Polymorphisms of F2 (G20210A), F5 (G1691A), F 7 (G10976A), F 13(G13T), FGB, ITGA2, ITGB3, PAI-I genes and its association with thrombotic complications in patients suffering from Takayasu aortoarteritis of Urals population.

OBJECTIVE: Cardiovascular complications, especially thrombotic events, are characteristic for Takayasu arteritis (TA). These events significantly deteriorate the patients' quality of life and cause disability and preterm death. Coagulation factor II (F2, G20210A), coagulation factor V (F5, G1691A, Leiden), coagulation factor VII (F7, G10976A), coagulation factor XIII (F13, G13T), fibrinogen (FGB), platelet alpha subunit of transmembrane receptor for collagens and related proteins (ITGA2), platelet glycoprotein (ITGB3), and plasminogen activator inhibitor-1 (PAI-I) gene polymorphisms coexist with TA, and their pathophysiologic interaction needs to be studied. METHODS: A total of 43 patients with TA were examined for nucleotides polymorphism in F2 (G20210A), F5 (G1691A, Leiden), F7 (G10976A), F13 (G13T), FGB, ITGA2, ITGB3, and PAI-I genes using polymerase chain reaction. Moreover, 130 sex- and age-adjusted healthy controls without a history of any thrombotic complications were enrolled. RESULTS: Among the patients with TA, there were 34 women aged between 17 and 77 (mean 49, median 49; Q1-Q3, 36-61) years and 9 men aged between 20 and 66 (mean 37.8, median 38; Q1-Q3: 31-45) years. Thrombotic complications were recorded in 22 (51%) patients with TA. Comparison of thrombophilia markers genotypes in patients with TA and healthy controls revealed homozygous and heterozygous mutation in ITGA2 (p<0.0001) and PAI-I genes (p=0.026). The frequency of occurrence of hereditary thrombophilia markers in patients with TA was assessed. Detection of the PAI-I gene mutation was significantly more frequent (p=0.032) in patients with TA with a history of thrombotic events than in those with no thrombosis history. Detection of multiple (more than 4 genes) simultaneous mutations of thrombophilia markers was significantly (p=0.0001) more frequent in patients with TA with a history of thrombotic events. CONCLUSION: Assessment of hereditary thrombophilia genetic markers reveals additional (genetic) risk markers of thrombotic complications in patients with TA and may help in decision making for antiplatelet and/or anticoagulant treatment in patients with TA to reduce the risk of thrombotic complications.

APOL1 Genetic Variants Are Associated With Increased Risk of Coronary Atherosclerotic Plaque Rupture in the Black Population.

[Figure: see text].

Identification of common and divergent gene expression signatures in patients with venous and arterial thrombosis using data from public repositories.

STRENGTHS AND LIMITATIONS OF THIS STUDY: Our results represent the first comparison of venous and arterial thrombosis at the transcriptomic level.Our main result was the demonstration that immunothrombosis pathways are important to the pathophysiology of these conditions, also at the transcriptomic level.A specific signature for venous and arterial thrombosis was described, and validated in independent cohorts.The limited number of public repositories with gene expression data from patients with venous thromboembolism limits the representation of these patients in our analyses.In order to gather a meaningful number of studies with gene expression data we had to include patients in different time-points since the index thrombotic event, which might have increased the heterogeneity of our population.

Comparison Between Clopidogrel and Prasugrel Associated With CYP2C19 Genotypes in Patients Receiving Percutaneous Coronary Intervention in a Japanese Population.

