuous medical record data spanning 1 year preinjury and postinjury. Patients were sorted into three treatment groups based on injury management: (1) Neither triage to trauma team activation (TTA) nor inpatient admission (Early Discharge), (2) Triage but no inpatient admission (Triage Only), and (3) Triage and In-patient. Outcomes included time from injury to first VOE, annual VOE counts requiring an emergency department (ED) visit, and ED length of stay (LOS) for the first VOE after injury. Results: Early Discharge individuals experienced a VOE event within 2.93 days of injury, significantly shorter time to event than Triage and In-patient individuals at 52.375 days and Triage Only individuals at 100.16 days (p=0.0058). No difference in annual VOE counts was noted postinjury across all groups. However, a significant increase in VOE LOS preinjury (16.1 hours) to postinjury (77.4 hours) was noted only for the Triage Only group (p=0.038). Cox regression model showed that shortened time to VOE events was marginally associated with TTA status (p=0.06). Conclusion: Despite minimal changes in long-term VOE outcomes after injury, traumatic injuries may accelerate the time-to-VOE among the Early Discharge group. Therefore, future research is warranted to analyze whether the absence of postinjury triage assessment and intervention may cause unforeseen physiologic stressors contributing to VOE outcomes. Level of evidence: Level IV: retrospective case-control study with three negative criteria.

Prehospital tranexamic acid is associated with a dose-dependent decrease in syndecan-1 after trauma: A secondary analysis of a prospective randomized trial.

BACKGROUND: In the Study of Tranexamic Acid During Air and Ground Prehospital Transport (STAAMP) Trial, prehospital tranexamic acid (TXA) was associated with lower mortality in specific patient subgroups. The underlying mechanisms responsible for a TXA benefit remain incompletely characterized. We hypothesized that TXA may mitigate endothelial injury and sought to assess whether TXA was associated with decreased endothelial or tissue damage markers among all patients enrolled in the STAAMP Trial. METHODS: We collected blood samples from STAAMP Trial patients and measured markers of endothelial function and tissue damage including syndecan-1, soluble thrombomodulin (sTM), and platelet endothelial cell adhesion molecule-1 at hospital admission (0 hours) and 12 hours, 24 hours, and 72 hours after admission. We compared these marker values for patients in each treatment group during the first 72 hours, and modeled the relationship between TXA and marker concentration using regression analysis to control for potential confounding factors. RESULTS: We analyzed samples from 766 patients: 383 placebo, 130 abbreviated dosing, 119 standard dosing, and 130 repeat dosing. Lower levels of syndecan-1, TM, and platelet endothelial cell adhesion molecule measured within the first 72 hours of hospital admission were associated with survival at 30 days ( p < 0.001). At hospital admission, syndecan-1 was lower in the TXA group (28.30 [20.05, 42.75] vs. 33.50 [23.00, 54.00] p = 0.001) even after controlling for patient, injury, and prehospital factors ( p = 0.001). For every 1 g increase in TXA administered over the first 8 hours of prehospital transport and hospital admission, there was a 4-ng/mL decrease in syndecan-1 at 12 hours controlling for patient, injury, and treatment factors ( p = 0.03). CONCLUSION: Prehospital TXA was associated with decreased syndecan-1 at hospital admission. Syndecan-1 measured 12 hours after admission was inversely related to the dose of TXA received. Early prehospital and in-hospital TXA may decrease endothelial glycocalyx damage or upregulate vascular repair mechanisms in a dose-dependent fashion. LEVEL OF EVIDENCE: Therapeutic/Care Management; Level III.

Lipidomic signatures align with inflammatory patterns and outcomes in critical illness.

Alterations in lipid metabolism have the potential to be markers as well as drivers of pathobiology of acute critical illness. Here, we took advantage of the temporal precision offered by trauma as a common cause of critical illness to identify the dynamic patterns in the circulating lipidome in critically ill humans. The major findings include an early loss of all classes of circulating lipids followed by a delayed and selective lipogenesis in patients destined to remain critically ill. The previously reported survival benefit of early thawed plasma administration was associated with preserved lipid levels that related to favorable changes in coagulation and inflammation biomarkers in causal modelling. Phosphatidylethanolamines (PE) were elevated in patients with persistent critical illness and PE levels were prognostic for worse outcomes not only in trauma but also severe COVID-19 patients. Here we show selective rise in systemic PE as a common prognostic feature of critical illness.

