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OBJECTIVES Extracorporeal membrane oxygenation provides large surface exposure to human blood leading to coagulation activation. Only limited clinical data are available on contact activation and coagulation factor XII activity in extracorporeal membrane oxygenation patients. DESIGN Prospective cohort study. SETTING Three medical ICUs at the Medical University of Vienna. PATIENTS Adult patients receiving venovenous or venoarterial extracorporeal membrane oxygenation. INTERVENTIONS None. MEASUREMENTS AND MAIN RESULTS The primary outcome was the change in coagulation factor XII activity in response to extracorporeal membrane oxygenation. Secondary outcomes included the prevalence of reduced coagulation factor XII activity (< 60%) among patients receiving extracorporeal membrane oxygenation and association of coagulation factor XII activity with thromboembolic and bleeding complications. An exploratory endpoint was the association of coagulation factor XII activity and activated partial thromboplastin time in heparinase-treated samples in vitro. Fifty-one patients with a total of 117 samples were included in the study between July 2018 and February 2020. Fifty patients (98%) had reduced coagulation factor XII activity at any timepoint during extracorporeal membrane oxygenation. Median coagulation factor XII activity during extracorporeal membrane oxygenation treatment was 30% (interquartile range, 21.5-41%) and increased after discontinuation (p = 0.047). Patients with thromboembolic complications had higher median coagulation factor XII activity during extracorporeal membrane oxygenation (34% vs 23%; p = 0.023). The odds of a thromboembolic event increased by 200% per tertile of median coagulation factor XII activity (crude odds ratio, 3.034; 95% CI, 1.21-7.63). No association with bleeding was observed. In heparinase-treated samples, coagulation factor XII activity correlated well with activated partial thromboplastin time (r = -0.789; p = 0.007). CONCLUSIONS We observed a high prevalence of reduced coagulation factor XII activity in adult patients on extracorporeal membrane oxygenation, which may confound activated partial thromboplastin time measurements and limit its clinical usefulness for monitoring and titrating anticoagulation with unfractionated heparin. Lower coagulation factor XII activity was associated with less thromboembolic complications, which may highlight the potential of coagulation factor XII to serve as a target for anticoagulation in extracorporeal membrane oxygenation.

To review the evidence that the Alzheimer peptide β-amyloid interacts with the blood coagulation system and influences the pathophysiology of the disease.β-amyloid can interact with fibrinogen and blood coagulation factor XII and trigger ischemia and inflammation.β-amyloid interacts with fibrinogen and factor XII. These interactions can lead to increased clotting, abnormal clot formation, persistent fibrin deposition, and generation of proinflammatory molecules. These events can damage neurons and could contribute to the cognitive decline in Alzheimer's disease patients.

Background: Terminal complications of bacterial sepsis include development of disseminated intravascular consumptive coagulopathy. Bacterial constituents, including long-chain polyphosphates (polyP), have been shown to activate the contact pathway of coagulation in plasma. Recent work shows that activation of the contact pathway in flowing whole blood can promote thrombin generation and platelet activation and consumption distal to thrombus formation ex vivo and in vivo . Aim: Determine whether presence of long-chain polyP in the bloodstream promotes platelet activation and consumption in a coagulation factor (F)XII-dependent manner. Methods/Results: The addition of long-chain polyP to human whole blood promoted platelet P-selectin expression, microaggregate formation and platelet consumption in the bloodstream under shear in a FXII-dependent manner. Moreover, long-chain polyP accelerated thrombus formation on immobilized collagen surfaces under shear flow in a thrombin generation-dependent manner. Distal to the site of thrombus formation, platelet consumption was dramatically enhanced in the presence of long-chain polyP in the bloodstream. Inhibiting contact activation of coagulation using established and novel agents reduced fibrin formation on collagen as well as platelet consumption in the bloodstream distal to the site of thrombus formation. In vivo , FXII deficiency was protective against long-chain polyP occlusive lung thrombus formation in mice. Lastly, in a non-human primate model of sepsis, pretreatment of animals with an antibody blocking FXI activation by FXIIa (14E11) diminished LD 100 S. aureus -induced platelet and fibrinogen consumption. Conclusions: This study demonstrates that bacterial-type long-chain polyP promotes FXII-mediated thrombin generation and platelet activation in the flowing blood and could contribute to sepsis-associated thrombotic processes, consumptive coagulopathy and thrombocytopenia.