BACKGROUND: The association between cytochrome P450 (CYP) 2C19 genotypes and adverse events in patients treated with clopidogrel or prasugrel after percutaneous coronary intervention (PCI) in the Japanese population is unclear.Methods and Results:This study consisted of 1,580 patients whoseCYP2C19genotypes were assessed at Shiga University of Medical Science Hospital, and 193 clopidogrel-treated and 217 prasugrel-treated patients who were followed more than 1 year after receiving PCI were analyzed. Among 1,580 patients, the prevalence of normal, intermediate, and poor metabolizers was 32%, 49%, and 17%, respectively. Overall incidence of the primary outcome, defined as a composite of cardiovascular death, myocardial infarction, definite stent thrombosis, ischemic stroke, or major bleeding was not significantly different between the clopidogrel and prasugrel groups (adjusted hazard ratio [HR] 1.98, 95% confidence interval [CI] 0.85-4.61, P=0.12). Among patients with theCYP2C19loss-of-function (LOF) allele, however, the incidence of the primary outcome was significantly higher in the clopidogrel group (adjusted HR 3.19, 95% CI 1.10-9.24, P=0.03), whereas no difference was observed among patients without theCYP2C19LOF allele (adjusted HR 0.67, 95% CI 0.14-3.26, P=0.62). CONCLUSIONS: Among patients with theCYP2C19LOF allele, the use of clopidogrel was significantly associated with increased adverse events. Thus, further investigation is needed to establish the practical use ofCYP2C19genotyping.

Updated Expert Consensus Statement on Platelet Function and Genetic Testing for Guiding P2Y12 Receptor Inhibitor Treatment in Percutaneous Coronary Intervention.

Dual-antiplatelet therapy (DAPT) with aspirin and a P2Y12 receptor inhibitor is the standard treatment for patients undergoing percutaneous coronary intervention. The availability of different P2Y12 receptor inhibitors (clopidogrel, prasugrel, ticagrelor) with varying levels of potency has enabled physicians to contemplate individualized treatment regimens, which may include escalation or de-escalation of P2Y12-inhibiting therapy. Indeed, individualized and alternative DAPT strategies may be chosen according to the clinical setting (stable coronary artery disease vs. acute coronary syndrome), the stage of the disease (early- vs. long-term treatment), and patient risk for ischemic and bleeding complications. A tailored DAPT approach may be potentially guided by platelet function testing (PFT) or genetic testing. Although the routine use of PFT or genetic testing in percutaneous coronary intervention-treated patients is not recommended, recent data have led to an update in guideline recommendations that allow considering selective use of PFT for DAPT de-escalation. However, guidelines do not expand on when to implement the selective use of such assays into decision making for personalized treatment approaches. Therefore, an international expert consensus group of key leaders from North America, Asia, and Europe with expertise in the field of antiplatelet treatment was convened. This document updates 2 prior consensus papers on this topic and summarizes the contemporary updated expert consensus recommendations for the selective use of PFT or genotyping in patients undergoing percutaneous coronary intervention.

Association of the coronary artery disease risk gene GUCY1A3 with ischaemic events after coronary intervention.

AIM: A common genetic variant at the GUCY1A3 coronary artery disease locus has been shown to influence platelet aggregation. The risk of ischaemic events including stent thrombosis varies with the efficacy of aspirin to inhibit platelet reactivity. This study sought to investigate whether homozygous GUCY1A3 (rs7692387) risk allele carriers display higher on-aspirin platelet reactivity and risk of ischaemic events early after coronary intervention. METHODS AND RESULTS: The association of GUCY1A3 genotype and on-aspirin platelet reactivity was analysed in the genetics substudy of the ISAR-ASPI registry (n = 1678) using impedance aggregometry. The clinical outcome cardiovascular death or stent thrombosis within 30 days after stenting was investigated in a meta-analysis of substudies of the ISAR-ASPI registry, the PLATO trial (n = 3236), and the Utrecht Coronary Biobank (n = 1003) comprising a total 5917 patients. Homozygous GUCY1A3 risk allele carriers (GG) displayed increased on-aspirin platelet reactivity compared with non-risk allele (AA/AG) carriers [150 (interquartile range 91-209) vs. 134 (85-194) AU⋅min, P < 0.01]. More homozygous risk allele carriers, compared with non-risk allele carriers, were assigned to the high-risk group for ischaemic events (>203 AU⋅min; 29.5 vs. 24.2%, P = 0.02). Homozygous risk allele carriers were also at higher risk for cardiovascular death or stent thrombosis (hazard ratio 1.70, 95% confidence interval 1.08-2.68; P = 0.02). Bleeding risk was not altered. CONCLUSION: We conclude that homozygous GUCY1A3 risk allele carriers are at increased risk of cardiovascular death or stent thrombosis within 30 days after coronary stenting, likely due to higher on-aspirin platelet reactivity. Whether GUCY1A3 genotype helps to tailor antiplatelet treatment remains to be investigated.

