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Background Studies associating coagulopathic changes with burn injury have relied on limited tests such as partial thromboplastin time (PTT) and international normalized ratio (INR). Understanding the clotting dynamics and associated risk factors after burn injury could influence management. This work aimed to identify real-time changes in coagulation after burn injury not indicated by PTT or INR alone. Materials and methods Nine burn-injured patients at a regional burn center were enrolled for blood collection at admission and set intervals over 96 h. Patient demographics, management, and laboratory data (PTT and INR) were collected. Plasma assays determined factors II, V, VII, VIII, IX, X, XI, antithrombin, and protein C functional activity as well as PAP, D-Dimer, fibrin monomer, TFPI, IL-1b, IL-6, IL-10, IL-12p.70, and TNF-α concentrations. Results Overall, five patients died. These patients had higher mortality scores and were more acidotic. All patients had normal coagulation studies (INR < 1.5, PTT < 45 s) within 24 h of admission, and only two were abnormal after. Increased factor VIII and IX activity were identified in seven patients at admission. Decreased antithrombin and protein C activity were seen in all patients. Patients had increased PAP, D-Dimer, and fibrin monomer concentrations throughout their hospital course. At admission, increased fold changes were seen in IL-6 (2.5–117) and IL-10 (2.4–32), whereas IL-1b and TNF-α levels were depressed in all patients. Conclusions Extensive changes not identified by PTT or INR were seen after burn injury that may explain perturbed coagulation in these patients. This approach further characterizes the impact thermal injury has on coagulation. [ABSTRACT FROM AUTHOR]

While platelets are well known to play a central role in hemostasis and thrombosis, there is emerging experimental evidence to suggest that they also mediate tumor cell growth, dissemination, and angiogenesis. An increase in platelet number (thrombocytosis) and activity is seen in patients with a wide spectrum of malignancies, and the former is correlated with a decrease in overall survival and poorer prognosis. Preclinical data suggest that circulating tumor cell partnerships with platelets in the blood facilitate tumor metastases through direct interactions and secreted bioactive proteins. Platelets form aggregates with tumor cells, thereby protecting them from host immune surveillance through physical shielding and induction of 'platelet mimicry.' There is also laboratory evidence to suggest that activated platelets interact with cancer cells within the tumor microenvironment through paracrine signaling and direct contact, thereby promoting tumor cell growth and survival. For example, platelets release mediators of both tumor angiogenesis and osteoclast resorption. The interplay between platelets and tumor cells is complex and bidirectional with involvement of multiple other components within the tumor microenvironment, including immune cells, endothelial cells, and the extracellular matrix. We review the role of platelets in tumor progression, emphasizing the opportunity these interactions afford to target platelets and platelet function to improve patient outcomes in the cancer prevention and treatment setting. J. Cell. Physiol. 229: 1005-1015, 2014. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

The underlying cause of thrombosis in a large protein C (PC) deficient Vermont kindred appears to be multicausal and not explained by PC deficiency alone. We evaluated the contribution of coagulation factors to thrombin generation in this population utilizing a mathematical model that incorporates a mechanistic description of the PC pathway. Thrombin generation profiles for each individual were generated with and without the contribution of the PC pathway. Parameters that describe thrombin generation: maximum level (MaxL) and rate (MaxR), their respective times (TMaxL, TMaxR), area under the curve (AUC) and clotting time (CT) were examined in individuals ±PC mutation, ±prothrombin G20210A polymorphism and ±thrombosis history (DVT or PE). This family (n = 364) is shifted towards greater thrombin generation relative to the mean physiologic control. When this family was analyzed with the PC pathway, our results showed that: carriers of the PC mutation (n = 81) had higher MaxL and MaxR and greater AUC (all p<0.001) than non-carriers (n = 283); and individuals with a DVT and/or PE history (n = 13) had higher MaxL (p = 0.005) and greater AUC (p<0.001) than individuals without a thrombosis history (n = 351). These differences were further stratified by gender, with women in all categories generating more thrombin than males. These results show that all individuals within this family with or without PC deficiency have an increased baseline procoagulant potential reflective of increased thrombin generation. In addition, variations within the plasma composition of each individual can further segregate out increased procoagulant phenotypes, with gender-associated plasma compositional differences playing a large role. [ABSTRACT FROM AUTHOR]