High Dimensional Multiomics Reveals Unique Characteristics of Early Plasma Administration in Polytrauma Patients With TBI.

OBJECTIVES: The authors sought to identify causal factors that explain the selective benefit of prehospital administration of thawed plasma (TP) in traumatic brain injury (TBI) patients using mediation analysis of a multiomic database. BACKGROUND: The Prehospital Air Medical Plasma (PAMPer) Trial showed that patients with TBI and a pronounced systemic response to injury [defined as endotype 2 (E2)], have a survival benefit from prehospital administration of TP. An interrogation of high dimensional proteomics, lipidomics and metabolomics previously demonstrated unique patterns in circulating biomarkers in patients receiving prehospital TP, suggesting that a deeper analysis could reveal causal features specific to TBI patients. METHODS: A novel proteomic database (SomaLogic Inc., aptamer-based assay, 7K platform) was generated using admission blood samples from a subset of patients (n=149) from the PAMPer Trial. This proteomic dataset was combined with previously reported metabolomic and lipidomic datasets from these same patients. A 2-step analysis was performed to identify factors that promote survival in E2-TBI patients who had received early TP. First, features were selected using both linear and multivariate-latent-factor regression analyses. Then, the selected features were entered into the causal mediation analysis. RESULTS: Causal mediation analysis of observable features identified 16 proteins and 41 lipids with a high proportion of mediated effect (>50%) to explain the survival benefit of early TP in E2-TBI patients. The multivariate latent-factor regression analyses also uncovered 5 latent clusters of features with a proportion effect >30%, many in common with the observable features. Among the observable and latent features were protease inhibitors known to inhibit activated protein C and block fibrinolysis (SERPINA5 and CPB2), a clotting factor (factor XI), as well as proteins involved in lipid transport and metabolism (APOE3 and sPLA(2)-XIIA). CONCLUSIONS: These findings suggest that severely injured patients with TBI process exogenous plasma differently than those without TBI. The beneficial effects of early TP in E2-TBI patients may be the result of improved blood clotting and the effect of brain protective factors independent of coagulation.

Prehospital Lactate is Associated with the Need for Blood in Trauma.

Background: Traumatic hemorrhage is the leading cause of preventable death, and its effects are often evident within the first 24 hours of hospital admission. We investigated the relationship between prehospital lactate measurement and administration of hospital blood products and life-saving interventions (LSIs) within 24 hours of hospital admission.Methods: We included trauma patients with recorded prehospital venous lactate transported by a single critical care transport service to a Level I trauma center between 2012 and 2019. We abstracted vital signs, mission type, anatomic location of injury, prehospital administration of crystalloid and blood products, and hospital LSIs started within 24 hours of admission. We used logistic regression to determine the association of prehospital lactate and in-hospital administration of blood products in groups with or without hypotensive patients. We investigated the effect of prehospital lactate concentration on secondary outcomes such as LSIs and mortality.Results: We included 2,170 patients transported from the scene or emergency department (ED), of whom 1,821 (84%) were normotensive. The median concentration of prehospital lactate was 2.10 mmol/L for the main population (IQR = 1.40-3.30) and 2.00 mmol/L for the normotensive subgroup (IQR = 1.30-2.90). A higher prehospital lactate concentration was associated with higher odds of needing early hospital blood products in the whole study population (OR = 1.12, (95% CI 1.06-1.20), p < 0.01) and in the normotensive subgroup (OR = 1.13, (95% CI 1.03-1.22), p = 0.01). These positive associations were also observed with the secondary outcome of hospital LSIs, and higher prehospital lactate was also associated with higher odds of mortality (OR = 1.32, (95% CI 1.20-1.45), p < 0.01).Conclusions: Higher concentrations of prehospital lactate were associated with the need for in-hospital blood transfusion within 24 hours of admission. The relationship between lactate and blood transfusion persisted among normotensive patients. Further work is needed to incorporate prehospital lactate into decision support tools for prehospital blood administration.

