Fibrinolytic Activation in Patients with Progressive Intracranial Hemorrhage after Traumatic Brain Injury.

Progression of intracranial hemorrhage (PICH) is a significant cause of secondary brain injury in patients with traumatic brain injury (TBI). Previous studies have implicated a variety of mediators that contribute to PICH. We hypothesized that patients with PICH would display either a hypocoagulable state, hyperfibrinolysis, or both. We conducted a prospective study of adult trauma patients with isolated TBI. Blood was obtained for routine coagulation assays, platelet count, fibrinogen, thrombelastography, markers of thrombin generation, and markers of fibrinolysis at admission and 6, 12, 24, and 48 h. Univariate analyses were performed to compare baseline characteristics between groups. Linear regression models were created, adjusting for baseline differences, to determine the relationship between individual assays and PICH. One hundred forty-one patients met entry criteria, of whom 71 had hemorrhage progression. Patients with PICH had a higher Injury Severity Score and Abbreviated Injury Scale score (head), a lower Glasgow Coma Scale score, and lower plasma sodium on admission. Patients with PICH had higher D-dimers on admission. After adjusting for baseline differences, elevated D-dimers remained significantly associated with PICH compared to patients without PICH at admission. Hypocoagulation was not significantly associated with PICH in these patients. The association between PICH and elevated D-dimers early after injury suggests that fibrinolytic activation may contribute to PICH in patients with TBI.

Tranexamic acid administration in the field does not affect admission thromboelastography after traumatic brain injury.

BACKGROUND: No Food and Drug Administration-approved medication improves outcomes following traumatic brain injury (TBI). A forthcoming clinical trial that evaluated the effects of two prehospital tranexamic acid (TXA) dosing strategies compared with placebo demonstrated no differences in thromboelastography (TEG) values. We proposed to explore the impact of TXA on markers of coagulation and fibrinolysis in patients with moderate to severe TBI. METHODS: Data were extracted from a placebo-controlled clinical trial in which patients 15 years or older with TBI (Glasgow Coma Scale, 3-12) and systolic blood pressure of ≥90 mm Hg were randomized prehospital to receive placebo bolus/placebo infusion (placebo), 1 g of TXA bolus/1 g of TXA infusion (bolus maintenance), or 2 g of TXA bolus/placebo infusion (bolus only). Thromboelastography was performed, and coagulation measures including prothrombin time, activated partial thromboplastin time, international ratio, fibrinogen, D-dimer, plasmin-antiplasmin (PAP), thrombin antithrombin, tissue plasminogen activator, and plasminogen activator inhibitor 1 were quantified at admission and 6 hours later. RESULTS: Of 966 patients receiving study drug, 700 had laboratory tests drawn at admission and 6 hours later. There were no statistically significant differences in TEG values, including LY30, between groups (p > 0.05). No differences between prothrombin time, activated partial thromboplastin time, international ratio, fibrinogen, thrombin antithrombin, tissue plasminogen activator, and plasminogen activator inhibitor 1 were demonstrated across treatment groups. Concentrations of D-dimer in TXA treatment groups were less than placebo at 6 hours (p < 0.001). Concentrations of PAP in TXA treatment groups were less than placebo on admission (p < 0.001) and 6 hours (p = 0.02). No differences in D-dimer and PAP were observed between bolus maintenance and bolus only. CONCLUSION: While D-dimer and PAP levels reflect a lower degree of fibrinolysis following prehospital administration of TXA when compared with placebo in a large prehospital trial of patients with TBI, TEG obtained on admission and 6 hours later did not demonstrate any differences in fibrinolysis between the two TXA dosing regimens and placebo. LEVEL OF EVIDENCE: Diagnostic test, level III.

Early Tranexamic Acid Administration After Traumatic Brain Injury Is Associated With Reduced Syndecan-1 and Angiopoietin-2 in Patients With Traumatic Intracranial Hemorrhage.