Factor (f)Xa is a critical enzyme in blood coagulation that is responsible for the initiation and propagation of thrombin generation. Previously we have shown that analysis of computationally generated thrombin profiles is a tool to investigate hemostasis in various populations. In this study, we evaluate the potential of computationally derived time courses of fXa generation as another approach for investigating thrombotic risk. Utilizing the case (n = 473) and control (n = 426) population from the Leiden Thrombophilia Study and each individual's plasma protein factor composition for fII, fV, fVII, fVIII, fIX, fX, antithrombin and tissue factor pathway inhibitor, tissue factor-initiated total active fXa generation was assessed using a mathematical model. FXa generation was evaluated by the area under the curve (AUC), the maximum rate (MaxR) and level (MaxL) and the time to reach these, TMaxR and TMaxL, respectively. FXa generation was analyzed in the entire populations and in defined subgroups (by sex, age, body mass index, oral contraceptive use). The maximum rates and levels of fXa generation occur over a 10- to 12- fold range in both cases and controls. This variation is larger than that observed with thrombin (3-6 fold) in the same population. The greatest risk association was obtained using either MaxR or MaxL of fXa generation; with an ∼2.2 fold increased risk for individuals exceeding the 90th percentile. This risk was similar to that of thrombin generation(MaxR OR 2.6). Grouping defined by oral contraceptive (OC) use in the control population showed the biggest differences in fXa generation; a >60% increase in the MaxR upon OC use. FXa generation can distinguish between a subset of individuals characterized by overlapping thrombin generation profiles. Analysis of fXa generation is a phenotypic characteristic which may prove to be a more sensitive discriminator than thrombin generation among all individuals. [ABSTRACT FROM AUTHOR]

The function of tissue factor (Tf)-initiated coagulation is hemorrhage control through the formation and maintenance of an impermeable platelet-fibrin barrier. The catalytic processes involved in the clot maintenance function are not well defined, although the rebleeding problems characteristic of individuals with hemophilias A and B suggest a link between specific defects in the Tf-initiated process and defects in the maintenance function. We have previously demonstrated, using a methodology of "flow replacement" (or resupply) of ongoing Tf-initiated reactions with fresh reactants, that procoagulant complexes are produced during Tf-initiated coagulation, which are capable of reinitiating coagulation without input from extrinsic factor Xase activity (Orfeo, T., Butenas, S., Brummel-Ziedins, K. E., and Mann, K. G. (2005)1. Biol. Chem. 280, 42887-42896). Here we used Tf-initiated reactions in normal and hemophilia blood or in their corresponding proteome mixtures as sources of pro-coagulant end products and then varied the resupplying mate- rial to determine the identity of the catalysts that drive the new cycle of thrombin formation. The central findings are as follows: 1) the prothrombinase complex (fVa-fXa-Ca2+-membrane) accumulated during the episode of Tf-initiated coagulation is the primary catalyst responsible for the observed pattern of prothrombin activation after resupply; 2) impairments in intrinsic factor Xase function, i.e. hemophilias A and B, result in an impaired capacity to mount a resupply response; and 3) in normal hemostasis the intrinsic factor Xase function contributes to the durability of the resupply response. [ABSTRACT FROM AUTHOR]

Abstract: Global assays and computational models have advanced research into the realm of individualized profiling of hemostatic states. This brief review will describe one computational approach which utilizes an integrated method that evaluates the dynamics of thrombin generation by defining interactions of the pro and anticoagulant proteins, enzymes and cofactors based upon individualized concentrations of select factors. This plasma composition based computational modeling can provide a mechanism based bridge between empirical global assays of coagulation and individualized risk prediction. [Copyright &y& Elsevier]