Protective/reparative cytokines are suppressed at high injury severity in human trauma.

BACKGROUND: Trauma elicits a complex inflammatory response that, among multiple presenting factors, is greatly impacted by the magnitude of injury severity. Herein, we compared the changes in circulating levels of mediators with known proinflammatory roles to those with known protective/reparative actions as a function of injury severity in injured humans. METHODS: Clinical and biobank data were obtained from 472 (trauma database-1 (TD-1), University of Pittsburgh) and 89 (trauma database-2 (TD-2), Indiana University) trauma patients admitted to the intensive care unit (ICU) and who survived to discharge. Injury severity was estimated based on the Injury Severity Score (ISS), and this was used as both a continuous variable and for the purpose of grouping patients into severity-based cohorts. Samples within the first 24 hours were obtained from all patients and then daily up to day 7 postinjury in TD-1. Sixteen cytokines were assayed using Luminex and were analyzed using two-way analysis of variance (p<0.05). RESULTS: Patients with higher ISSs had longer ICU and hospital stays, days on mechanical ventilation and higher rates of nosocomial infection when compared with the mild and moderate groups. Time course analysis and correlations with ISS showed that 11 inflammatory mediators correlated positively with injury severity, consistent with previous reports. However, five mediators (interleukin (IL)-9, IL-21, IL-22, IL-23 and IL-17E/25) were suppressed in patients with high ISS and inversely correlated with ISS. DISCUSSION: These findings suggest that severe injury is associated with a suppression of a subset of cytokines known to be involved in tissue protection and regeneration (IL-9, IL-22 and IL-17E/25) and lymphocyte differentiation (IL-21 and IL-23), which in turn correlates with adverse clinical outcomes. Thus, patterns of proinflammatory versus protective/reparative mediators diverge with increasing ISS.

Age of thawed plasma does not affect clinical outcomes or biomarker expression in patients receiving prehospital thawed plasma: a PAMPer secondary analysis.

BACKGROUND: Prehospital plasma administration during air medical transport reduces the endotheliopathy of trauma, circulating pro-inflammatory cytokines, and 30-day mortality among traumatically injured patients at risk of hemorrhagic shock. No clinical data currently exists evaluating the age of thawed plasma and its association with clinical outcomes and biomarker expression post-injury. METHODS: We performed a secondary analysis from the prehospital plasma administration randomized controlled trial, PAMPer. We dichotomized the age of thawed plasma creating three groups: standard-care, YOUNG (day 0-1) plasma, and OLD (day 2-5) plasma. We generated HRs and 95% CIs for mortality. Among all patients randomized to plasma, we compared predicted biomarker values at hospital admission (T0) and 24 hours later (T24) controlling for key difference between groups with a multivariable linear regression. Analyses were repeated in a severely injured subgroup. RESULTS: Two hundred and seventy-one patients were randomized to standard-care and 230 to plasma (40% YOUNG, 60% OLD). There were no clinically or statistically significant differences in demographics, injury, admission vital signs, or laboratory values including thromboelastography between YOUNG and OLD. Compared with standard-care, YOUNG (HR 0.66 (95% CI 0.41 to 1.07), p=0.09) and OLD (HR 0.64 (95% CI 0.42 to 0.96), p=0.03) plasma demonstrated reduced 30-day mortality. Among those randomized to plasma, plasma age did not affect mortality (HR 1.04 (95% CI 0.60 to 1.82), p=0.90) and/or adjusted serum markers by plasma age at T0 or T24 (p>0.05). However, among the severely injured subgroup, OLD plasma was significantly associated with increased adjusted inflammatory and decreased adjusted endothelial biomarkers at T0. DISCUSSION: Age of thawed plasma does not result in clinical outcome or biomarker expression differences in the overall PAMPer study cohort. There were biomarker expression differences in those patients with severe injury. Definitive investigation is needed to determine if the age of thawed plasma is associated with biomarker expression and outcome differences following traumatic injury. LEVEL OF EVIDENCE: II.

Lipidomic Signatures Align with Inflammatory Patterns and Outcomes in Critical Illness.

