Aminotransferases disorders associated with venous thromboembolic events in patients infected with COVID-19.

INTRODUCTION AND OBJECTIVES: Since the outbreak of the COVID-19 pandemic, increasing evidence suggests that infected patients present a high incidence of venous thromboembolic (VTE) events and elevated aminotransferases (AT).The objective of this work was to evaluate the incidence of aminotransferases disorders in patients infected with COVID-19 and to manage the VTE events associated with elevated AT. PATIENTS OR MATERIALS AND METHODS: We report a retrospective study of 46 patients admitted for COVID-19 infection. Venous duplex ultrasound of lower limbs was performed in all patients at Day 0 and Day 5. All patients had antithrombotic-prophylaxis upon admission using low molecular weight heparin with Enoxaparin. Demographics, comorbidities and laboratory parameters were collected and analyzed. RESULTS: Elevated AT were reported in 28 patients (61%). 10 had acute VTE events of which eight (17.4%) had aminotransferases disorders. They had been treated with curative Enoxaparin. After a follow-up of 15 and/or 30 days, six of them were controlled, and treated with direct oral anticoagulant (DOACs) after normalization of aminotransferases. CONCLUSIONS: The incidence of aminotransferases disorders associated with acute VTE events in patients infected with COVID-19 is significant. The use of DOACs appear pertinent in these patients. Monitoring of the liver balance should therefore be considered at a distance from the acute episode in the perspective of DOACs relay.

Correlations between methylenetetrahydrofolate reductase gene polymorphisms and venous thromboembolism: A meta-analysis of 99 genetic association studies.

We performed this meta-analysis to better assess the relationship between methylenetetrahydrofolate reductase gene ( MTHFR) polymorphisms and the risk of venous thromboembolism. Eligible studies were searched in PubMed, Medline, Embase, and Web of Science. Odds ratios with 95% confidence intervals were used to assess associations of MTHFR polymorphisms with venous thromboembolism. A total of 99 genetic association studies were enrolled for analyses. Although no positive results were detected in overall analyses for the rs1801131 polymorphism. Further subgroup analyses according to ethnicity of participants and type of disease revealed that the rs1801131 polymorphism was significantly correlated with the risk of pulmonary embolism. For the rs1801133 polymorphism, significant association with the risk of venous thromboembolism was found in the dominant, recessive, and allele models. Further subgroup analyses according to ethnicity of participants revealed that the rs1801133 polymorphism was significantly associated with the risk of venous thromboembolism in Caucasians, East Asians, and West Asians. When we stratified available data according to type of disease, we found that the rs1801133 polymorphism was also significantly correlated with the risk of deep vein thrombosis and pulmonary embolism. In conclusion, our findings indicate that the MTHFR rs1801133 polymorphism may serve as a potential biological marker for venous thromboembolism in Caucasians, East Asians, and West Asians. Moreover, the MTHFR rs1801133 polymorphism may be implicated in the development of deep vein thrombosis and pulmonary embolism, while the MTHFR rs1801131 polymorphism may contribute to the development of pulmonary embolism.

Implications of Janus Kinase 2 Mutation in Embolic Stroke of Unknown Source.

The role of genetic mutations in cerebral ischemia is not completely understood. Among these genetic variations, Philadelphia-negative gain-of-function mutation in the janus kinase 2 (JAK2) protein leads to overexpression of the genes involved in cell growth and proliferation, and has been linked to development of hematological malignancies, specifically, myeloproliferative neoplasms (MPNs; essential thrombocythemia [ET], polycythemia vera [PV], and primary myelofibrosis). Overt ET and PV are known to induce a prothrombotic state that leads to development of vascular complications, including cerebral arterial or venous thrombosis. Thromboembolism can precede overt presentation of an MPN by 2-3 years. As such, for the selected cases of embolic stroke or cerebrovascular sinus thrombosis with otherwise undetermined source and persistent thrombocytosis or polycythemia, in the absence of a confirmed MPN diagnosis, screening for JAK2 mutation may be reasonable, as early diagnosis and appropriate treatment can influence outcome by preventing recurrent thrombotic events. In this article, we review the literature on the genetics, pathogenesis, clinical manifestations, and treatment of JAK2-associated thrombosis, and present 2 cases of JAK2-associated cerebral arterial infarction and cerebral and systemic venous thromboembolism with otherwise negative etiology workup for stroke.