Whole genome sequencing of an individual completely devoid of plasma- and platelet-derived factor V (FV) identified 167 variants in his F5 gene including previously identified and damaging missense mutations at rs6027 and Leu90Ser. Because the administration of fresh frozen plasma (FFP) prevents gastrointestinal bleeding in this individual, its effects on his plasma- and platelet-derived FV concentrations were assessed. The patient's plasma FV levels peaked by 2 hours following FFP administration and were undetectable 96 hours later. In contrast, increased platelet-derived FV/Va concentrations were observed within 6 hours, peaked at 24 hours, decreased slowly over 7 days, and originated from megakaryocyte endocytosis and intracellular processing of plasma FV. Ten days after transfusion, no thrombin was generated in a tissue taclor-initiated whole blood clotting assay unless exogenous FV was added, consistent with the complete absence of plasma FV. In marked contrast, release of the patient's platelet-derived FV/Va (7% of normal) following platelet activation resulted in robust thrombin generation, similar to that in an individual with normal plasma- and platelet-derived FV concentrations. Thus, total FV deficiency can be corrected by plasma administration, which partially repletes and sustains the platelet cofactor pool, thereby highlighting the critical role of platelet-derived FV/Va in ensuring hemostatic competence. [ABSTRACT FROM AUTHOR]

Proposed mechanisms of acute traumatic coagulopathy (ATC) include decreased clotting potential due to factor consumption and proteolytic inactivation of factor V (FV) and activated factor V (FVa) by activated protein C (aPC). The role of FV/FVa depletion or inactivation in burn-induced coagulopathy is not well characterized. This study evaluates FV dynamics following burn and nonburn trauma. Burn and trauma patients were prospectively enrolled. Western blotting was performed on admission plasma to quantitate levels of FV antigen and to assess for aPC or other proteolytically derived FV/FVa degradation products. Statistical analysis was performed with Spearman's, Chi-square, Mann–Whitney U test, and logistic regression. Burn (n = 60) and trauma (n = 136) cohorts showed similar degrees of FV consumption with median FV levels of 76% versus 73% (P = 0.65) of normal, respectively. Percent total body surface area (TBSA) was not correlated with FV, nor were significant differences in median FV levels observed between low and high TBSA groups. The injury severity score (ISS) in trauma patients was inversely correlated with FV (ρ = −0.26; P = 0.01) and ISS ≥ 25 was associated with a lower FV antigen level (64% versus. 93%; P = 0.009). The proportion of samples showing proteolysis-derived FV was greater in trauma than burn patients (42% versus. 16%; P = 0.0006). Increasing traumatic injury severity is associated with decreased FV antigen levels, and a greater proportion of trauma patient samples exhibit proteolytically degraded FV fragments. These associations are not present in burns, suggesting that mechanisms underlying FV depletion in burn and nonburn trauma are not identical. [ABSTRACT FROM AUTHOR]

Formation of thrombin is triggered when membrane-localized tissue factor (TF) is exposed to blood. In closed models of this process, thrombin formation displays an initiation phase (low rates of thrombin production cause platelet activation and fibrinogen clotting), a propagation phase (>95% of thrombin production occurs), and a termination phase (prothrombin activation ceases and free thrombin is inactivated). A current controversy centers on whether the TF stimulus requires supplementation from a circulating pool of blood TF to sustain an adequate procoagulant response. We have evaluated the requirement for TF during the progress of the blood coagulation reaction and have extended these analyses to assess the requirement for TF during resupply (‘flow replacement’). Elimination of TF activity at various times during the initiation phase indicated: a period of absolute dependence (<10s); a transitional period in which the dependence on TF is partial and decreases as the reaction proceeds (10–240 s); and a period in which the progress of the reaction is TF independent (>240s). Resupply of reactions late during the termination phase with fresh reactants, but no TF, yielded immediate bursts of thrombin formation similar in magnitude to the original propagation phases. Our data show that independence from the initial TF stimulus is achieved by the onset of the propagation phase and that the ensemble of coagulation products and intermediates that yield this TF independence maintain their prothrombin activating potential for considerable time. These observations support the hypothesis that the transient, localized expression of TF is sufficient to sustain a TF-independent procoagulant response as long as flow persists. [ABSTRACT FROM AUTHOR]

