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Background: The inhibitory effect of adenosine on platelet aggregation is abrogated after the addition of adenosine-deaminase. Inosine is a naturally occurring nucleoside degraded from adenosine. Objectives: The mechanisms of antiplatelet action of adenosine and inosine in vitro and in vivo, and their differential biological effects by molecular modeling were investigated. Results: Adenosine (0.5, 1 and 2 mmol/L) inhibited phosphatidylserine exposure from 52±4% in the control group to 44±4 (p<0.05), 29±2 (p<0.01) and 20±3% (p<0.001). P-selectin expression in the presence of adenosine 0.5, 1 and 2 mmol/L was inhibited from 32±4 to 27±2 (p<0.05), 14±3 (p<0.01) and 9±3% (p<0.001), respectively. At the concentrations tested, only inosine to 4 mmol/L had effect on platelet P-selectin expression (p<0.05). Adenosine and inosine inhibited platelet aggregation and ATP release stimulated by ADP and collagen. Adenosine and inosine reduced collagen-induced platelet adhesion and aggregate formation under flow. At the same concentrations adenosine inhibited platelet aggregation, decreased the levels of sCD40L and increased intraplatelet cAMP. In addition, SQ22536 (an adenylate cyclase inhibitor) and ZM241385 (a potent adenosine receptor A2A antagonist) attenuated the effect of adenosine on platelet aggregation induced by ADP and intraplatelet level of cAMP. Adenosine and inosine significantly inhibited thrombosis formation in vivo (62±2% occlusion at 60 min [n = 6, p<0.01] and 72±1.9% occlusion at 60 min, [n = 6, p<0.05], respectively) compared with the control (98±2% occlusion at 60 min, n = 6). A2A is the adenosine receptor present in platelets; it is known that inosine is not an A2A ligand. Docking of adenosine and inosine inside A2A showed that the main difference is the formation by adenosine of an additional hydrogen bond between the NH2 of the adenine group and the residues Asn253 in H6 and Glu169 in EL2 of the A2A receptor. Conclusion: Therefore, adenosine and inosine may represent novel agents lowering the risk of arterial thrombosis. [ABSTRACT FROM AUTHOR]

Polymorphisms affecting platelet receptors and intracellular proteins have been extensively studied in relation to their potential influence in thrombosis and haemorrhages. However, few reports have addressed their impact on platelet function, with contradictory results. Limitations of these studies include, among others, small number of patients, the platelet functional parameters analyzed and their known variability in the healthy population. We studied the effect of six polymorphisms [ ITGB3 1565T > C (HPA-1), GPIBA variable number tandem repeat and 524C > T (HPA-2), ITGA2 807C > T, ADRA2A 1780A > G, and TUBB1 Q43P] on platelet function in 286 healthy subjects and their potential pathogenetic role in 160 patients with hereditary mucocutaneous bleeding of unknown cause. We found no effect of any of these polymorphisms on platelet aggregation, secretion, PFA-100®, and thrombin generation in platelet rich plasma. Furthermore, patients and controls showed no significant differences in the frequency of any of these polymorphisms. Thus, our study demonstrated that polymorphisms in genes affecting platelet function do not influence significantly major platelet functions and appear irrelevant in the pathogenesis of bleeding disorders. [ABSTRACT FROM AUTHOR]

It is known that polymorphisms of β 2-glycoprotein I ( β 2GPI) in exon 7 affect interaction between the phospholipid binding site and the antibodies, and that other polymorphisms in exon 8 increase the generation of antibodies. In this study, we analyzed genetic polymorphisms of β 2GPI in unselected Chilean patients to determine the prevalence of β 2GPI polymorphisms in the phospholipid domain in patients with venous and arterial thrombosis and the clinical correlation with thromboembolic complications. This study comprised 149 patients with venous and arterial thrombosis (62 with venous thrombosis and 87 with arterial thrombosis) and 160 healthy controls with no previous history of thrombosis. Polymorphisms of exons 7 and 8 of β 2GPI, which encode for its fifth domain, were determined by PCR-RFLP. The presence of aPL or anti- β 2GPI in the patients was detected by ELISA. Anti- β 2GPI were present in 8/149 patients (5.4%); of these, five had aCL antibodies of low titer. The allele containing Val/Leu247 and Trp/Ser316 was significantly more frequent in patients with thrombosis than in the control group (OR=3.1, CI 1.6–6.0, p=0.0003; OR=2.9, CI 1.1–8.6, p=0.027, respectively). These polymorphisms did not correlate with aPL or anti- β 2GPI but significant differences were observed with venous thrombosis ( p=<0.0001) and arterial thrombosis ( p=0.026). In conclusion, the β 2GPI polymorphisms Val/Leu247 and Trp/Ser316 are not related to the presence of anti- β 2GPI antibodies in unselected Chilean patients with venous and arterial thrombosis, but they are significantly associated with venous and arterial thrombosis. [ABSTRACT FROM AUTHOR]