OBJECTIVE: To evaluate the effect of early tranexamic acid (TXA) administration on circulating markers of endotheliopathy. SETTING: Twenty trauma centers in the United States and Canada. PARTICIPANTS: Patients with moderate-to-severe traumatic brain injury (TBI) (MS-TBI) and intracranial hemorrhage who were not in shock (systolic blood pressure ≥90 mm Hg). DESIGN: TXA (2 g) or placebo administered prior to hospital arrival, less than 2 hours postinjury. Blood samples and head computed tomographic scan collected upon arrival. Plasma markers measured using Luminex analyte platform. Differences in median marker levels evaluated using t tests performed on log-transformed variables. Comparison groups were TXA versus placebo and less than 45 minutes versus 45 minutes or more from time of injury to treatment administration. MAIN MEASURES: Plasma levels of angiopoietin-1, angiopoietin-2, syndecan-1, thrombomodulin, thrombospondin-2, intercellular adhesion molecule 1, vascular adhesion molecule 1. RESULTS: Demographics and Injury Severity Score were similar between the placebo (n = 129) and TXA (n = 158) groups. Levels of syndecan-1 were lower in the TXA group (median [interquartile range or IQR] = 254.6 pg/mL [200.7-322.0] vs 272.4 pg/mL [219.7-373.1], P = .05. Patients who received TXA less than 45 minutes postinjury had significantly lower levels of angiopoietin-2 (median [IQR] = 144.3 pg/mL [94.0-174.3] vs 154.6 pg/mL [110.4-209.8], P = .05). No differences were observed in remaining markers. CONCLUSIONS: TXA may inhibit early upregulation of syndecan-1 and angiopoietin-2 in patients with MS-TBI, suggesting attenuation of protease-mediated vascular glycocalyx breakdown. The findings of this exploratory analysis should be considered preliminary and require confirmation in future studies.

Use of bilobed partial resuscitative endovascular balloon occlusion of the aorta is logistically superior in prolonged management of a highly lethal aortic injury.

BACKGROUND: Resuscitative endovascular balloon occlusion of the aorta (REBOA) is a viable technique for management of noncompressible torso hemorrhage. The major limitation of the current unilobed fully occlusive REBOA catheters is below-the-balloon ischemia-reperfusion complications. We hypothesized that partial aortic occlusion with a novel bilobed partial (p)REBOA-PRO would result in the need for less intraaortic balloon adjustments to maintain a distal goal perfusion pressure as compared with currently available unilobed ER-REBOA. METHODS: Anesthetized (40-50 kg) swine randomized to control (no intervention), ER-REBOA, or pREBOA-PRO underwent supraceliac aortic injury. The REBOA groups underwent catheter placement into zone 1 with initial balloon inflation to full occlusion for 10 minutes followed by gradual deflation to achieve and subsequently maintain half of the baseline below-the-balloon mean arterial pressure (MAP). Physiologic data and blood samples were collected at baseline and then hourly. At 4 hours, the animals were euthanized, total blood loss and urine output were recorded, and tissue samples were collected. RESULTS: Baseline physiologic data and basic laboratories were similar between groups. Compared with control, interventions similarly prolonged survival from a median of 18 minutes to over 240 minutes with comparable mortality trends. Blood loss was similar between partial ER-REBOA (41%) and pREBOA-PRO (51%). Partial pREBOA-PRO required a significantly lower number of intraaortic balloon adjustments (10 ER-REBOA vs. 3 pREBOA-PRO, p < 0.05) to maintain the target below-the-balloon MAP. The partial ER-REBOA group developed significantly increased hypercapnia, fibrin clot formation on TEG, liver inflammation, and IL-10 expression compared with pREBOA-PRO. CONCLUSION: In this highly lethal aortic injury model, use of bilobed pREBOA-PRO for a 4-hour partial aortic occlusion was logistically superior to unilobed ER-REBOA. It required less intraaortic balloon adjustments to maintain target MAP and resulted in less inflammation.

γ' fibrinogen levels are associated with blood clot strength in traumatic brain injury patients.