Alterations in lipid metabolism have the potential to be markers as well as drivers of the pathobiology of acute critical illness. Here, we took advantage of the temporal precision offered by trauma as a common cause of critical illness to identify the dynamic patterns in the circulating lipidome in critically ill humans. The major findings include an early loss of all classes of circulating lipids followed by a delayed and selective lipogenesis in patients destined to remain critically ill. Early in the clinical course, Fresh Frozen Plasma administration led to improved survival in association with preserved lipid levels that related to favorable changes in coagulation and inflammation biomarkers. Late over-representation of phosphatidylethanolamines with critical illness led to the validation of a Lipid Reprogramming Score that was prognostic not only in trauma but also severe COVID-19 patients. Our lipidomic findings provide a new paradigm for the lipid response underlying critical illness.

Multi-omic analysis in injured humans: Patterns align with outcomes and treatment responses.

Trauma is a leading cause of death and morbidity worldwide. Here, we present the analysis of a longitudinal multi-omic dataset comprising clinical, cytokine, endotheliopathy biomarker, lipidome, metabolome, and proteome data from severely injured humans. A "systemic storm" pattern with release of 1,061 markers, together with a pattern suggestive of the "massive consumption" of 892 constitutive circulating markers, is identified in the acute phase post-trauma. Data integration reveals two human injury response endotypes, which align with clinical trajectory. Prehospital thawed plasma rescues only endotype 2 patients with traumatic brain injury (30-day mortality: 30.3 versus 75.0%; p = 0.0015). Ubiquitin carboxy-terminal hydrolase L1 (UCHL1) was identified as the most predictive circulating biomarker to identify endotype 2-traumatic brain injury (TBI) patients. These response patterns refine the paradigm for human injury, while the datasets provide a resource for the study of critical illness, trauma, and human stress responses.

Association of Prehospital Plasma With Survival in Patients With Traumatic Brain Injury: A Secondary Analysis of the PAMPer Cluster Randomized Clinical Trial.

Importance: Prehospital plasma administration improves survival in injured patients at risk for hemorrhagic shock and transported by air ambulance. Traumatic brain injury (TBI) is a leading cause of death following trauma, but few early interventions improve outcomes. Objective: To assess the association between prehospital plasma and survival in patients with TBI. Design, Setting, and Participants: The Prehospital Air Medical Plasma (PAMPer) trial was a pragmatic, multicenter, phase 3, cluster randomized clinical trial involving injured patients who were at risk for hemorrhagic shock during air medical transport to a trauma center. The trial was conducted at 6 US sites with 9 level-I trauma centers (comprising 27 helicopter emergency services bases). The original trial analyzed 501 patients, including 230 patients who were randomized to receive plasma and 271 randomized to standard care resuscitation. This secondary analysis of a predefined subgroup included patients with TBI. Data analysis was performed from October 2019 to February 2020. Interventions: Patients were randomized to receive standard care fluid resuscitation or 2 units of thawed plasma. Main Outcomes and Measures: The primary outcome was mortality at 30 days. Patients with TBI were prespecified as a subgroup for secondary analysis and for measurement of markers of brain injury. The 30-day survival benefit of prehospital plasma in subgroups with and without TBI as diagnosed by computed tomography was characterized using Kaplan-Meier survival analysis and Cox proportional hazard regression. Results: In total, 166 patients had TBI (median [interquartile range] age, 43.00 [25.00-59.75] years; 125 men [75.3%]). When compared with the 92 patients who received standard care, the 74 patients with TBI who received prehospital plasma had improved 30-day survival even after adjustment for multiple confounders and assessment of the degree of brain injury with clinical variables and biomarkers (hazard ratio [HR], 0.55; 95% CI, 0.33-0.94; P = .03). Receipt of prehospital plasma was associated with improved survival among patients with TBI with a prehospital Glasgow Coma Scale score of less than 8 (HR, 0.56; 95% CI, 0.35-0.91) and those with polytrauma (HR, 0.50; 95% CI, 0.28-0.89). Patients with TBI transported from the scene of injury had improved survival following prehospital plasma administration (HR, 0.45; 95% CI, 0.26-0.80; P = .005), whereas patients who were transferred from an outside hospital showed no difference in survival for the plasma intervention (HR, 1.00; 95% CI, 0.33-3.00; P = .99). Conclusions and Relevance: These findings are exploratory, but they suggest that receipt of prehospital plasma is associated with improved survival in patients with computed tomography-positive TBI. The prehospital setting may be a critical period to intervene in the care of patients with TBI. Future studies are needed to confirm the clinical benefits of early plasma resuscitation following TBI and concomitant polytrauma. Trial Registration: ClinicalTrials.gov Identifier: NCT01818427.