Introduction. Antiphospholipid Antibodies (aPL) has been associated with thrombotic events. The objective of this study was to investigate the prevalence of aPL in a group of Chilean patients with venous and arterial thrombosis. Patients and Methods. Two hundred twenty six patients with venous and arterial thrombosis and 95 healthy controls were included in this study. Anticardiolipin (aCL), Anti-β2Glycoprotein I (anti-β2GPI) and Antiprothrombin (aPT) antibodies, were determined by ELISA using "in-house" assays. Results. Eighty eight out of 226 (38.9%) patients with thrombosis had some type of aPL. The prevalence of aCL, anti-β2GPI and aPT in patients with TV y TA, and in control group, shows in the following table: Twelve patients (5.3%) were positive for mixture of aPL. IgG, IgM and IgA isotypes were observed in aCL, Anti-β2GPI and aPT antibodies. Twenty four out of 78 (30.8%) patients with deep venous thrombosis, 2/14 (14.3%) with retinal venous thrombosis 17/68 (25.0%) with ischemic stroke and 14/66 (21.2%) with acute myocardial infarction, were positive for aCL antibodies. Anti-β2GPI and aPT antibodies were also present in these groups although in minor proportion. Conclusion. Our results show an important prevalence of aPL in patients with thrombosis, and aCL and aPT seem to be a risk factor of venous and arterial thrombosis. [ABSTRACT FROM AUTHOR]

Background and aims High plasma LDL-cholesterol (LDL-C) and platelet responses have major pathogenic roles in atherothrombosis. Thus, statins and anti-platelet drugs constitute mainstays in cardiovascular prevention/treatment. However, the role of platelet tissue factor-dependent procoagulant activity (TF-PCA) has remained unexplored in hypercholesterolemia. We aimed to study platelet TF-PCA and its relationship with membrane cholesterol in vitro and in 45 hypercholesterolemic patients (HC-patients) (LDL-C >3.37 mmol/L, 130 mg/dL) and 37 control subjects (LDL-C <3.37 mmol/L). The effect of 1-month administration of 80 mg/day atorvastatin (n = 21) and 20 mg/day rosuvastatin (n = 24) was compared. Methods Platelet TF-PCA was induced by GPIbα activation with VWF-ristocetin. Results Cholesterol-enriched platelets in vitro had augmented aggregation/secretion and platelet FXa generation (1.65-fold increase, p = 0.01). HC-patients had 1.5-, 2.3- and 2.5-fold increases in platelet cholesterol, TF protein and activity, respectively; their platelets had neither hyper-aggregation nor endogenous thrombin generation (ETP). Rosuvastatin, but not atorvastatin, normalized platelet cholesterol, TF protein and FXa generation. It also increased slightly the plasma HDL-C levels, which correlated negatively with TF-PCA. Conclusions Platelets from HC-patients were not hyper-responsive to low concentrations of classical agonists and had normal PRP-ETP, before and after statin administration. However, washed platelets from HC-patients had increased membrane cholesterol, TF protein and TF-PCA. The platelet TF-dependent PCA was specifically expressed after VWF-induced GPIbα activation. Rosuvastatin, but not atorvastatin treatment, normalized the membrane cholesterol, TF protein and TF-PCA in HC-patients, possibly unveiling a new pleiotropic effect of rosuvastatin. Modulation of platelet TF-PCA may become a novel target to prevent/treat atherothrombosis without increasing bleeding risks. [ABSTRACT FROM AUTHOR]

Inflammation, not hyperhomocysteinemia, is related to oxidative stress and hemostatic and endothelial dysfunction in uremia. Background. Several cardiovascular risk factors are present in patients with chronic renal failure (CRF), among which are systemic inflammation and hyperhomocysteinemia. Increased oxidative stress, endothelial activation/dysfunction, and coagulation activation are considered integral components of the inflammatory response, but have also been proposed as mediators of plasma homocysteine (tHcy)-induced cell damage. Using correlation analysis, we assessed the relative contributions of inflammation and hyperhomocysteinemia in the abnormal oxidative stress, endothelial activation/dysfunction, and hemostasis activation in patients with CRF. Methods. The relationships of inflammatory proteins and tHcy with plasma markers of these processes were studied in 64 patients with CRF (serum creatinine 526 ± 319 μmol/L) on conservative treatment, comparing the results with healthy controls (N = 15 to 40, depending on the measured variable) of similar sex and age. Results. Patients had significant increases in inflammatory cytokines (TNF-α and IL-8) and acute-phase proteins (C-reactive protein, fibrinogen and α1-antitrypsin). tHcy was increased in 87.5% of patients (mean = 27.1 μmol/L, range 6.5 to 118). Patients had significant increases in (1) indices of oxidative stress: TBARS (thiobarbituric acid-reactive species), a marker of lipid peroxidation and AOPP (advanced oxidation protein products), a marker of protein oxidation; (2) endothelial cell markers such as von Willebrand factor (vWF:Ag), soluble ICAM-1 and soluble thrombomodulin (sTM); (3) markers of intravascular thrombin generation: thrombin-antithrombin complexes (TAT) and prothrombin fragment F1+2 (PF1+2); and (4) indices of activation of fibrinolysis: plasmin-antiplasmin complexes (PAP), fibrin degradation products (FnDP) and fibrinogen... [ABSTRACT FROM AUTHOR]