Combined role of the Lewis antigenic system, Chlamydia pneumoniae, and C-reactive protein in unstable angina

AbstractObjectivesThe goal of this study was to assess the prognostic role of the Lewis antigenic system, Chlamydia pneumoniae(CP) seropositivity (CP+), and C-reactive protein (CRP) levels in unstable angina (UA).BackgroundThe role of CP infection in acute coronary syndromes is contradictory. The Lewis antigenic system, a genetically determined blood group system associated with infections and several disorders, including ischemic heart disease, might influence the susceptibility to CP infection, inflammatory response, and risk of cardiac ischemic events.MethodsThe CRP levels, Lewis antigens, and CP+ were measured in 54 patients with Braunwald’s class IIIB UA. All patients were followed up for one year, and the occurrence of new coronary events (coronary death, myocardial infarction, and recurrence of instability) were recorded.ResultsTwenty-five coronary events occurred during follow-up. At univariate analysis CRP >3 mg/l (CRP+) (p = 0.0056), Lewis antigen b (Leb+) (p = 0.028), and the combination of Leb+ and CP+ (p = 0.006) and of CRP+ and Leb+ (p = 0.003) were associated with new coronary events, while CP+ alone was not. At multivariate analysis, CRP+ (p = 0.008) and combined Leb+CP+ (p = 0.03) were independent predictors of worse outcome. The event rate was 64% in CRP+ patients, 67% in Leb+CP+ patients, and 86% in CRP+Leb+CP+ patients. Combined Leb+CP+, but not Leb+ and CP+ alone, was related to CRP levels (p = 0.03). Among CP+ patients, CRP levels were higher in Leb+ than Leb- (p = 0.028).ConclusionsOur data demonstrate that in UA the Lewis antigenic system plays an important role, probably determining individual susceptibility to the detrimental effects of CP infection and by determining an enhanced inflammatory response

Surgical bleeding after pre-operative unfractionated heparin and low molecular weight heparin for coronary bypass surgery

Background and Objectives Since the impairment of platelet function may cause excess peri-operative bleeding, pre-operative discontinuation of aspirin and heparin bridging are common for cardiac surgery. We evaluated the impact of pre-operative administration of enoxaparin and unfractionated heparin (UFH) on coagulation parameters and peri-operative bleeding in patients undergoing elective coronary artery bypass grafting (CABG) surgery after discontinuation of aspirin. Design and Methods Forty-three patients with three-vessel coronary artery disease undergoing elective CABG surgery discontinued aspirin and were randomized to receive either UFH 180 UI/Kg × 2/day s.c. or enoxaparin 100 UI/Kg × 2/day s.c. until 12 h before surgery (median pre-operative treatment 8 days, range 6–12 days). Surgery was performed as usual with UFH. Neither UFH nor any low molecular weight heparin was given in the immediate post-operative period. The effects of UFH and enoxaparin were monitored by the activated partial thromboplastin time (aPTT) and the Enox-test (sensitive to factor Xa inhibition) using a Rapidpoint® Coagulation Analyzer. aPTT and factor Xa activity were also measured by standard methods. Peri-operative bleeding and the nadirs of hemoglobin concentration, hematocrit and platelet count were monitored post-operatively. Results Patients in the two groups were similar for number of bypasses, on-pump time, total surgery time, and time from the last heparin administration. Coagulation parameters increased significantly and similarly at 30 min and 6 h with both treatments, but returned within the normal range at 12 h. Hemoglobin, hematocrit and platelet counts significantly decreased to the same extent after CABG and re-normalized at the same time. Transfusional requirements of blood and plasma units were similar in the two groups. Interpretation and Conclusions From the kinetics of coagulation parameters and the evaluation of bleeding, enoxaparin is a safe alternative to UFH as a bridging therapy to CABG after discontinuation of aspirin

Ibrutinib does not affect ristocetin-induced platelet aggregation evaluated by light transmission aggregometry in chronic lymphocytic leukemia patients

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Evaluation of Novel Guanidino-Containing Isonipecotamide Inhibitors of Blood Coagulation Factors against SARS-CoV-2 Virus Infection