Characterization of unexpected survivors following a prehospital plasma randomized trial.

BACKGROUND: Prehospital plasma improves survival for severely injured trauma patients transported by air ambulance. We sought to characterize the unexpected survivors, patients who survived despite having high predicted mortality after traumatic injury. METHODS: The Prehospital Air Medical Plasma trial randomized severely injured patients (n = 501) to receive either standard care (crystalloid) or two units of prehospital plasma followed by standard care fluid resuscitation. We built a generalized linear model to estimate patient mortality. Area under the receiver operating characteristic curve was used to evaluate model performance. We defined unexpected survivors as patients who had a predicted mortality greater than 50% and survived to 30 days. We characterized patient demographics, clinical features, and outcomes of the unexpected survivors. Observed to expected (O/E) ratios and Z-statistics were calculated using model-estimated mortality for each cohort. RESULTS: Our model predicted mortality better than Injury Severity Score or Revised Trauma Score parameters and identified 36 unexpected survivors. Compared with expected survivors, unexpected survivors were younger (33 years [24, 52 years] vs. 47 years [32, 59 years], p = 0.013), were more severely injured (Injury Severity Score 34 [22, 50] vs. 18 [10, 27], p < 0.001), had worse organ dysfunction and hospital resource outcomes (multiple organ failure, intensive care unit, hospital length of stay, and ventilator days), and were more likely to receive prehospital plasma (67 vs. 46%, p = 0.031). Nonsurvivors with high predicted mortality were more likely to receive standard care resuscitation (p < 0.001). Unexpected survivors who received prehospital plasma had a lower observed to expected mortality than those that received standard care resuscitation (O/E 0.56 [0.33-0.84] vs. 1.0 [0.73-1.32]). The number of prehospital plasma survivors (24) exceeded the number of predicted survivors (n = 10) estimated by our model (p < 0.001). CONCLUSION: Prehospital plasma is associated with an increase in the number of unexpected survivors following severe traumatic injury. Prehospital interventions may improve the probability of survival for injured patients with high predicted mortality based on early injury characteristics, vital signs, and resuscitation measures. LEVEL OF EVIDENCE: Therapeutic Level III.

Prehospital plasma is associated with distinct biomarker expression following injury.

BACKGROUNDPrehospital plasma improves survival in severely injured patients transported by air ambulance. We hypothesized that prehospital plasma would be associated with a reduction in immune imbalance and endothelial damage.METHODSWe sampled blood from 405 trauma patients enrolled in the Prehospital Air Medical Plasma (PAMPer) trial upon hospital admission (0 hours) and 24 hours post admission across 6 U.S. sites. We assayed samples for 21 inflammatory mediators and 7 markers associated with endothelial function and damage. We performed hierarchical clustering analysis (HCA) of these biomarkers of the immune response and endothelial injury. Regression analysis was used to control for differences across study and to assess any association with prehospital plasma resuscitation.RESULTSHCA distinguished two patient clusters with different injury patterns and outcomes. Patients in cluster A had greater injury severity and incidence of blunt trauma, traumatic brain injury, and mortality. Cluster A patients that received prehospital plasma showed improved 30-day survival. Prehospital plasma did not improve survival in cluster B patients. In an adjusted analysis of the most seriously injured patients, prehospital plasma was associated with an increase in adiponectin, IL-1ß, IL-17A, IL-23, and IL-17E upon admission, and a reduction in syndecan-1, TM, VEGF, IL-6, IP-10, MCP-1, and TNF-α, and an increase in IL-33, IL-21, IL-23, and IL-17E 24 hours later.CONCLUSIONPrehospital plasma may ameliorate immune dysfunction and the endotheliopathy of trauma. These effects of plasma may contribute to improved survival in injured patients.TRIAL REGISTRATIONNCT01818427.FUNDINGDepartment of Defense; National Institutes of Health, U.S. Army.