Assessment of the procoagulant potential after laparoscopic sleeve gastrectomy: a potential role for extended thromboprophylaxis.

BACKGROUND: Bariatric surgery is associated with increased thromboembolic risk, which may extend well beyond hospital stay. The hemostatic mechanisms implicated in this risk are not well established. OBJECTIVES: We aimed to determine the dynamics of hemostatic changes and procoagulant potential among patients undergoing laparoscopic sleeve gastrectomy, during both the early and late postoperative periods. SETTING: A university hospital. METHODS: Patients who underwent laparoscopic sleeve gastrectomy were recruited consecutively to this study. Blood samples were taken preoperatively, before discharge (postoperative day [POD] 3), and at the first follow-up visit (POD10). All samples were tested for complete blood count, C-reactive protein, von Willebrand factor, factor VIII, fibrinogen, and thrombin generation. RESULTS: The median preoperative body mass index of the 26 participants was 41.3 (38.7-43.3) kg/m2. Compared with preoperative evaluation, fibrinogen, von Willebrand factor antigen and activity, and factor VIII levels were significantly higher at POD3 and POD10 (P<.0001 for all comparisons). Peak thrombin levels and endogenous thrombin potential (ETP) were higher at POD3 (P = .005 for both comparisons) and POD10 (P = .0009 and<.0001) compared with baseline. ETP and peak thrombin, as well as fibrinogen, von Willebrand factor, and factor VIII levels, were comparable between POD3 and POD10. Multivariate analysis showed that the only predictor of postoperative ETP was the preoperative ETP level (ß = .55, P = .007). CONCLUSIONS: As determined by thrombin generation, laparoscopic sleeve gastrectomy was associated with hypercoagulability, which persisted during POD10. This finding suggests a possible benefit of extended thromboprophylaxis. Nevertheless, our results should be interpreted with caution due to the lack of a control group.

MTHFR A1298C and C677T Polymorphisms Are Associated with Increased Risk of Venous Thromboembolism: A Retrospective Chart Review Study.

BACKGROUND: Methylene tetrahydrofolate reductase (MTHFR) is a key enzyme in homocysteine metabolism. This study aims to determine the impact of MTHFR polymorphisms on plasma homocysteine levels and risks of venous thromboembolism (VTE). METHODS: This retrospective chart review study included a total of 188 subjects who were tested for MTHFR polymorphisms at Metrowest Coagulation Laboratory between April 2011 and April 2016. Two independent coders were trained to extract relevant clinical data for statistical analysis. RESULTS: VTE occurred in 50% of patients with compound mutation, compared with only 28.6% of subjects from the wild-type group. Patients with heterozygous or homozygous A1298C or C677T variants had an intermediate risk of VTE. The median homocysteine level in the wild-type group was slightly lower than that of heterozygous or homozygous MTHFR variants. The difference, however, was not significant (p = 0.6193). Moreover, there was no difference in plasma homocysteine level between patients with VTE versus VTE-free (p = 0.4923). CONCLUSIONS: Heterozygous or homozygous MTHFR variants, especially a compound mutation, are associated with increased risk of VTE. Hyperhomocysteinemia does not correlate with MTHFR polymorphisms or VTE risk. Hence, MTHFR genotyping provides more consistent assessment of VTE risk. This information can be incorporated into risk stratification for early intervention and prophylaxis of VTE.

The ENTPD1 promoter polymorphism -860 A > G (rs3814159) is associated with increased gene transcription, protein expression, CD39/NTPDase1 enzymatic activity, and thromboembolism risk.