Background: Terminal complications of bacterial sepsis include development of disseminated intravascular consumptive coagulopathy. Bacterial constituents, including long-chain polyphosphates (polyP), have been shown to activate the contact pathway of coagulation in plasma. Recent work shows that activation of the contact pathway in flowing whole blood can promote thrombin generation and platelet activation and consumption distal to thrombus formation ex vivo and in vivo . Aim: Determine whether presence of long-chain polyP in the bloodstream promotes platelet activation and consumption in a coagulation factor (F)XII-dependent manner. Methods/Results: The addition of long-chain polyP to human whole blood promoted platelet P-selectin expression, microaggregate formation and platelet consumption in the bloodstream under shear in a FXII-dependent manner. Moreover, long-chain polyP accelerated thrombus formation on immobilized collagen surfaces under shear flow in a thrombin generation-dependent manner. Distal to the site of thrombus formation, platelet consumption was dramatically enhanced in the presence of long-chain polyP in the bloodstream. Inhibiting contact activation of coagulation using established and novel agents reduced fibrin formation on collagen as well as platelet consumption in the bloodstream distal to the site of thrombus formation. In vivo , FXII deficiency was protective against long-chain polyP occlusive lung thrombus formation in mice. Lastly, in a non-human primate model of sepsis, pretreatment of animals with an antibody blocking FXI activation by FXIIa (14E11) diminished LD 100 S. aureus -induced platelet and fibrinogen consumption. Conclusions: This study demonstrates that bacterial-type long-chain polyP promotes FXII-mediated thrombin generation and platelet activation in the flowing blood and could contribute to sepsis-associated thrombotic processes, consumptive coagulopathy and thrombocytopenia.

Background: Terminal complications of bacterial sepsis include development of consumptive coagulopathy referred to as disseminated intravascular coagulation. Bacterial constituents, including long-chain polyphosphates (polyP), have been shown to activate the contact pathway of coagulation in plasma. Recent work shows that activation of the contact pathway is capable of promoting thrombin generation and platelet activation and consumption in whole blood distal to thrombus formation under shear ex vivo and in vivo . Aim: Test the hypothesis that the presence of long-chain polyP in the bloodstream promotes platelet activation and consumption in a coagulation factor (F)XII-dependent manner. Methods and Results: Presence of long-chain polyP in whole blood promoted platelet aggregation on immobilized collagen surfaces under shear flow. Long-chain polyP enhanced fibrin formation and shortened clotting times of plasma and whole blood. The addition of long-chain polyP promoted platelet P-selectin expression, microaggregate formation and platelet consumption in the bloodstream under shear in a FXII-dependent manner. Moreover, long-chain polyP accelerated thrombus formation on immobilized collagen surfaces under shear flow. Distal to the sites of thrombus formation, platelet consumption was dramatically enhanced in the presence of long-chain polyP in the bloodstream. Inhibiting contact activation of the coagulation pathway reduced fibrin formation on collagen as well as platelet consumption in the bloodstream distal to the site of thrombus formation. Conclusions: This study demonstrates that bacterial-type long-chain polyP promotes FXII-mediated thrombin generation and platelet activation in the flowing blood and could exaggerate sepsis-associated thrombotic processes, consumptive coagulopathy and thrombocytopenia.

Objective— Activation of neutrophils by microbial or inflammatory stimuli results in the release of neutrophil extracellular traps (NETs) that are composed of DNA, histones, and antimicrobial proteins. In purified systems, cell-free DNA (CFDNA) activates the intrinsic pathway of coagulation, whereas histones promote thrombin generation through platelet-dependent mechanisms. However, the overall procoagulant effects of CFDNA/histone complexes as part of intact NETs are unknown. In this study, we examined the procoagulant potential of intact NETs released from activated neutrophils. We also determined the relative contribution of CFDNA and histones to thrombin generation in plasmas from patients with sepsis. Approach and Results— NETs released from phorbyl myristate–activated neutrophils enhance thrombin generation in platelet-poor plasma. This effect was DNA dependent (confirmed by DNase treatment) and occurred via the intrinsic pathway of coagulation (confirmed with coagulation factor XII– and coagulation factor XI–depleted plasma). In platelet-rich plasma treated with corn trypsin inhibitor, addition of phorbyl myristate–activated neutrophils increased thrombin generation and shortened the lag time in a toll-like receptor-2– and toll-like receptor-4–dependent mechanism. Addition of DNase further augmented thrombin generation, suggesting that dismantling of the NET scaffold increases histone-mediated, platelet-dependent thrombin generation. In platelet-poor plasma samples from patients with sepsis, we found a positive correlation between endogenous CFDNA and thrombin generation, and addition of DNase attenuated thrombin generation. Conclusions— These studies examine the procoagulant activities of CFDNA and histones in the context of NETs. Our studies also implicate a role for the intrinsic pathway of coagulation in sepsis pathogenesis.