BACKGROUND: γ' fibrinogen is an alternatively-spliced fibrinogen variant that displays different coagulation parameters in vitro than the major form of fibrinogen. Purified γ' fibrinogen has slower clotting kinetics than unfractionated fibrinogen, but forms clots that are stronger and resistant to fibrinolysis. However, these properties have only been investigated in human populations in a limited number of studies. We therefore performed a retrospective analysis to test the hypothesis that γ' fibrinogen levels influence coagulation in vivo. METHODS: In the present study, we utilized blood samples that were collected from traumatic brain injury patients to probe the relationship between γ' fibrinogen levels and traditional coagulation parameters. RESULTS: The results show that the levels of γ' fibrinogen were inversely associated with clotting kinetics, indicated by a shortened INR. In addition, the levels of γ' fibrinogen were associated with stronger clots by thrombelastography. However, these changes were not associated with significant changes in hemorrhage progression. CONCLUSIONS: These findings verify that γ' fibrinogen properties observed in purified systems result in similar properties in a clinical setting, and may affect coagulation.

The effects of cryopreserved red blood cell transfusion on tissue oxygenation in obese trauma patients.

BACKGROUND: Low tissue oxygenation (StO2) is associated with poor outcomes in obese trauma patients. A novel treatment could be the transfusion of cryopreserved packed red blood cells (CPRBCs), which the in vitro biochemical profile favors red blood cell (RBC) function. We hypothesized that CPRBC transfusion improves StO2 in obese trauma patients. METHODS: Two hundred forty-three trauma patients at five Level I trauma centers who required RBC transfusion were randomized to receive one to two units of liquid packed RBCs (LPRBCs) or CPRBCs. Demographics, injury severity, StO2, outcomes, and biomarkers of RBC function were compared in nonobese (body mass index [BMI] < 30) and obese (BMI ≥ 30) patients. StO2 was also compared between obese patients with BMI of 30 to 34.9 and BMI ≥ 35. StO2 was normalized and expressed as % change after RBC transfusion. A p value less than 0.05 indicated significance. RESULTS: Patients with BMI less than 30 (n = 141) and BMI of 30 or greater (n = 102) had similar Injury Severity Score, Glasgow Coma Scale, and baseline StO2. Plasma levels of free hemoglobin, an index of RBC lysis, were lower in obese patients after CPRBC (125 [72-259] µg/mL) versus LPRBC transfusion (230 [178-388] µg/mL; p < 0.05). StO2 was similar in nonobese patients regardless of transfusion type, but improved in obese patients who received CPRBCs (104 ± 1%) versus LPRPCs (99 ± 1%, p < 0.05; 8 hours after transfusion). Subanalysis showed improved StO2 after CPRBC transfusion was specific to BMI of 35 or greater, starting 5 hours after transfusion (p < 0.05 vs. LPRBCs). CPRBCs did not improve clinical outcomes in either group. CONCLUSION: CPRBC transfusion is associated with increased StO2 and lower free hemoglobin levels in obese trauma patients, but did not improve clinical outcomes. Future studies are needed to determine if CPRBC transfusion in obese patients attenuates hemolysis to improve StO2. LEVEL OF EVIDENCE: Therapeutic, level IV.

Thrombelastography-Based Dosing of Enoxaparin for Thromboprophylaxis in Trauma and Surgical Patients: A Randomized Clinical Trial.