Ectonucleoside triphosphate diphosphohydrolase 1 (NTPDase1) degrades the purines ATP and ADP that are key regulators of inflammation and clotting. We hypothesized that NTPDase1 polymorphisms exist and that they regulate this pathway. We sequenced the ENTPD1 gene (encoding NTPDase1) in 216 subjects then assessed genotypes in 2 cohorts comprising 2213 humans to identify ENTPD1 polymorphisms associated with venous thromboembolism (VTE). The G allele of the intron 1 polymorphism rs3176891 was more common in VTE vs. controls (odds ratio 1.26-1.9); it did not affect RNA splicing, but it was in strong linkage disequilibrium with the G allele of the promoter polymorphism rs3814159, which increased transcriptional activity by 8-fold. Oligonucleotides containing the G allele of this promoter region bound nuclear extracts more avidly. Carriers of rs3176891 G had endothelial cells with increased NTPDase1 activity and protein expression, and had platelets with enhanced aggregation. Thus, the G allele of rs3176891 marks a haplotype associated with increased clotting and platelet aggregation attributable to a promoter variant associated with increased transcription, expression, and activity of NTPDase1. We term this gain-of-function phenotype observed with rs3814159 G "CD39 Denver."-Maloney, J. P., Branchford, B. R., Brodsky, G. L., Cosmic, M. S., Calabrese, D. W., Aquilante, C. L., Maloney, K. W., Gonzalez, J. R., Zhang, W., Moreau, K. L., Wiggins, K. L., Smith, N. L., Broeckel, U., Di Paola, J. The ENTPD1 promoter polymorphism -860 A > G (rs3814159) is associated with increased gene transcription, protein expression, CD39/NTPDase1 enzymatic activity, and thromboembolism risk.

Elevated CETP Lipid Transfer Activity is Associated with the Risk of Venous Thromboembolism.

AIM: Cholesteryl ester transfer protein (CETP) is an important lipid transfer factor in plasma that enhances prothrombinase activity in purified systems. This study was conducted to test the association of plasma CETP activity with venous thrombosis (VTE) and to address the procoagulant mechanism of CETP activity in prothrombinase assays. METHODS: We measured CETP lipid transfer activity in plasmas of 49 male VTE patients and in plasmas of matched controls. CETP procoagulant activity was tested in purified prothrombinase systems. RESULTS: CETP lipid transfer activity levels were significantly higher in VTE patients than in controls (p=0.0008). A subset of patients carrying the CETP mutations Ala373Pro and Arg451Gln, which were also linked to the VTE risk, showed significantly higher plasma CETP activity than the noncarriers. The plasma CETP activity negatively correlated with APTT, suggesting that the CETP activity is associated with plasma coagulability. Recombinant (r) CETP bound to both factor Xa (Kd=15 nM) and Gla-domainless factor Xa (Kd=59 nM), whereas rCETP enhanced prothrombin activation by factor Xa, but not by Gla-domainless factor Xa. rCETP also required factor Va for enhancement of prothrombinase activity. When we addressed the effects of mutations in CETP on prothrombinase activity, Gln451-rCETP was found to have five-fold higher thrombin generation activity than wt-rCETP or Pro373-rCETP. CONCLUSIONS: Elevated CETP lipid transfer activity in plasma was associated with the risk of VTE. Gln451-CETP, which is linked to VTE, has much higher procoagulant activity than wt-CETP. CETP might act as a physiologic procoagulant by mechanisms that involve its direct binding to factor Xa.

Defective thrombus formation in mice lacking endogenous factor VII activating protease (FSAP).

Factor VII (FVII) activating protease (FSAP) is a circulating protease with a putative function in blood coagulation and fibrinolysis. Genetic epidemiological studies have implied a role for FSAP in carotid stenosis, stroke and thrombosis. To date, no in vivo evidence is available to support these claims. We have, for the first time, used FSAP-/- mice to define its role in thrombosis and haemostasis in vivo and to characterise the molecular mechanisms involved. FeCl3-induced arterial thrombosis in carotid and mesenteric artery revealed that the occlusion time was significantly increased in FSAP-/- mice (p< 0.01) and that some FSAP-/- mice did not occlude at all. FSAP-/- mice were protected from lethal pulmonary thromboembolism induced by collagen/ epinephrine infusion (p< 0.01). Although no spontaneous bleeding was evident, in the tail bleeding assay a re-bleeding pattern was observed in FSAP-/- mice. To explain these observations at a mechanistic level we then determined how haemostasis factors and putative FSAP substrates were altered in FSAP-/- mice. Tissue factor pathway inhibitor (TFPI) levels were higher in FSAP-/- mice compared to WT mice whereas FVIIa levels were unchanged. Other coagulation factors as well as markers of platelet activation and function revealed no significant differences between WT and FSAP-/- mice. This phenotype of FSAP-/- mice could be reversed by application of exogenous FSAP. In conclusion, a lack of endogenous FSAP impaired the formation of stable, occlusive thrombi in mice. The underlying in vivo effect of FSAP is more likely to be related to the modulation of TFPI rather than FVIIa.