Background— Blood coagulation is a tightly regulated process of sequentially activated serine proteases culminating in fibrin formation, which is critical for limiting posttraumatic blood loss but also may contribute to acute thrombotic diseases, most notably myocardial infarction and stroke. Recent studies with factor XII–deficient mice revealed that the factor XII–induced intrinsic coagulation pathway is essential for pathological thrombus formation but dispensable for hemostasis. Consequently, these findings led to the hypothesis that factor XII could be a promising pharmacological target for safe antithrombotic therapy. Methods and Results— The complementary DNA of the previously described factor XIIa inhibitor Infestin-4, cloned from the midgut of Triatoma infestans , was fused to recombinant human albumin (rHA) and analyzed in vitro. The resulting protein rHA-Infestin-4 specifically inhibits factor XIIa and causes prolonged activated partial thromboplastin time in human, mouse, and rat plasma. To assess its inhibitory potency in vivo, mice and rats were injected with rHA-Infestin-4 and challenged in pathological thrombus formation models. In addition, bleeding assays were performed. rHA-Infestin-4 completely abolished occlusive arterial thrombus formation in mice and rats while leaving hemostasis fully intact. Furthermore, rHA-Infestin-4 was highly protective in a murine model of ischemic stroke. Conclusion— These results identify rHA-Infestin-4 as a promising agent to achieve powerful protection from ischemic cardiovascular and cerebrovascular events without affecting hemostasis.

In a subgroup of hereditary angioedema (HAE) patients with normal C1-esterase inhibitor levels, HAE is caused by a Thr309Lys mutation in the coagulation factor XII (F12) gene. The aim of this study was to examine elements of the kallikrein-kinin system ('contact system') and the downstream-linked coagulation, complement and fibrinolytic systems in the plasma of six patients with HAE caused by the Thr309Lys mutation and healthy probands. Blood samples were taken from participants during the symptom-free interval between attacks. Samples were analyzed for activity and concentrations of components of the kallikrein-kinin system and linked enzyme systems. The mean FXII clotting activity was 90% in patients with the FXII mutation, and the concentration of FXIIa was 4.1 ng/ml; this did not differ from healthy probands. Mean prekallikrein amidolytic activity and high-molecular-weight kininogen clotting activity were 130 and 144%, respectively, both higher than in healthy probands. The mean kallikrein-like activity of the HAE patients was 11.4 U/l and did not differ from healthy probands. There was no difference in FXII surface activation by silicon dioxide or in kallikrein-like activity with and without activation by dextran sulfate. Contrary to the results of a recently published study, no indication that the Thr309Lys mutation causes a 'gain-of-function' of FXIIa was observed in this investigation.