Introduction The development of methods that generate individualized assessments of the procoagulant potential of burn patients could improve their treatment. Beyond its role as an essential intermediate in the formation of thrombin, factor (F)Xa has systemic effects as an agonist to inflammatory processes. In this study, we use a computational model to study the FXa dynamics underlying tissue factor-initiated thrombin generation in a small cohort of burn patients. Materials and Methods Plasma samples were collected upon admission (Hour 0) from nine subjects (five non-survivors) with major burn injuries and then at 48 hours. Coagulation factor concentrations (II, V, VII, VIII, IX, X, TFPI, antithrombin (AT), protein C (PC)) were measured and used in a computational model to generate time course profiles for thrombin (IIa), FXa, extrinsic tenase , intrinsic tenase and prothrombinase complexes upon a 5 pM tissue factor stimulus in the presence of 1 nM thrombomodulin. Parameters were extracted from the thrombin and FXa profiles (including max rate (MaxRIIa and MaxRFXa) and peak level (MaxLIIa and MaxLFXa)). Procoagulant potential was also evaluated by determining the concentration of the complexes at select times. Parameter values were compared between survivors and non-survivors in the burn cohort and between the burn cohort and a simulation based on the mean physiological (100%) concentration for all factor levels. Results Burn patients differed at Hour 0 (p < 0.05) from 100% mean physiological levels for all coagulation factor levels except FV and FVII. The concentration of FX, FII, TFPI, AT and PC was lower; FIX and FVIII were increased. The composition differences resulted in all nine burn patients at Hour 0 displaying a procoagulant phenotype relative to 100% mean physiological simulation (MaxLIIa (306 ± 90 nM vs. 52 nM), MaxRIIa (2.9 ± 1.1 nM/s vs. 0.3 nM/s), respectively p < 0.001); MaxRFXa and MaxLFXa were also an order of magnitude greater than 100% mean physiological simulation (p < 0.001). When grouped by survival status and compared at the time of admission, non-survivors had lower PC levels (56 ± 18% vs. 82 ± 9%, p < 0.05), and faster MaxRFXa (29 ± 6 pM/s vs. 18 ± 6 pM/s, p < 0.05) than those that survived; similar trends were observed for all other procoagulant parameters. At 48 hours when comparing non-survivors to survivors, TFPI levels were higher (108 ± 18% vs. 59 ± 18%, p < 0.05), and MaxRIIa (1.5 ± 1.4 nM/s vs. 3.6 ± 0.7 nM/s, p < 0.05) and MaxRFXa (13 ± 12 pM/s vs. 35 ± 4 pM/s, p < 0.05) were lower; similar trends were observed with all other procoagulant parameters. Overall, between admission and 48 hours, procoagulant potential, as represented by MaxR and MaxL parameters for thrombin and FXa, in non-survivors decreased while in survivors they increased (p < 0.05). In patients that survived, there was a positive correlation between FX levels and MaxLFXa (r = 0.96) and reversed in mortality (r = −0.91). Conclusions Thrombin and FXa generation are increased in burn patients at admission compared to mean physiological simulations. Over the first 48 hours, burn survivors became more procoagulant while non-survivors became less procoagulant. Differences between survivors and non-survivors appear to be present in the underlying dynamics that contribute to FXa dynamics. Understanding how the individual specific balance of procoagulant and anticoagulant proteins contributes to thrombin and FXa generation could ultimately guide therapy and potentially reduce burn injury-related morbidity and mortality. [ABSTRACT FROM AUTHOR]