Association between MTHFR C677T polymorphism and venous thromboembolism risk in the Chinese population: a meta-analysis of 24 case-controlled studies.

The association between methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms and venous thromboembolism (VTE) risk in the Chinese population has been widely reported, but results were inconsistent and underpowered. To elucidate the variable results, a meta-analysis and systematic review were performed from all case-controlled studies relating MTHFR C677T polymorphism by pooling data on them. We estimated the pooled odds ratio with its 95% confidence intervals to assess this possible association. Finally, a total of 24 studies with 2339 cases and 4048 controls were included in the current meta-analysis. Significant association was found with VTE risk for all genetic models. Subgroup analyses by type of VTE further identified the above-mentioned association in deep vein thrombosis/pulmonary embolism and splanchnic vein thrombosis. The findings from our meta-analysis support the associations of MTHFR C677T polymorphism with VTE risk in the Chinese population.

Dysfunction of protein C anticoagulant system, main genetic risk factor for venous thromboembolism in northeast Asians.

Venous thromboembolism (VTE) is a life threatening medical disorder worldwide. A great deal of evidence suggests that prevalence of VTE varies significantly among ethnic populations, with consistently lower incidence found in Asians. While the distribution of genetic risk factors may vary among races, genetic risk factors can play a major role among individuals with different genetic backgrounds. Two clinically evaluated low-frequency genetic mutations that predispose to VTE--the factor V Leiden mutation and prothrombin G20210A mutation--are found predominantly in Caucasians, and virtually never in Asians. The findings of a recent genetic study of VTE in northeast Asians, which greatly advanced our knowledge in this area, indicate that the most frequent genetic risk factors for VTE in northeast Asians can be attributed to a dysfunction of the protein C anticoagulant system. Several low-frequency genetic mutations, PROS1 p.Lys196Glu in Japanese and PROC p.Arg189Trp and p.Lys193del in Chinese, are significantly associated with increased risk for VTE, with odds ratio more than 2 through the reduced protein C anticoagulant activity. Construction of a multifactorial model based on the genetic risk factors in the protein C anticoagulant system could facilitate genetic counseling for VTE risk in these populations. The influence of prevalent genetic mutations on the risk of VTE should be further investigated in Asian countries.

Lipoprotein-associated phospholipase A2 and venous thromboembolism: a prospective study.

INTRODUCTION: Plasma lipoprotein-associated phospholipase A2 (Lp-PLA2) is an inflammatory marker associated positively with atherothrombotic risk. Whether Lp-PLA2 is related to risk of venous thromboembolism (VTE) is incompletely studied. METHODS: We assessed Lp-PLA2 activity in 10,687 Atherosclerosis Risk in Communities (ARIC) Study participants and followed them a median of 8.3 years (from 1996-98 through 2005) for VTE occurrence (n=226). RESULTS: There was no significant association between baseline Lp-PLA2 quartiles and risk of VTE, neither overall nor stratified as provoked or unprovoked. Adjusted for other risk factors, the hazard ratios (95% confidence interval) of total VTE across quartiles of Lp-PLA2 were 1.0 (reference), 0.95 (0.64, 1.42), 1.03 (0.69, 1.56), and 1.26 (0.83, 1.91). In the subset of participants with LDL-cholesterol ≥130 mg/dL, hazard ratios of total VTE were 1.00, 1.39 (0.44, 4.44), 2.45 (0.84, 7.11), and 2.84 (0.99, 8.14). CONCLUSION: Our study does not support the overall hypothesis that elevated Lp-PLA2 contributes to VTE occurrence in the general population. However, in the presence of high LDL-cholesterol there was some evidence that Lp-PLA2 may increase VTE risk.