Importance: Prophylactic enoxaparin is used to prevent venous thromboembolism (VTE) in surgical and trauma patients. However, VTE remains an important source of morbidity and mortality, potentially exacerbated by antithrombin III or anti-Factor Xa deficiencies and missed enoxaparin doses. Recent data suggest that a difference in reaction time (time to initial fibrin formation) greater than 1 minute between heparinase and standard thrombelastogram (TEG) is associated with a decreased risk of VTE. Objective: To evaluate the effectiveness of TEG-adjusted prophylactic enoxaparin dosing among trauma and surgical patients. Design, Setting, and Participants: This randomized clinical trial, conducted from October 2012 to May 2015, compared standard dosing (30 mg twice daily) with TEG-adjusted enoxaparin dosing (35 mg twice daily) for 185 surgical and trauma patients screened for VTE at 3 level I trauma centers in the United States. Main Outcomes and Measures: The incidence of VTE, bleeding complications, anti-Factor Xa deficiency, and antithrombin III deficiency. Results: Of the 185 trial participants, 89 were randomized to the control group (median age, 44.0 years; 55.1% male) and 96 to the intervention group (median age, 48.5 years; 74.0% male). Patients in the intervention group received a higher median enoxaparin dose than control patients (35 mg vs 30 mg twice daily; P < .001). Anti-Factor Xa levels in intervention patients were not higher than levels in control patients until day 6 (0.4 U/mL vs 0.21 U/mL; P < .001). Only 22 patients (11.9%) achieved a difference in reaction time greater than 1 minute, which was similar between the control and intervention groups (10.4% vs 13.5%; P = .68). The time to enoxaparin initiation was similar between the control and intervention groups (median [range] days, 1.0 [0.0-2.0] vs 1.0 [1.0-2.0]; P = .39), and the number of patients who missed at least 1 dose was also similar (43 [48.3%] vs 54 [56.3%]; P = .30). Rates of VTE (6 [6.7%] vs 6 [6.3%]; P > .99) were similar, but the difference in bleeding complications (5 [5.6%] vs 13 [13.5%]; P = .08) was not statistically significant. Antithrombin III and anti-Factor Xa deficiencies and hypercoagulable TEG parameters, including elevated coagulation index (>3), maximum amplitude (>74 mm), and G value (>12.4 dynes/cm2), were prevalent in both groups. Identified risk factors for VTE included older age (61.0 years vs 46.0 years; P = .04), higher body mass index (calculated as weight in kilograms divided by height in meters squared; 30.6 vs 27.1; P = .03), increased Acute Physiology and Chronic Health Evaluation II score (8.5 vs 7.0; P = .03), and increased percentage of missed doses per patient (14.8% vs 2.5%; P = .05). Conclusions and Relevance: The incidence of VTE was low and similar between groups; however, few patients achieved a difference in reaction time greater than 1 minute. Antithrombin III deficiencies and hypercoagulable TEG parameters were prevalent among patients with VTE. Low VTE incidence may be due to an early time to enoxaparin initiation and an overall healthier and less severely injured study population than previously reported. Trial Registration: clinicaltrials.gov Identifier: NCT00990236.

Transfusion of cryopreserved packed red blood cells is safe and effective after trauma: a prospective randomized trial.

OBJECTIVES: To determine the safety and efficacy of cryopreserved packed red blood cell (CPRBC) transfusion in trauma patients. BACKGROUND: Liquid packed red blood cells (LPRBCs) have an abbreviated shelf-life and worsening storage lesion with age. CPRBCs are frozen 2 to 6 days after donation, stored up to 10 years, and are available for 14 days after thawing and washing. CPRBCs can be utilized in diverse settings, but the effect on clinical outcomes is unknown. METHODS: We performed a prospective, randomized, double-blind study at 5 level 1 trauma centers. Stable trauma patients requiring transfusion were randomized to young LPRBCs (≤14 storage days), old LPRBCs (>14 storage days), or CPRBCs. Tissue oxygenation (StO2), biochemical and inflammatory mediators were measured, and clinical outcomes were determined. RESULTS: Two hundred fifty-six patients with well-matched injury severity and demographics (P > 0.2) were randomized (84 young, 86 old, and 86 CPRBCs). Pretransfusion and final hematocrits were similar (P > 0.68). Patients in all groups received the same number of units postrandomization (2 [1-4]; P > 0.05). There was no difference in the change in tissue oxygenation between groups. CPRBCs contained less α2-macrogobulin, haptoglobin, C-reactive protein, and serum amyloid P (P < 0.001). Organ failure, infection rate, and mortality did not differ between groups (P > 0.2). CONCLUSIONS: Transfusion of CPRBCs is as safe and effective as transfusion of young and old LPRBCs and provides a mechanism to deliver PRBCs in a wide variety of settings.

Effect of ascorbic acid concentrations on hemodynamics and inflammation following lyophilized plasma transfusion.