Introduction: The development of methods that generate individualized assessments of the procoagulant potential of burn patients could improve their treatment. Beyond its role as an essential intermediate in the formation of thrombin, factor (F)Xa has systemic effects as an agonist to inflammatory processes. In this study, we use a computational model to study the FXa dynamics underlying tissue factor-initiated thrombin generation in a small cohort of burn patients.Materials and Methods: Plasma samples were collected upon admission (Hour 0) from nine subjects (five non-survivors) with major burn injuries and then at 48 hours. Coagulation factor concentrations (II, V, VII, VIII, IX, X, TFPI, antithrombin (AT), protein C (PC)) were measured and used in a computational model to generate time course profiles for thrombin (IIa), FXa, extrinsic tenase, intrinsic tenase and prothrombinase complexes upon a 5 pM tissue factor stimulus in the presence of 1 nM thrombomodulin. Parameters were extracted from the thrombin and FXa profiles (including max rate (MaxRIIa and MaxRFXa) and peak level (MaxLIIa and MaxLFXa)). Procoagulant potential was also evaluated by determining the concentration of the complexes at select times. Parameter values were compared between survivors and non-survivors in the burn cohort and between the burn cohort and a simulation based on the mean physiological (100%) concentration for all factor levels.Results: Burn patients differed at Hour 0 (p < 0.05) from 100% mean physiological levels for all coagulation factor levels except FV and FVII. The concentration of FX, FII, TFPI, AT and PC was lower; FIX and FVIII were increased. The composition differences resulted in all nine burn patients at Hour 0 displaying a procoagulant phenotype relative to 100% mean physiological simulation (MaxLIIa (306 ± 90 nM vs. 52 nM), MaxRIIa (2.9 ± 1.1 nM/s vs. 0.3 nM/s), respectively p < 0.001); MaxRFXa and MaxLFXa were also an order of magnitude greater than 100% mean physiological simulation (p < 0.001). When grouped by survival status and compared at the time of admission, non-survivors had lower PC levels (56 ± 18% vs. 82 ± 9%, p < 0.05), and faster MaxRFXa (29 ± 6 pM/s vs. 18 ± 6 pM/s, p < 0.05) than those that survived; similar trends were observed for all other procoagulant parameters. At 48 hours when comparing non-survivors to survivors, TFPI levels were higher (108 ± 18% vs. 59 ± 18%, p < 0.05), and MaxRIIa (1.5 ± 1.4 nM/s vs. 3.6 ± 0.7 nM/s, p < 0.05) and MaxRFXa (13 ± 12 pM/s vs. 35 ± 4 pM/s, p < 0.05) were lower; similar trends were observed with all other procoagulant parameters. Overall, between admission and 48 hours, procoagulant potential, as represented by MaxR and MaxL parameters for thrombin and FXa, in non-survivors decreased while in survivors they increased (p < 0.05). In patients that survived, there was a positive correlation between FX levels and MaxLFXa (r = 0.96) and reversed in mortality (r= -0.91).Conclusions: Thrombin and FXa generation are increased in burn patients at admission compared to mean physiological simulations. Over the first 48 hours, burn survivors became more procoagulant while non-survivors became less procoagulant. Differences between survivors and non-survivors appear to be present in the underlying dynamics that contribute to FXa dynamics. Understanding how the individual specific balance of procoagulant and anticoagulant proteins contributes to thrombin and FXa generation could ultimately guide therapy and potentially reduce burn injury-related morbidity and mortality. [ABSTRACT FROM AUTHOR]