Coagulation factor Xa (fXa) and thrombin (thr) are widely expressed in pulmonary tissues, where they may catalyze, together with the transmembrane serine protease 2 (TMPRSS2), the coronaviruses spike protein (SP) cleavage and activation, thus enhancing the SP binding to ACE2 and cell infection. In this study, we evaluate in vitro the ability of approved (i.e., dabigatran and rivaroxaban) and newly synthesized isonipecotamide-based reversible inhibitors of fXa/thr (cmpds 1-3) to hinder the SARS-CoV-2 infectivity of VERO cells. Nafamostat, which is a guanidine/amidine antithrombin and antiplasmin agent, disclosed as a covalent inhibitor of TMPRSS2, was also evaluated. While dabigatran and rivaroxaban at 100 mu M concentration did not show any effect on SARS-CoV-2 infection, the virus preincubation with new guanidino-containing fXa-selective inhibitors 1 and 3 did decrease viral infectivity of VERO cells at subtoxic doses. When the cells were pre-incubated with 3, a reversible nanomolar inhibitor of fXa (K-i = 15 nM) showing the best in silico docking score toward TMPRSS2 (pdb 7MEQ), the SARS-CoV-2 infectivity was completely inhibited at 100 mu M (p < 0.0001), where the cytopathic effect was just about 10%. The inhibitory effects of 3 on SARS-CoV-2 infection was evident (ca. 30%) at lower concentrations (3-50 mu M). The covalent TMPRSS2 and the selective inhibitor nafamostat mesylate, although showing some effect (15-20% inhibition), did not achieve statistically significant activity against SARS-CoV-2 infection in the whole range of test concentrations (3-100 mu M). These findings suggest that direct inhibitors of the main serine proteases of the blood coagulation cascade may have potential in SARS-CoV-2 drug discovery. Furthermore, they prove that basic amidino-containing fXa inhibitors with a higher docking score towards TMPRSS2 may be considered hits for optimizing novel small molecules protecting guest cells from SARS-CoV-2 infection

Abnormal proplatelet formation and emperipolesis in cultured human megakaryocytes from gray platelet syndrome patients

none10siThe Gray Platelet Syndrome (GPS) is a rare inherited bleeding disorder characterized by deficiency of platelet α-granules, macrothrombocytopenia and marrow fibrosis. The autosomal recessive form of GPS is linked to loss of function mutations in NBEAL2, which is predicted to regulate granule trafficking in megakaryocytes, the platelet progenitors. We report the first analysis of cultured megakaryocytes from GPS patients with NBEAL2 mutations. Megakaryocytes cultured from peripheral blood or bone marrow hematopoietic progenitor cells from four patients were used to investigate megakaryopoiesis, megakaryocyte morphology and platelet formation. In vitro differentiation of megakaryocytes was normal, whereas we observed deficiency of megakaryocyte α-granule proteins and emperipolesis. Importantly, we first demonstrated that platelet formation by GPS megakaryocytes was severely affected, a defect which might be the major cause of thrombocytopenia in patients. These results demonstrate that cultured megakaryocytes from GPS patients provide a valuable model to understand the pathogenesis of GPS in humans.openDi Buduo, Christian A.; Alberelli, Maria Adele; Glembostky, Ana C.; Podda, Gianmarco; Lev, Paola R.; Cattaneo, Marco; Landolfi, Raffaele; Heller, Paula G.; Balduini, Alessandra; De Candia, EricaDI BUDUO, CHRISTIAN ANDREA; Alberelli, Maria Adele; Glembostky, Ana C.; Podda, Gianmarco; Lev, Paola R.; Cattaneo, Marco; Landolfi, Raffaele; Heller, Paula G.; Balduini, Alessandra; De Candia, Eric

Detection of Platelet-Activating Antibodies Associated with Vaccine-Induced Thrombotic Thrombocytopenia by Flow Cytometry: An Italian Experience