Update on pathological platelet activation in coronary thrombosis.

Although coronary thrombosis (CT) is integral to cardiovascular outcomes, the underlying pathophysiological mechanisms remain unclear. CT may occur in case of atherosclerotic plaque erosion/rupture, or even after stenting implantation. Platelets (PLT) activation is the keystone of atherothrombosis and depends on many dysregulated elements, including endothelial dysfunction, oxidized lipoproteins, and immune response. Besides the classical view of PLT as an effector of hemostatic response, a new repertoire of PLT activities is emerging. PLT lipidome oxidation is a self-maintaining process which promotes PLT reactivity, coagulation cascade, and inflammatory cell activation. PLT-innate immune cell interaction is also sustained by neutrophil extracellular traps and NLRP3 inflammasome pathways. Other noteworthy emerging mechanisms are implicated in the crosstalk between PLT and surrounding cells. Especially, microvesicles (MVs) released from PLT may extend their signaling network far beyond the classical cell-cell interactions. Moreover, the recognition of noncoding RNA in PLT MVs introduce another layer of complexity in terms of intercellular signaling by a direct regulation of messenger RNA profile and gene expression in the recipient cells. The aim of this narrative review is to update the recent advance in CT and intracoronary stent thrombosis, including causal factors and potential translation of experimental evidence into the clinical setting.

Sub-Chronic Stress Exacerbates the Pro-Thrombotic Phenotype in BDNFVal/Met Mice: Gene-Environment Interaction in the Modulation of Arterial Thrombosis.

Brain-Derived Neurotrophic Factor (BDNF) Val66Met polymorphism has been associated with increased susceptibility to develop mood disorders and recently it has been also linked with cardiovascular disease (CVD). Interestingly, stressful conditions unveil the anxious/depressive-like behavioral phenotype in heterozygous BDNFVal66Met (BDNFVal/Met) mice, suggesting an important relationship in terms of gene-environment interaction (GxE). However, the interplay between stress and BDNFVal/Met in relation to CVD is completely unknown. Here, we showed that BDNFVal/Met mice display a greater propensity to arterial thrombosis than wild type BDNFVal/Val mice after 7 days of restraint stress (RS). RS markedly increased the number of leukocytes and platelets, and induced hyper-responsive platelets as showed by increased circulating platelet/leukocyte aggregates and enhanced expression of P-selectin and GPIIbIIIa in heterozygous mutant mice. In addition, stressed BDNFVal/Met mice had a greater number of large and reticulated platelets but comparable number and maturation profile of bone marrow megakaryocytes compared to BDNFVal/Val mice. Interestingly, RS led to a significant reduction of BDNF expression accompanied by an increased activity of tissue factor in the aorta of both BDNFVal/Val and BDNFVal/Met mice. In conclusion, we provide evidence that sub-chronic stress unveils prothrombotic phenotype in heterozygous BDNF Val66Met mice affecting both the number and functionality of blood circulating cells, and the expression of key thrombotic molecules in aorta. Human studies will be crucial to understand whether this GxE interaction need to be taken into account in risk stratification of coronary artery disease (CAD) patients.

Advocating cardiovascular precision medicine with P2Y12 receptor inhibitors.