BACKGROUND 'New pressor protein' was observed after tryptic activation of human and rat plasma in vitro, which is done conventionally for prorenin measurements. RESULTS It is potently pressor, heat labile, possesses enzyme activity, and has a relative molecular mass > 30 kDa with isoelectric point(s) 4.7-4.9. New pressor protein equivalent to only 0.01 ml human, or rat, plasma injected intravenously quickly raises systolic blood pressure in 300 g anesthetized, ganglion-blocked, bioassay rats by about 15 mmHg. For unknown reasons, this is potentiated to about 45 mmHg after treatment with angiotensin I converting enzyme inhibitors (such as captopril and enalapril). New pressor protein activity in rats remains normal 24 h after bilateral nephrectomy, suggesting that it has an extrarenal origin and, furthermore, excluding the possibility of an association with renin-angiotensin system. Systolic blood pressure elevation is greater than the diastolic one, implicating cardiotonic effects. Human plasma new pressor protein was purified using standard biochemical techniques and its N-terminal sequence (19 residues) found to be homologous with the beta factor XIIa fragment of coagulation factor XII. This was supported by demonstrating inhibition of new pressor protein activity in vitro using the factor XII-specific corn trypsin inhibitor. Also, human new pressor protein activity in humans congenitally deficient in coagulation factor XII is very low. The high potency and multiphasic, cardiotonic effects of injected new pressor protein suggest that it interacts synergistically with other systems in the body. This was confirmed by showing that, within 10 min of total bilateral adrenalectomy, responses to new pressor protein decreased markedly. CONCLUSIONS New pressor protein's action requires adrenal (medullary?) involvement, but its mechanism of action and that of its potentiation by angiotensin converting enzyme inhibitors remain unknown. The physiologic and clinical relevance of these observations depends on whether activation of new pressor protein can occur in vivo.

Background: Currently, a majority of evidence suggests that the tissue factor-mediated extrinsic coagulation pathway is most important in generating thrombin in vivo , although accumulating evidence implicates the coagulation factor XII/factor XI-mediated intrinsic system as well. The endogenous thrombin potential (ETP), a measure of thrombin generation, is gaining importance in assessing risk of hemorrhage and thrombosis, but genetic determinants of ETP have not been investigated in population-based studies. Methods: Total ETP (ETP T ) was measured in citrated plasma by TechnoClone TGA assay in a random subset of Cardiovascular Health Study (CHS) participants ( n =1,000). Extrinsic pathway ETP (ETP EX ) was measured by the addition of anti-FXIa antibody to the assay; Intrinsic pathway ETP (ETP IN ) was calculated by subtracting ETP EX from ETP T . Genotyping was performed in the NHLBI’s Candidate-gene Association Resource (CARe) Study on 49,320 SNPs using Illumina’s custom IBCv2 genotyping array in 866 participants. Associations of SNPs with age- and sex-adjusted ETP phenotypes were evaluated by linear regression using PLINK; results were considered statistically significant if p≤ 2x 10 -6 . Results: Two SNPs on the IBC array were significantly associated with lower ETP T : rs2545801 (β=-45.87 nM ± 7.3 nM; p=5.7x10 -10 ; minor allele frequency (MAF) =0.24), and rs1801020 (β=-45.76 nM ± 7.3 nM; p=7.4x10 -10 ; MAF= 0.23). Both SNPs are located in the coagulation factor XII gene ( F12 ); rs2545801is located in a 5’ region upstream of the transcriptional start site, and rs1801020 in a 5’ untranslated region; the SNPs are in strong linkage disequilibrium (LD) (r 2 = 0.92). Each SNP had a significant effect on ETP thrombin peak height. The mean (SD) thrombin peak heights were 408 nM (152nM) vs.523 nM (123 nM) and 404 nM (147 nM) vs. 523 nM (123 nM) for those homozygous for the minor and major alleles of rs2545801 and rs1801020, respectively. Addition of corn trypsin inhibitor (CTI; an inhibitor of activated FXII (FXIIa)) had no effect on ETP, suggesting that the F12 SNPs may affect the FXII-dependent generation of activated FXI (FXIa) in vivo. Therefore, we inhibited FXIa by the addition of anti-FXIa antibody, which resulted in a large and variable effect on ETP thrombin peak height. Consistent with these observations, analyses of SNPs associated with ETP IN revealed increased significance of the p-values for the F12 SNPs (p=1.40 x 10 -15 and p=1.60 x 10 -14 for rs1801020 and rs254580, respectively). Conclusion: These data support the importance of F12 SNPs on thrombin generation, and suggest that these SNPs affect the amount of FXIa generated in vivo . Additional work is needed to confirm this hypothesis, and to examine relationships of these biomarkers and SNPs with thrombotic diseases.

Three patients with liver disease and prolonged activated partial thromboplastine time (APTT) on routine tests are presented. One woman had metastatic liver disease from gastric carcinoma, a second woman had autoimmune hepatitis, and one man had severe chronic hepatitis B. APTT was not corrected after mixing experiments with 25%, 50%, and 75% of normal pool plasma, indicating the presence of an acquired inhibitor. In all three cases, factor XII coagulant activity was reduced