Background: Beyond localized damage to the circulatory system and surrounding tissue, trauma stresses endothelial cells throughout the vasculature, potentially leading to hemorrhagic or thrombotic complications away from the injury site.Objective: Use a whole blood endothelial cell model to define the effects of crystalloid fluid therapy on protein C pathway regulation of tissue factor-initiated coagulation.Methods: Tissue factor-initiated coagulation was studied in the presence of EA.hy926 cells. Blood was diluted to 70% or 40% using normal saline or lactated ringers. Analyses of coagulation dynamics included clot times, thrombin formation (thrombin-antithrombin complex), FV activation/inactivation, fibrinogen consumption, FXIII activation, and platelet activation.Results: In all donors, the onset of thrombin generation was not altered in 70% blood using either diluent; with the blood component reduced to 40%, clot time was prolonged two-fold when normal saline was utilized but was unchanged with lactated ringers. The timing of the activations of FV, fibrinogen, and platelets paralleled the effects of dilution on clot times. Extensive inactivation of FVa was observed in undiluted blood and where lactated ringers was the diluent but not in trials with 40% blood/60% normal saline.Conclusion: Feedback inhibition of tissue factor-initiated coagulation by the protein C pathway is not compromised by hemodilution with crystalloids. [ABSTRACT FROM AUTHOR]

Background: A functional coagulation assay was used to investigate the extrinsic pathway of coagulation on citrated whole blood samples from healthy adult male Sprague Dawley rats using the mini cup and pin system. Methods: Reference values for coagulation parameters from forty-three animals were calculated using data obtained from the ROTEM® delta hemostasis analyzer with the EXTEM test. Results: The following ranges, presented as the 2.5-97.5 percentiles, were established: CT [18-77], CFT [20-80], α [73-86], MCF [53-70], and ML [1-22], along with others. Conclusions: These reference ranges can be used in future studies in rats to identify clinically significant coagulopathies. [ABSTRACT FROM AUTHOR]

The standard clinical coagulation assays, activated partial thromboplastin time (aPTT) and prothrombin time (PT) cannot predict thrombotic or bleeding risk. Since thrombin generation is central to haemorrhage control and when unregulated, is the likely cause of thrombosis, thrombin generation assays (TGA) have gained acceptance as "global assays" of haemostasis. These assays generate an enormous amount of data including four key thrombin parameters (lag time, maximum rate, peak and total thrombin) that may change to varying degrees over time in longitudinal studies. Currently, each thrombin parameter is averaged and presented individually in a table, bar graph or box plot; no method exists to visualize comprehensive thrombin generation data over time. To address this need, we have created a method that visualizes all four thrombin parameters simultaneously and can be animated to evaluate how thrombin generation changes over time. This method uses all thrombin parameters to intrinsically rank individuals based on their haemostatic status. The thrombin generation parameters can be derived empirically using TGA or simulated using computational models (CM). To establish the utility and diverse applicability of our method we demonstrate how warfarin therapy (CM), factor VIII prophylaxis for haemophilia A (CM), and pregnancy (TGA) affects thrombin generation over time. The method is especially suited to evaluate an individual's thrombotic and bleeding risk during "normal" processes (e.g pregnancy or aging) or during therapeutic challenges to the haemostatic system. Ultimately, our method is designed to visualize individualized patient profiles which are becoming evermore important as personalized medicine strategies become routine clinical practice. [ABSTRACT FROM AUTHOR]