Antiplatelet therapy with P2Y12-receptor inhibitors has become the cornerstone of medical treatment in patients with acute coronary syndrome treated with percutaneous coronary intervention (PCI). With over 100 million prescriptions filled since its approval, clopidogrel is the most widely used P2Y12-receptor inhibitor. Dual antiplatelet therapy with clopidogrel plus aspirin has been associated with a lower rate of major cardiovascular events in patients after PCI than aspirin monotherapy. However, an alarmingly high number of clopidogrel-treated patients experience adverse thrombotic events. Insufficient P2Y12-inhibition or high on-treatment platelet reactivity to adenosine diphosphate has stimulated the increased use of more potent P2Y12 inhibitors: prasugrel (a third generation thienopyridine) and ticagrelor (a cyclopentyl-triazolo-pyrimidine). However, more potent platelet inhibition and low on-treatment platelet reactivity has resulted in increased major bleeding and higher costs. These limitations have suggested the need for an individualized antiplatelet approach in order to decrease thrombotic events and minimize bleeding. This model of personalized medicine integrates a patient's demographic and biological data (pharmacodynamic, genomic, epigenomic, transcriptomic, and metabolic information) to target therapy in order to maximize efficacy while minimizing bleeding and costs. This review discusses the role of diagnostic tools such as platelet function and pharmacogenomic testing to personalize antiplatelet therapy.

The role of microRNAs in prethrombotic status associated with coronary artery disease.

The acute cardiovascular events following thrombus formation is a primary cause of morbidity and mortality of patients with coronary artery disease (CAD). Numerous studies have shown that a prethrombotic status, which can be defined as an imbalance between the procoagulant and anticoagulant conditions, would exist for a period of time before thrombogenesis. Therefore, early diagnosis and intervention of prethrombotic status are important for reducing acute cardiovascular events. However, none of prethrombotic indicators have been identified as golden standard for diagnosis of prethrombotic status to date. MicroRNAs (miRNAs), a class of short non-coding RNAs, have been shown to be involved in pathophysiologic processes related to prethrombotic status, such as endothelial dysfunction, platelet activation, impaired fibrinolysis and elevated procoagulant factors, etc. Owing to their multiple and fine-tuning impacts on gene expression, miRNAs raise a novel understanding in the underlying mechanism of prethrombotic status. This review aims to discuss the role of miRNAs in prethrombotic status, especially the differently expressed miRNAs in CAD, which may be meaningful for developing promising diagnostic biomarkers and therapeutic strategies for CAD patients in future.

Granulocytes in coronary thrombus evolution after myocardial infarction--time-dependent changes in expression of matrix metalloproteinases.

BACKGROUND: Remodeling of extracellular matrix is a key process during wound healing, which is strictly regulated by matrix metalloproteinases (MMPs) and their tissue inhibitors [tissue inhibitors of metalloproteinases (TIMPs)]. In this study, we evaluated intrathrombotic MMPs and TIMPs and their cellular origin during thrombus evolution after disruption of coronary atherosclerotic plaque. MATERIALS AND METHODS: Thrombectomy materials (N=120) obtained from patients with acute myocardial infarction were histologically classified in three groups based on thrombus age: fresh (<1day), lytic (1-5days), or organized (>5days) thrombi; materials showing a heterogeneous composition were classified according to oldest part. Presence and cellular origin of MMPs (MMP-1, MMP-2, MMP-8, MMP-9, and MMP-14) and TIMPs (TIMP-1, TIMP-2, and TIMP-3) was evaluated with immunostains (double) and with polymerase chain reaction. RESULTS AND CONCLUSION: MMPs and TIMPs were present in all the thrombectomy samples. A distinct temporal change in extent and cellular origin of MMPs and TIMPs during thrombus evolution was observed. In the early (fresh and lytic) stages of thrombus, high numbers of neutrophilic granulocytes occupy the thrombus mass and produce large amounts of MMPs and TIMPs. However, with progression of thrombus evolution (organizing stage) and diminishment of neutrophil granulocytes, there is disappearance of MMP-8 and MMP-9, steep decline of MMP-1 and TIMP-2, and progressive decrease of TIMP-3. In contrast, intrathrombotic MMP-2 and MMP-14 are present at a constant high level during the entire process of thrombus evolution. These temporal changes indicate a complex time-dependent function of MMPs, which are largely granulocyte derived, in the healing process of thrombus after plaque disruption.

