ALBERT

Description: Background. It has been observed that trauma patients often display an elevated procoagulant activity. This activity has been hypothesized to be caused in part by tissue factor (TF) located on the endothelium, blood cells and microparticles. We also observed in our previous studies that trauma patients with thermal, blunt and penetrating injuries have active FIXa and FXIa in their plasma, often persistent for several days to weeks. What is not known is the effect of injury type and severity on TF release after trauma. In the current study we evaluated how the severity of an injury with or without accompanying shock affects the frequency and concentration of TF, FIXa and FXIa in plasma from trauma patients. Methods. Eighty trauma patients presenting to a major urban level 1 trauma center (47 with blunt trauma and 33 with penetrating trauma) were enrolled. 62 patients were male and 18 female. The age of patients varied between 18 and 90 years (average 42.6±19.2 years) and the Injury Severity Score (ISS) varied between 1 and 75 (average 19.3±17.2). Blood was collected immediately upon emergency department arrival prior to any resuscitation or blood product transfusion. For analysis our cohort was divided into 4 groups (20 patients/group) based on their ISS and presence of shock (base deficit BD): Group 1: Non-severe injury, no shock (ISS≤15; BD〉-6) Group 2: Non-severe injury, with shock (ISS≤15; BD≤-6) Group 3: Severe injury, no shock (ISS〉15; BD〉-6) Group 4: Severe injury, with shock (ISS〉15; BD≤-6) Citrate plasma was prepared, frozen and stored at -80°C. No additional freeze/thaw cycles were involved prior to FXIa, FIXa and TF activity assays. The assays were based on a response of thrombin generation to corresponding monoclonal inhibitory antibodies (αFXIa-2, αFIXa-91 and αTF-5, respectively; all at 0.1 mg/mL). Results. The frequency of TF in less injured patients was relatively low (table) and increased by at least 3-fold in patients with severe injury (groups 3 and 4). The concentration of TF varied in a wide range in all 4 groups, but was the lowest in group 1 patient samples and the highest in groups 3 and 4. FXIa was observed in 91% of plasma samples and was high in all 4 groups of patients. Similar to TF, the concentration of FXIa varied in a wide range in the entire study population. Unlike TF however, FXIa was present in patients with shock at a higher concentration and did not correlate with the severity of injury. Only 9 of 20 group 1 plasma samples had detectable FIXa, whereas in groups 2-4 only 1 or 2 plasma samples had no FIXa activity. The median concentration of this protein varied in a relatively narrow range between all 4 groups, although the lowest was observed in group 1. For the entire cohort, there was a good correlation between FIXa and FXIa concentrations (R2 =0.33), but no correlation between TF and FIXa or FIXa. These data suggest that FXIa in trauma patient blood is generated primarily through the contact pathway, although the input of the TF pathway to FXI activation cannot be excluded. Table 1.Patient GroupTFFXIaFIXaFrequencyConcentration (pM)FrequencyConcentration (pM)FrequencyConcentration (pM)RangeMedianRangeMedianRangeMedian14/20 (20%)0.1 - 〉6.40.2818/20 (90%)0.9 - 〉647.39/20 (45%)73 - 〉100017523/20 (15%)0.5 - 〉6.41.0019/20 (95%)3.2- 〉6423.418/20 (90%)38 - 〉1000270312/20 (60%)0.3 - 〉6.43.5017/20 (85%)0.7 - 〉6411.118/20 (90%)75 - 〉1000252414/20 (70%)0.2 - 〉6.42.4819/20 (95%)1.8 - 〉6426.519/20 (95%)26 - 〉1000205Overall33/80 (41%)0.1 - 〉6.41.0173/80 (91%)0.7 - 〉10017.064/80 (80%)26 - 〉1000250 Conclusions. 1) Frequency and concentration of TF is higher in patients with a higher trauma severity, but it is independent of shock; 2) The vast majority of plasma samples from trauma patients contain active FIXa and FXIa; 3) Concentration of FXIa is higher in patients with shock and does not appear to be affected by the trauma severity. Taken together, these data suggest separate mechanisms for contact pathway activation after injury driven by shock and TF pathway activation driven by tissue injury. Disclosures No relevant conflicts of interest to declare.

Description: BACKGROUND: Trauma is the leading cause of death in individuals under the age of 44. Acute traumatic coagulopathy (ATC) occurs when severe injury is combined with shock and results in a hypocoagulable state resulting in increased bleeding, increased resuscitation requirements and worsened outcomes including a 4-fold increase in mortality. Our group has implicated protein C activation as a likely mechanism for ATC. We have evaluated whether activated protein C (APC) induces coagulopathy through cleavage of factor (f)V/Va by systematically evaluating 4 groups of increasing level of traumatic coagulopathy. METHODS: From a large cohort of plasma samples collected from trauma patients at time of admission to a major level 1 trauma sample, 80 plasma samples distributed equally between the following trauma severity groupings were selected: Group 1: ISS ≤ 15 and BD 〉 -6 (less injured, no shock); Group 2: ISS ≤ 15 and BD ≤ -6 (less injured, with shock); Group 3:ISS 〉 15 and BD 〉 -6 (more injured, no shock); Group 4:ISS 〉 15 and BD ≤ -6 (more injured, with shock). Each plasma sample was evaluated for plasma composition (functional activity of fII, fV, fVII, fVIII, fIX, fX, antithrombin, tissue factor pathway inhibitor, protein C), plasmin-antiplasmin complex (PAP), α-thrombin-antithrombin complex (TAT), and mathematically modeled thrombin generation. Quantitative Western blot analyses were performed using an anti-fV antibody directed at the fV heavy chain (residues 307-506). Intact fV levels were quantified via densitometry to determine whether levels were the within range characterizing healthy individuals and the presence of APC derived and other degradation products assessed via mobility comparison to purified standards. Demographic, resuscitation, and outcomes data were collected in parallel. RESULTS: All data were quantitated and evaluated for having concentrations above and/or below the normal range for each analyte. For PAP, Groups 3 (18/20) and 4 (17/20) had a greater percentage of individuals over Groups 1 (5/20) and 2 (6/20) that were above the normal range. Groups 3 (12/20) and 4 (12/20) also had a higher percentage of detectable α-TAT over Groups 1 (5/20) and 2 (2/20). For fV antigen, Group 4 had 16/20 individuals that were below the lower limit of normal. Groups 1, 2 and 3 were all similar with 5/20 below the limit of normal but had 3-5/20 above the upper limit of normal. Products associated with activated protein C were found (fV 30kD fragment) but also other cleavage products not associated with APC were observed. There was more fV fragmentation in Group 4, with the 30kD fragment being detected in 10 individuals. No detectable fV 30kD fragments were detected in either Groups 1 or 2. In terms of the overall plasma composition there is a large variation in functional activity, especially fVIII. Group 4 showed a pattern of all factors reduced except for TFPI versus Groups 1,2, 3. These pro and anticoagulant differences between individuals results in heterogenous modeled thrombin generation profiles with the majority all being procoagulant versus a normal control. Group 4 had a higher percentage of individuals that had a prolonged clot time and a pattern towards diminished thrombin generation. CONCLUSIONS: These data suggest that the combination of injury and shock were associated with altered PAP, TAT and functional activity changes of the plasma composition resulting in hemorrhagic thrombin generation profiles. In addition the most severely injured and shocked patients had less fVa activity, less intact fV and increased degradation of fV/fVa suggesting the presence of increased APC activity in mediating ATC. Disclosures No relevant conflicts of interest to declare.