Paleozoic diversification of terrestrial chitin-degrading bacterial lineages.

BACKGROUND: Establishing the divergence times of groups of organisms is a major goal of evolutionary biology. This is especially challenging for microbial lineages due to the near-absence of preserved physical evidence (diagnostic body fossils or geochemical biomarkers). Horizontal gene transfer (HGT) can serve as a temporal scaffold between microbial groups and other fossil-calibrated clades, potentially improving these estimates. Specifically, HGT to or from organisms with fossil-calibrated age estimates can propagate these constraints to additional groups that lack fossils. While HGT is common between lineages, only a small subset of HGT events are potentially informative for dating microbial groups. RESULTS: Constrained by published fossil-calibrated studies of fungal evolution, molecular clock analyses show that multiple clades of Bacteria likely acquired chitinase homologs via HGT during the very late Neoproterozoic into the early Paleozoic. These results also show that, following these HGT events, recipient terrestrial bacterial clades likely diversified ~ 300-500 million years ago, consistent with established timescales of arthropod and plant terrestrialization. CONCLUSIONS: We conclude that these age estimates are broadly consistent with the dispersal of chitinase genes throughout the microbial world in direct response to the evolution and ecological expansion of detrital-chitin producing groups. The convergence of multiple lines of evidence demonstrates the utility of HGT-based dating methods in microbial evolution. The pattern of inheritance of chitinase genes in multiple terrestrial bacterial lineages via HGT processes suggests that these genes, and possibly other genes encoding substrate-specific enzymes, can serve as a "standard candle" for dating microbial lineages across the Tree of Life.

Methane cycling. Nonequilibrium clumped isotope signals in microbial methane.

Methane is a key component in the global carbon cycle, with a wide range of anthropogenic and natural sources. Although isotopic compositions of methane have traditionally aided source identification, the abundance of its multiply substituted "clumped" isotopologues (for example, (13)CH3D) has recently emerged as a proxy for determining methane-formation temperatures. However, the effect of biological processes on methane's clumped isotopologue signature is poorly constrained. We show that methanogenesis proceeding at relatively high rates in cattle, surface environments, and laboratory cultures exerts kinetic control on (13)CH3D abundances and results in anomalously elevated formation-temperature estimates. We demonstrate quantitatively that H2 availability accounts for this effect. Clumped methane thermometry can therefore provide constraints on the generation of methane in diverse settings, including continental serpentinization sites and ancient, deep groundwaters.

Measurement of a doubly substituted methane isotopologue, ¹³CH3D, by tunable infrared laser direct absorption spectroscopy.

Methane is an important energy resource and significant long-lived greenhouse gas. Carbon and hydrogen isotope ratios have been used to better constrain the sources of methane but interpretations based on these two parameters alone can often be inconclusive. The precise measurement of a doubly substituted methane isotopologue, (13)CH3D, is expected to add a critical new dimension to source signatures by providing the apparent temperature at which methane was formed or thermally equilibrated. We have developed a new method to precisely determine the relative abundance of (13)CH3D by using tunable infrared laser direct absorption spectroscopy (TILDAS). The TILDAS instrument houses two continuous wave quantum cascade lasers; one tuned at 8.6 µm to measure (13)CH3D, (12)CH3D, and (12)CH4, and the other at 7.5 µm to measure (13)CH4. With the use of an astigmatic Herriott cell with an effective path length of 76 m, a precision of 0.2‰ (2σ) was achieved for the measurement of (13)CH3D abundance in ca. 10 mL STP (i.e., 0.42 mmol) pure methane samples. Smaller quantity samples (ca. 0.5 mL STP) can be measured at lower precision. The accuracy of the Δ(13)CH3D measurement is 0.7‰ (2σ), evaluated by thermally equilibrating methane with a range of δD values. The precision of ±0.2‰ corresponds to uncertainties of ±7 °C at 25 °C and ±20 °C at 200 °C for estimates of apparent equilibrium temperatures. The TILDAS instrument offers a simple and precise method to determine (13)CH3D in natural methane samples to distinguish geological and biological sources of methane in the atmosphere, hydrosphere, and lithosphere.