IFN-γ-producing Th17 cells bias by HMGB1-T-bet/RUNX3 axis might contribute to progression of coronary artery atherosclerosis.

BACKGROUND: IFN-γ-producing Th17 cells have been implicated in autoimmune disorders, but their properties in humans are known only partially. The molecular mechanisms and external factors that govern IFN-γ-producing Th17-cell bias are incompletely understood. The present work was to clarify whether (i) IFN-γ-producing Th17 cells are present in the peripheral circulation of patients with coronary atherosclerosis (CA); (ii) high mobility group box (HMGB)1 in circulation is associated with IFN-γ-producing Th17-cell bias. METHODS: Thirty-six patients (17 females and 19 males; 45-84 years) diagnosed as having atherosclerosis after coronary angiography for suspected or known CA were included the study cohort. Samples of peripheral blood were collected from healthy volunteers and patients, and classical tests (flow cytometry, RT-qPCR) were used to measure blood components. RESULTS AND CONCLUSION: Our results clearly demonstrated that HMGB1 were up-regulated in different progressive CA patients: 5.38 ± 1.48 ng/ml, 6.30 ± 1.53 ng/ml and 5.86 ± 1.12 ng/ml vs1.45 ± 0.65 ng/ml for only atherosclerotic plaque (AP), atherosclerotic plaque and some plaque rupture, no thrombosis (PR), plaque rupture and accompanying thrombosis (TH) and volunteers, respectively, p < 0.05. The frequency of IFN-γ-producing Th17 cells was 2.33 ± 0.58%, 1.93 ± 0.2% and 2.21 ± 0.65% vs 0.38 ± 0.21% for AP, PR, TH and volunteers, p < 0.05, respectively. Furthermore, HMGB1 contributed to IFN-γ-producing Th17-cell bias by controlling expression of T-bet and RUNX3. We demonstrated, for the first time, that HMGB1 is a potential inducer of IFN-γ-producing Th17-cell bias, and that IFN-γ-producing Th17 cells might be one of the pathogenic factors in atherosclerosis.

[The molecular mechanisms of platelets activation in patients with cerebrovascular disease].

Cerebrovascular disease is a main cause of mortality and one of the big medical problems. After the vascular wall's damage the endothelial cells secrete the von Willebrand factor which then connects with its platelet's receptor GP Ib-V-IX. There are two polymorphisms Thr145Met and T(-5)C of the GP Iba gene associated with arterial thrombosis development. Also the difference in platelets' genes expressions was shown in patients with various clinical course of ischemic heart disease. The aim of this study was to investigate the role of platelet's receptor for von Willebrand factor in platelets' activation in patients with cerebrovascular disease. 123 patients with cerebrovascular disease and 97 healthy donors were included into the study. We analyzed the level of receptor for von Willebrand factor on platelet's membrane by flow cytometry, Thr145Met and T(-5)C GP Iba polymorphiams by PCR-RFLP, the GP Iba gene expression by RT-PCR and ADP-induced platelet aggregation by Born method. We have shown: 1) the 145Met GP Iba allele prevalence in patients with atherotrombotic stroke development due to macroangiopathy; 2) the pre-mRNA transform into the mature mRNA in activated platelets and this process may be stopped by the antiplatelet therapy by acetylsalicylic acid.

Human coronary thrombus formation is associated with degree of plaque disruption and expression of tissue factor and hexokinase II.