Increased risk of venous thromboembolism is associated with genetic variation in heme oxygenase-1 in Blacks.

BACKGROUND: Venous thromboembolism (VTE) affects as many as 1 in 1000 individuals in the United States. Although Blacks are disproportionately affected by VTE, few genetic risk factors have been identified in this population. The inducible heme oxygenase-1 (HMOX1) gene encodes a key cytoprotective enzyme with anti-inflammatory, antioxidant and anticoagulant activity acting in the vascular system. A (GT)(n) microsatellite located in the promoter of the HMOX1 gene influences the level of response. METHODS AND RESULTS: Using the Genetic Attributes and Thrombosis Epidemiology (GATE) study, we examined the association between HMOX1 repeat length and VTE events in 883 Black and 927 White patients and matched controls. We found no association between HMOX1 genotypes and VTE in Whites. However, in Black patients, carrying two long (L) alleles (≥ 34 repeats) was significantly associated with provoked (odds ratio (OR) 1.86, 95% confidence interval (CI): 1.19-2.90) or recurrent (OR 3.13, 95% CI: 1.77-5.53) VTE events. CONCLUSIONS: We have demonstrated for the first time an association between genetic variation in HMOX1, and VTE in Blacks. Our results support a key role for the heme oxygenase system in protecting patients at increased risk for thrombosis and suggest a potential mechanism for targeted screening and intervention.

Identification and characterisation of monoclonal antibodies that impair the activation of human thrombin activatable fibrinolysis inhibitor through different mechanisms.

Thrombin activatable fibrinolysis inhibitor (TAFI) forms a molecular link between coagulation and fibrinolysis and is a putative target to develop profibrinolytic drugs. Out of a panel of monoclonal antibodies (MA) raised against TAFI-ACIIYQ, we selected MA-TCK11A9, MA-TCK22G2 and MA-TCK27A4, which revealed high affinity towards human TAFI-TI-wt. MA-TCK11A9 was able to inhibit mainly plasmin-mediated TAFI activation, MA-TCK22G2 inhibited plasmin- and thrombin-mediated TAFI activation and MA-TCK27A4 inhibited TAFI activation by plasmin, thrombin and thrombin/thrombomodulin (T/TM) in a dose-dependent manner. These MA did not interfere with TAFIa activity. Using an eight-fold molar excess of MA over TAFI, all three MA were able to reduce clot lysis time significantly, i.e. in the presence of exogenous TM, MA-TCK11A9, MA-TCK22G2 and MA-TCK27A4 reduced clot lysis time by 47 ± 9.1%, 80 ± 8.6% and 92 ± 14%, respectively, compared to PTCI. This effect was even more pronounced in the absence of TM i.e. MA-TCK11A9, MA-TCK22G2 and MA-TCK27A4 reduced clot lysis time by 90 ± 14%, 140 ± 12% and 147 ± 29%, respectively, compared to PTCI. Mutagenesis analysis revealed that residues at position 268, 272 and 276 are involved in the binding of MA-TCK11A9, residues 147 and 148 in the binding of MA-TCK22G2 and residue 113 in the binding of MA-TCK27A4. The present study identified three MA, with distinct epitopes, that impair the activation of human TAFI and demonstrated that MA-TCK11A9 which mainly impairs plasmin-mediated TAFI activation can also reduce significantly clot lysis time in vitro.

Absolute risk of venous and arterial thromboembolism in thrombophilic families is not increased by high thrombin-activatable fibrinolysis inhibitor (TAFI) levels.