BACKGROUND: Compared with lyophilized plasma (LP) buffered with other acids, LP with ascorbic acid (AA) attenuates systemic inflammation and DNA damage in a combat relevant polytrauma swine model. We hypothesize that increasing concentrations of AA in transfused LP will be safe, will be hemodynamically well tolerated, and will attenuate systemic inflammation following polytraumatic injury and hemorrhage in swine. METHODS: This prospective, randomized, blinded study involved 52 female swine. Forty animals were subjected to our validated polytrauma model and resuscitated with LP. Baseline control sham (n = 6), operative control sham (n = 6), low-AA (n = 10), medium-AA (n = 10), high-AA (n = 10) groups, and a hydrochloric acid control (HCL, n = 10) were randomized. Hemodynamics, thrombelastography, and blood chemistries were assessed. Inflammatory cytokines (tumor necrosis factor α, interleukin 6 [IL-6], C-reactive protein, and IL-10) and DNA damage were measured at baseline, 2 hours, and 4 hours after liver injury. Significance was set at p < 0.05, with a Bonferroni correction for multiple comparisons. RESULTS: Hemodynamics, shock, and blood loss were similar between groups. All animals had robust procoagulant activity 2 hours following liver injury. Inflammation was similar between groups at baseline, and AA groups remained similar to HCL following liver injury. IL-6 and tumor necrosis factor α were increased at 2 hours and 4 hours compared with baseline within all groups (p < 0.008). DNA damage increased at 2 hours compared with baseline in all groups (p < 0.017) and further increased at 4 hours compared with baseline in HCL, low-, and high-AA groups (p < 0.005). C-reactive protein was similar between and within groups. IL-10 increased at 2 hours compared with baseline in low- and high-AA groups and remained elevated at 4 hours compared with baseline in the low-AA group (all, p < 0.017). CONCLUSION: Concentrations of AA were well tolerated and did not diminish the procoagulant activity of LP. Within our tested range of concentrations, AA can safely be used to buffer LP.

Reconstitution fluid type does not affect pulmonary inflammation or DNA damage following infusion of lyophilized plasma.

BACKGROUND: Dysfunctional inflammation following traumatic hemorrhage can lead to multiple-organ failure and death. In our polytrauma swine model, lyophilized plasma (LP) reconstituted with sterile water and ascorbic acid suppressed systemic inflammation and attenuated DNA damage. However, it remains unknown whether the inflammatory response is affected by the type of fluid used to reconstitute LP. We hypothesized that common resuscitation fluids such as normal saline (LP-NS), lactated Ringer's solution (LP-LR), Hextend (LP-HX), or sterile water (LP-SW) would yield similar inflammation profiles and DNA damage following LP reconstitution and transfusion. METHODS: This was a randomized, prospective, blinded animal study. LP was reconstituted to 50% of original volume with NS, LR, HX, or SW buffered with 15-mM ascorbic acid. Forty swine were subjected to a validated model of polytrauma, hemorrhagic shock, and Grade V liver injury and resuscitated with LP. Serum interleukin 6 (IL-6), IL-10, plasma C-reactive protein, and 8-hydroxy-2-deoxyguanosine concentrations were assessed for systemic inflammation and DNA damage at baseline, 2 hours, and 4 hours following liver injury. Lung inflammation was evaluated by Real Time Polymerize Chain Reaction (RT-PCR). RESULTS: Reconstituted LP pH was similar between groups before resuscitation. IL-6 and IL-10 increased at 2 hours and 4 hours compared with baseline in all groups (p < 0.017). DNA damage increased at 2 hours and 4 hours compared with baseline and from 2 hours to 4 hours in the LP-NS, LP-LR, and LP-SW groups (all p < 0.017). Animals resuscitated with LP-HX not only demonstrated increased DNA damage at 4 hours versus baseline but also had the lowest C-reactive protein level at 2 hours and 4-hours (p < 0.017). Overall, differences between groups were similar for DNA damage and lung inflammation. CONCLUSION: Reconstitution fluid type does not affect inflammatory cytokine profiles or DNA damage following LP transfusion in this swine polytrauma model. Based on universal availability, these data suggest that sterile water is the most logical choice for LP reconstitution in humans. LEVEL OF EVIDENCE: Prognostic, level II.

Contribution of a membrane estrogen receptor to the estrogenic regulation of body temperature and energy homeostasis.