Rare cases of thrombocytopenia and thrombosis after anti-COVID-19 adenovirus-associated mRNA vaccines (VITT) due to platelet-activating anti-platelet-factor 4 (PF4)/polyanion antibod-ies have been reported. VITT laboratory diagnosis, similarly to heparin-induced thrombocytope-nia (HIT) diagnosis, requires immunoassays for anti-PF4/polyanion antibodies identification, such as ELISA assays and platelet-activating functional tests, such as heparin-induced platelet activation test (HIPA), to confirm their pathogenicity. We compared the flow cytometry (FC) measurement of platelet p-selectin exposure to the gold standard functional test HIPA for diagno-sis confirmation in 13 patients with a clinical VITT syndrome (6M/7F; median age 56 (33–78)) who resulted positive to anti-PF4/polyanion antibodies ELISA assays (12/13). FC and HIPA sim-ilarly identified three different patterns: (1) a typical non-heparin-dependent VITT pattern (seven and six patients by FC and HIPA, respectively); (2) low/no platelet activation in patients under IvIg therapy (five out of five and two out of four patients by FC and HIPA, respectively); (3) a HIT pattern. Antibodies investigated by FC became negative after 7, 17, and 24 days of therapy in three patients. FC measurement of P-selectin exposure was as sensitive as HIPA but simpler to de-tect anti-PF4/polyanion antibodies in VITT patients. FC could reliably discriminate VITT from HIT, thus helping for the choice of the anticoagulant

Platelets and Neurodegenerative Diseases: Current Knowledge and Future Perspectives

: Platelets have a fundamental role in mediating hemostasis and thrombosis. However, more recently, a new idea is making headway, highlighting the importance of platelets as significant actors in modulating immune and inflammatory responses. In particular, platelets have an important role in the development of vascular amyloid-b-peptide(ab) deposits, known to play a relevant role in Alzheimer's disease (AD) through accumulation and deposition within the frontal cortex and hippocampus in the brain. The involvement of platelets in the pathogenesis of AD opens up the highly attractive possibility of applying antiplatelet therapy for the treatment and/or prevention of AD, but conclusive results are scarce. Even less is known about the potential role of platelets in mild cognitive impairment (MCI). The aim to this brief review is to summarize current knowledge on this topic and to introduce the new perspectives on the possible role of platelet activation as therapeutic target both in AD and MCI

Analysis of 339 pregnancies in 181 women with 13 different forms of inherited thrombocytopenia

Pregnancy in women with inherited thrombocytopenias is a major matter of concern as both the mothers and the newborns are potentially at risk of bleeding. However, medical management of this condition cannot be based on evidence because of the lack of consistent information in the literature. To advance knowledge on this matter, we performed a multicentric, retrospective study evaluating 339 pregnancies in 181 women with 13 different forms of inherited thrombocytopenia. Neither the degree of thrombocytopenia nor the severity of bleeding tendency worsened during pregnancy and the course of pregnancy did not differ from that of healthy subjects in terms of miscarriages, fetal bleeding and pre-term births. The degree of thrombocytopenia in the babies was similar to that in the mother. Only 7 of 156 affected newborns had delivery-related bleeding, but 2 of them died of cerebral hemorrhage. The frequency of delivery-related maternal bleeding ranged from 6.8% to 14.2% depending on the definition of abnormal blood loss, suggesting that the risk of abnormal blood loss was increased with respect to the general population. However, no mother died or had to undergo hysterectomy to arrest bleeding. The search for parameters predicting delivery-related bleeding in the mother suggested that hemorrhages requiring blood transfusion were more frequent in women with history of severe bleedings before pregnancy and with platelet count at delivery below 50 × 10(9)/L.Fil: Noris, Patrizia. Istituti di Ricovero e Cura a Carattere Scientifico. Policlinico San Matteo di Pavia; Italia. Università degli Studi di Pavia; ItaliaFil: Schlegel, Nicole. Université Paris Diderot - Paris 7; FranciaFil: Klersy, Catherine. Istituti di Ricovero e Cura a Carattere Scientifico. Policlinico San Matteo di Pavia; ItaliaFil: Heller, Paula Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Civaschi, Elisa. Università degli Studi di Pavia; ItaliaFil: Pujol Moix, Nuria. Universitat Autònoma de Barcelona; EspañaFil: Fabris, Fabrizio. Università di Padova; ItaliaFil: Favier, Remi. Inserm; Francia. Armand Trousseau Children’s Hospital; Francia. French Reference Center for Inherited Platelet disorders; FranciaFil: Gresele, Paolo. Università di Perugia; ItaliaFil: Latger Cannard, Véronique. Centre Hospitalo-Universitaire. Service d’Hématologie Biologique; Francia. Reference French Centre. Centre de Compétence Nord-Est des Pathologies Plaquettaires; FranciaFil: Cuker, Adam. University of Pennsylvania; Estados UnidosFil: Nurden, Paquita. Hôpital Xavier Arnozan; FranciaFil: Greinacher, Andreas. Institut für Immunologie und Transfusionsmedizin; AlemaniaFil: Cattaneo, Marco. Università degli Studi di Milano; ItaliaFil: De Candia, Erica. Università Cattolica del Sacro Cuore; ItaliaFil: Pecci, Alessandro. Università degli Studi di Pavia; ItaliaFil: Hurtaud Roux, Marie Françoise. Université Paris Diderot - Paris 7; FranciaFil: Glembotsky, Ana Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Muñiz Diaz, Eduardo. Banc de Sang i Teixits de Catalunya. Immunohematology Department; EspañaFil: Randi, Maria Luigia. Università di Padova; ItaliaFil: Trillot, Nathalie. Centre Hospitalier Régional Universitaire de Lille. Pôle Biologie Pathologie Génétique. Institut d’Hématologie-Transfusion; FranciaFil: Bury, Loredana. Università di Perugia; ItaliaFil: Lecompte, Thomas. Hôpitaux Universitaires de Genève; Suiza. Université de Genève. Faculté de Médecine; SuizaFil: Marconi, Caterina. Università di Bologna; ItaliaFil: Savoia, Anna. Università degli Studi di Trieste; ItaliaFil: Balduini, Carlo L.. Istituti di Ricovero e Cura a Carattere Scientifico Burlo Garofolo. Institute for Maternal and Child Health; Italia. Università degli Studi di Pavia; ItaliaFil: European Hematology Association Scientific Working Group on Thrombocytopenias and Platelet Function Disorders. No especifica