BACKGROUND: Atherosclerotic plaque thrombogenicity is a critical factor that affects thrombus formation and the onset of acute myocardial infarction (AMI). The aim of this study was to identify the vascular factors involved in thrombus formation and AMI onset. METHODS AND RESULTS: Culprit lesions in 40 coronary arteries with thrombi at autopsy after lethal AMI and non-cardiac death (asymptomatic plaque disruption) were analyzed on histology. Thrombus size, ratio of thrombus to lumen area, length of plaque disruption, and immunopositive areas for tissue factor (TF) and hexokinase (HK)-II were significantly larger in coronary arteries with AMI than with asymptomatic plaque disruption. The size of coronary thrombus positively correlated with the length of plaque disruption (r=0.80) and with immunopositive areas for TF (r=0.38) and HK-II (r=0.40). Because both M1 and M2 macrophages express TF and HK-II in symptomatic plaques, we assessed TF and HK-II expression in M1- and M2-polarized macrophages. The expression of TF was increased and that of HK-II was decreased in M2-, compared with M1-polarized THP-1 macrophages. Inhibiting glycolysis enhanced TF expression in the macrophages partly via hypoxia inducible factor-1α. CONCLUSIONS: The degree of plaque disruption and expression of TF and HK-II appear to be important vascular factors for AMI onset, and polarized macrophages make a distinct contribution to thrombogenicity and glucose metabolism.

Cytochrome P450 CYP 2C19*2 Associated with Adverse 1-Year Cardiovascular Events in Patients with Acute Coronary Syndrome.

BACKGROUND: The cytochrome P450 (CYP450) 2C19 681 genotypes affect the antiplatelet activity of clopidogrel. We investigated the correlation of CYP 2C19 681G > A mutation with clopidogrel resistance (CR). Additionally, we studied the effect of CR on clinical prognosis of patients with acute coronary syndrome (ACS). METHODS: One hundred ten ACS patients undergoing percutaneous coronary intervention, who were followed-up for 1 year, were included in the study. The patients were co-administered aspirin 100 mg/d and clopidogrel 75mg/d following a loading dose of 300 mg. CR was assessed on the basis of polymorphism observed in the CYP2C19 subgroup. RESULTS: Patients in GG genotype group exhibited greater inhibition of platelet aggregation than patients in GA and AA genotype groups (16.2 ± 10.1%; 10.2 ± 9.9%; 8.0 ± 5.9%, respectively, p < 0.01). CYP2C19 681GG genotype group was associated with lower CR than CYP2C19 681A allele (GA + AA) group (9/59 vs. (12+5)/51; p = 0.009). Over a follow-up of 12 months, the incidence of recurrent angina, acute myocardial infarction, and intra-stent thrombosis in CYP2C19 681 GG carriers was significantly lower than that in CYP2C19 681A allele (GA + AA) group (2/59 vs. 8/51, 1/59 vs. 6/51, 0 vs. 4/51, respectively, p < 0.05). CONCLUSION: CYP 2C19*2 is associated with reduced clopidogrel antiplatelet activity and might be an important marker for poor prognosis of ACS.

Genetic testing and antiplatelet treatment: Still way to go?

Despite medical and technical advancements stent thrombosis continued to poses a significant risk in patients after drug eluting stent (DES) implantation and clopidogrel resistance has been recognized as an important determinant of this risk. Novel antiplatelets such as prasugrel and ticagrelor can be used in cases of clopidogrel resistance however bleeding complications remain the Achilles' heel of antiplatelet therapy. Several genetic polymorphisms affect the clopidogrel absorption and bio-activation in the active form of the drug. CYP2C19 is responsible for most of the clopidogrel bio-transformation and loss of function as well as, gain of function polymorphisms of this enzyme has been recognized. Early studies have linked CYP2C19 genetic polymorphisms with clinical events, although, these findings were not confirmed by later studies. However, when the estimated thrombotic risk is high a combination of genetic information and functional platelet tests should be essential for clinicians to access clopidogrel effectiveness and to recommend alternative antiplatelet management.