High levels of thrombin-activatable fibrinolysis inhibitor (TAFI) are a supposed risk factor for thrombosis. However, results from previous studies are conflicting. We assessed the absolute risk of venous and arterial thromboembolism in subjects with high TAFI levels (>126 U/dl) versus subjects with normal levels, and the contribution of other concomitant thrombophilic defects. Relatives from four identical cohort studies in families with either deficiencies of antithrombin, protein C or protein S, prothrombin 20210A, high factor VIII levels, or hyperhomocysteinemia were pooled. Probands were excluded. Of 1,940 relatives, 187 had high TAFI levels. Annual incidences of venous thromboembolism were 0.23% in relatives with high TAFI levels versus 0.26% in relatives with normal TAFI levels (adjusted relative risk [RR] 0.8; 95% confidence interval [CI], 0.5-1.3). For arterial thrombosis these were 0.31% versus 0.23% (adjusted RR 1.4; 95% CI, 0.9-2.2). High levels of factor VIII, IX and XI were observed more frequently in relatives with high TAFI levels. Only high factor VIII levels were associated with an increased risk of venous and arterial thrombosis, independently of TAFI levels. None of these concomitant defects showed interaction with high TAFI levels. High TAFI levels were not associated with an increased risk of venous and arterial thromboembolism in thrombophilic families.

Association of ADAMDEC1 haplotype with high factor VIII levels in venous thromboembolism.

A suggestive locus on chromosome 8 could be shown to be associated with familial high factor VIII (FVIII) levels in venous thromboembolism. The ADAMDEC 1 gene is a candidate expressing an ectodomain sheddase. However, the ectodomain of the clearance receptor for FVIII, the low-density lipoprotein receptor-related protein (LRP), is subject to proteolysis by metalloproteases like ADAMDEC1. Other LRP-interacting proteins are lipoprotein lipase (LPL) and t-PA. For an association study, 165 thrombotic patients with high FVIII levels (from the MAISTHRO, i.e. Main-Isar-thrombosis register) were included. All patients with known causes for high FVIII levels had been previously excluded. The patients were compared with 214 healthy blood donors. Polymorphisms with usually a minor allele frequency >5%, i.e. 24 SNPs and two insertion/deletion polymorphisms of LPL gene, eight SNPs of the t-PA gene, and five SNPs of the ADAMDEC1 gene, were analyzed. Haplotype differences were calculated using PHASE. A new polymorphism in intron 7 of the t-PA gene with a minor allele frequency of 2.2% was identified. Analysis of each SNP by the Cochrane-Armitage trend test did not show any significant association between genotype and disease status. Interestingly, the ADAMDEC1 haplotype (rs12674766, rs10087305, rs2291577, rs2291578, rs3765124) differed between cases and controls (p = 0.04). In particular, the TGTGG haplotype showed a difference. In conclusion, the ADAMDEC 1 haplotype may indicate an underlying mechanism for high FVIII levels. The only moderate linkage disequilibrium may be due to a possible causal polymorphism in distant introns or the promoter region against a polygenic background.

Genetic variation in heme oxygenase 1 (HMOX1) and the risk of recurrent venous thromboembolism.

BACKGROUND/OBJECTIVE: Products of heme oxygenase 1 (HO1) possess antithrombotic properties, and impairment of HO1 activity may contribute to thrombus formation. Transcriptional activity of long GT-repeat alleles in HO1 gene (HMOX1) is lower as compared with short alleles. We hypothesize that these long alleles are associated with decreased HO1 anticoagulant activity and, thus, an increased risk of thrombosis.. DESIGN/METHODS: In a prospective cohort study, we followed 860 patients with a first VTE, and investigated the impact of a promoter GT-dinucleotid length polymorphism in HOMX1 on the risk of recurrent venous thromboembolism (VTE). RESULTS: Allele groups short (S), medium (M) and long (L) of the promoter GT-dinucleotide length polymorphism were distinguished. L-alleles, but not M- or S-alleles, were found to be more frequent among patients with recurrence. Heterozygous carriers of L-alleles had a two-fold higher relative risk of recurrence [(RR 2.2 (95% CI: 1.4-3.4)] as compared to wild type, which was independent of other thrombotic risk factors. At five years, the cumulative probability of recurrence was 18% (95% CI: 15%-22%) in patients without an L-allele compared to 32% (95% CI: 19%-46%) in patients heterozygous for the L-allele (P = .001). CONCLUSION: Patients with first VTE and long GT-repeat alleles in HMOX1 have an increased risk of recurrence. Genetically determined alterations in HO1 function may represent a new pathomechanism in VTE.