In terms of its soluble precursors, the coagulation proteome varies quantitatively among apparently healthy individuals. The significance of this variability remains obscure, in part because it is the backdrop against which the hemostatic consequences of more dramatic composition differences are studied. In this study we have defined the consequences of normal range variation of components of the coagulation proteome by using a mechanism-based computational approach that translates coagulation factor concentration data into a representation of an individual's thrombin generation potential. A novel graphical method is used to integrate standard measures that characterize thrombin generation in both empirical and computational models (e.g max rate, max level, total thrombin, time to 2 nM thrombin ("clot time")) to visualize how normal range variation in coagulation factors results in unique thrombin generation phenotypes. Unique ensembles of the 8 coagulation factors encompassing the limits of normal range variation were used as initial conditions for the computational modeling, each ensemble representing "an individual" in a theoretical healthy population. These "individuals" with unremarkable proteome composition was then compared to actual normal and "abnormal" individuals, i.e. factor ensembles measured in apparently healthy individuals, actual coagulopathic individuals or artificially constructed factor ensembles representing individuals with specific factor deficiencies. A sensitivity analysis was performed to rank either individual factors or all possible pairs of factors in terms of their contribution to the overall distribution of thrombin generation phenotypes. Key findings of these analyses include: normal range variation of coagulation factors yields thrombin generation phenotypes indistinguishable from individuals with some, but not all, coagulopathies examined; coordinate variation of certain pairs of factors within their normal ranges disproportionately results in extreme thrombin generation phenotypes, implying that measurement of a smaller set of factors may be sufficient to identify individuals with aberrant thrombin generation potential despite normal coagulation proteome composition. [ABSTRACT FROM AUTHOR]

Therapeutic agents that regulate blood coagulation are critical to the management of thrombotic disorders and surgical intervention. Many new compounds that were plausible in terms of target, relative specificity, mechanism and kinetic and thermodynamic properties have failed to perform as expected when assessed, at significant expense, in vivo. Our current deterministic mathematical model (Hockin et al. J. Biol. Chem. 2002, 277: 18322) describes the dynamic biochemical repair process that emerges in response to vascular injury and allows us to incorporate hypothetical inhibitors/enhancers at their proposed sites of interaction, thus providing an assessment of their efficacy. In conjunction with our coagulation proteome model (a synthetic reconstruction using purified protein of the components described in the computational model), we evaluated a set of anticoagulants currently in clinical use (unfractionated heparin, synthetic low molecular weight heparins, factor Xa inhibitors and thrombin inhibitors). The interactions of each anticoagulant with its target(s) were mathematically incorporated into the overall mathematical description of the reaction pathways. Outcomes for each anticoagulant at clinically relevant concentrations were generated and validated by comparisons with the synthetic coagulation proteome. Our results show that this type of mathematical methodology can lead to insights into drug performance and to rationalizing the differing properties that an ideal anticoagulant would require in therapeutic vs. prophylactic settings. [ABSTRACT FROM AUTHOR]

The view that clot time-based assays do not provide a sufficient assessment of an individual's hemostatic competence, especially in the context of anticoagulant therapy, has provoked a search for new metrics, with significant focus directed at techniques that define the propagation phase of thrombin generation. Here we use our deterministic mathematical model of tissue-factor initiated thrombin generation in combination with reconstructions using purified protein components to characterize how the interplay between anticoagulant mechanisms and variable composition of the coagulation proteome result in differential regulation of the propagation phase of thrombin generation. Thrombin parameters were extracted from computationally derived thrombin generation profiles generated using coagulation proteome factor data from warfarintreated individuals (N = 54) and matching groups of control individuals (N = 37). A computational clot time prolongation value (cINR) was devised that correlated with their actual International Normalized Ratio (INR) values, with differences between individual INR and cINR values shown to derive from the insensitivity of the INR to tissue factor pathway inhibitor (TFPI). The analysis suggests that normal range variation in TFPI levels could be an important contributor to the failure of the INR to adequately reflect the anticoagulated state in some individuals. Warfarin-induced changes in thrombin propagation phase parameters were then compared to those induced by unfractionated heparin, fondaparinux, rivaroxaban, and a reversible thrombin inhibitor. Anticoagulants were assessed at concentrations yielding equivalent cINR values, with each anticoagulant evaluated using 32 unique coagulation proteome compositions. The analyses showed that no anticoagulant recapitulated all features of warfarin propagation phase dynamics; differences in propagation phase effects suggest that anticoagulants that selectively target fXa or thrombin may provoke fewer bleeding episodes. More generally, the study shows that computational modeling of the response of core elements of the coagulation proteome to a physiologically relevant tissue factor stimulus may improve the monitoring of a broad range of anticoagulants. [ABSTRACT FROM AUTHOR]