The hypothalamus is a key region of the central nervous system involved in the control of homeostasis, including energy and core body temperature (Tc). 17ß-Estradiol (E2) regulates Tc, in part, via actions in the basal hypothalamus and preoptic area. E2 primarily controls hypothalamic functions via the nuclear steroid receptors, estrogen receptor α/ß. However, we have previously described an E2-responsive, Gq-coupled membrane receptor that reduces the postsynaptic inhibitory γ-aminobutyric acid-ergic tone and attenuates postovariectomy body weight gain in female guinea pigs through the administration of a selective Gq-mER ligand, STX. To determine the role of Gq-mER in regulating Tc, energy and bone homeostasis, ovariectomized female guinea pigs, implanted ip with temperature probes, were treated with STX or E2 for 7-8 wk. Tc was recorded for 4 wk, whereas food intake and body weight were monitored daily. Bone density and fat accumulation were determined postmortem. Both E2 and STX significantly reduced Tc in the females compared with controls. STX, similar to E2, reduced food intake and fat accumulation and increased tibial bone density. Therefore, a Gq-mER-coupled signaling pathway appears to be involved in maintaining homeostatic functions and may constitute a novel therapeutic target for treatment of hypoestrogenic symptoms.

17Beta-estradiol regulation of T-type calcium channels in gonadotropin-releasing hormone neurons.

T-type calcium channels are responsible for generating low-threshold spikes that facilitate burst firing and neurotransmitter release in neurons. Gonadotropin-releasing hormone (GnRH) neurons exhibit burst firing, but the underlying conductances are not known. Previously, we found that 17beta-estradiol (E2) increases T-type channel expression and excitability of hypothalamic arcuate nucleus neurons. Therefore, we used ovariectomized oil- or E2-treated EGFP (enhanced green fluorescent protein)-GnRH mice to explore the expression and E2 regulation of T-type channels in GnRH neurons. Based on single-cell reverse transcriptase-PCR and real-time PCR quantification of the T-type channel alpha(1) subunits, we found that all three subunits were expressed in GnRH neurons, with expression levels as follows: Cav3.3 > or = Cav3.2 > Cav3.1. The mRNA expression of the three subunits was increased with surge-inducing levels of E2 during the morning. During the afternoon, Cav3.3 mRNA expression remained elevated, whereas Cav3.1 and Cav3.2 were decreased. The membrane estrogen receptor agonist STX increased the expression of Cav3.3 but not Cav3.2 in GnRH neurons. Whole-cell patch recordings in GnRH neurons revealed that E2 treatment significantly augmented T-type current density at both time points and increased the rebound excitation during the afternoon. Although E2 regulated the mRNA expression of all three subunits in GnRH neurons, the increased expression combined with the slower inactivation kinetics of the T-type current indicates that Cav3.3 may be the most important for bursting activity associated with the GnRH/LH (luteinizing hormone) surge. The E2-induced increase in mRNA expression, which depends in part on membrane-initiated signaling, leads to increased channel function and neuronal excitability and could be a mechanism by which E2 facilitates burst firing and cyclic GnRH neurosecretion.

GPR30 contributes to estrogen-induced thymic atrophy.

The mechanisms by which prolonged estrogen exposures, such as estrogen therapy and pregnancy, reduce thymus weight, cellularity, and CD4 and CD8 phenotype expression, have not been well defined. In this study, the roles played by the membrane estrogen receptor, G protein-coupled receptor 30 (GPR30), and the intracellular estrogen receptors, estrogen receptor alpha (ERalpha) and beta (ERbeta), in 17beta-estradiol (E2)-induced thymic atrophy were distinguished by construction and the side-by-side comparison of GPR30-deficient mice with ERalpha and ERbeta gene-deficient mice. Our study shows that whereas ERalpha mediated exclusively the early developmental blockage of thymocytes, GPR30 was indispensable for thymocyte apoptosis that preferentially occurs in T cell receptor beta chain(-/low) double-positive thymocytes. Additionally, G1, a specific GPR30 agonist, induces thymic atrophy and thymocyte apoptosis, but not developmental blockage. Finally, E2 treatment attenuates the activation of nuclear factor-kappa B in CD25(-)CD4(-)CD8(-) double-negative thymocytes through an ERalpha-dependent yet ERbeta- and GPR30-independent pathway. Differential inhibition of nuclear factor-kappaB by ERalpha and GPR30 might underlie their disparate physiological effects on thymocytes. Our study distinguishes, for the first time, the respective contributions of nuclear and membrane E2 receptors in negative regulation of thymic development.