Abstract: Introduction: More than 80% of cerebrovascular events are ischemic and largely thromboembolic by nature. We evaluated whether plasma factor composition and thrombin generation dynamics might be a contributor to the thrombotic phenotype of ischemic cerebrovascular events. Materials and Methods: We studied (1) 100 patients with acute ischemic stroke (n=50) or transient ischemic attack (TIA) (n=50) within the first 24hours from symptom onset, and (2) 100 individuals 1 to 4years following ischemic stroke (n=50) or TIA (n=50). The tissue factor pathway to thrombin generation was simulated with a mathematical model using plasma levels of clotting factors (F)II, V, VII, VIII, IX, X, antithrombin and free tissue factor pathway inhibitor (TFPI). Results: The plasma levels of free TFPI, FII, FVIII, and FX were higher, while antithrombin was lower, in the acute patients compared to the previous event group (all p≤0.02). Thrombin generation during acute events was enhanced, with an 11% faster maximum rate, a 15% higher maximum level and a 26% larger total production (all p<0.01). The increased thrombin generation in acute patients was determined by higher FII and lower antithrombin, while increased free TFPI mediated this effect. When the groups are classified by etiology, all stroke sub-types except cardioembolic have increased TFPI and decreased AT and total thrombin produced. Conclusion: Augmented thrombin generation in acute stroke/TIA is to some extent determined by altered plasma levels of coagulation factors. [ABSTRACT FROM AUTHOR]

Deterministic mathematical models of biochemical processes operate as if the empirically derived rate constants governing the dynamics are known with certainty. Our objective in this study was to explore the sensitivity of a deterministic model of blood coagulation to variations in the values of its 44 rate constants. This was accomplished for each rate constant at a given time by defining a normalized ensemble standard deviation (wkif(t)) that accounted for the sensitivity of the predicted concentration of each protein species to variation in that rate constant (from 10 to 1000% of the accepted value). A mean coefficient of variation derived from (wkif(t)) values for all protein species was defined to quantify the overall variation introduced into the model's predictive capacity at that time by the assumed uncertainty in that rate constant. A time-average value of the coefficient of variation over the 20-min simulation for each rate constant was then used to rank rate constants. The model's predictive capacity is particularly sensitive (50% of the aggregate variation) to uncertainty in five rate constants involved in the regulation of the formation and function of the factor VIIa–tissue factor complex. Therefore, our analysis has identified specific rate constants to which the predictive capability of this model is most sensitive and thus where improvements in measurement accuracy will yield the greatest increase in predictive capability. [ABSTRACT FROM PUBLISHER]

Background Coagulation activity among persons with HIV is associated with end-organ disease risk, but the pathogenesis is not well characterized. We tested a hypothesis that hypercoagulation contributes to disease risk, in part, via upregulation of inflammation. Methods Treatment effects of edoxaban (30 mg), a direct factor Xa inhibitor, vs placebo were investigated in a randomized, double-blind crossover trial among participants with HIV and viral suppression and D-dimer levels ≥100 ng/mL. During each 4-month crossover period, blood measures of coagulation, inflammation, and immune activation were assessed. Analyses of change on edoxaban vs change on placebo used linear mixed models. Results Forty-four participants were randomized, and 40 completed at least 1 visit during each study period. The mean age was 49 years, and the CD4+ count was 739 cells/mm3. Edoxaban treatment led to declines in D-dimer (44%) and thrombin-antithrombin complex (26%) but did not lower inflammatory or immune activation measures. More bruising or bleeding events occurred during edoxaban (n = 28) than during placebo or no drug periods (n = 15). Conclusions The direct factor Xa inhibitor edoxaban led to a substantial reduction in coagulation but no effect on inflammation or immune activation. These results do not support that hypercoagulation contributes to ongoing inflammation during chronic antiretroviral therapy–treated HIV disease. [ABSTRACT FROM AUTHOR]