Analysis of 339 pregnancies in 181 women with 13 different forms of inherited thrombocytopenia

65Pregnancy in women with inherited thrombocytopenias is a major matter of concern as both the mothers and the newborns are potentially at risk of bleeding. However, medical management of this condition cannot be based on evidence because of the lack of consistent information in the literature. To advance knowledge on this matter, we performed a multicentric, retrospective study evaluating 339 pregnancies in 181 women with 13 different forms of inherited thrombocytopenia. Neither the degree of thrombocytopenia nor the severity of bleeding tendency worsened during pregnancy and the course of pregnancy did not differ from that of healthy subjects in terms of miscarriages, fetal bleeding and pre-term births. The degree of thrombocytopenia in the babies was similar to that in the mother. Only 7 of 156 affected newborns had delivery-related bleeding, but 2 of them died of cerebral hemorrhage. The frequency of delivery-related maternal bleeding ranged from 6.8% to 14.2% depending on the definition of abnormal blood loss, suggesting that the risk of abnormal blood loss was increased with respect to the general population. However, no mother died or had to undergo hysterectomy to arrest bleeding. The search for parameters predicting delivery-related bleeding in the mother suggested that hemorrhages requiring blood transfusion were more frequent in women with history of severe bleedings before pregnancy and with platelet count at delivery below 50 × 10(9)/L.openopenPatrizia Noris; Nicole Schlegel; Catherine Klersy; Paula G. Heller; Elisa Civaschi; Nuria Pujol-Moix; Fabrizio Fabris; Remi Favier; Paolo Gresele; Véronique Latger-Cannard; Adam Cuker; Paquita Nurden; Andreas Greinacher; Marco Cattaneo; Erica De Candia; Alessandro Pecci; Marie-Françoise Hurtaud-Roux; Ana C. Glembotsky; Eduardo Muñiz-Diaz; Maria Luigia Randi; Nathalie Trillot; Loredana Bury; Thomas Lecompte; Caterina Marconi; Anna Savoia; Carlo L. Balduini; Sophie Bayart; Anne Bauters; Schéhérazade Benabdallah-Guedira; Françoise Boehlen; Jeanne-Yvonne Borg; Roberta Bottega; James Bussel; Daniela De Rocco; Emmanuel de Maistre; Michela Faleschini; Emanuela Falcinelli; Silvia Ferrari; Alina Ferster; Tiziana Fierro; Dominique Fleury; Pierre Fontana; Chloé James; Francois Lanza; Véronique Le Cam Duchez; Giuseppe Loffredo; Pamela Magini; Dominique Martin-Coignard; Fanny Menard; Sandra Mercier; Annamaria Mezzasoma; Pietro Minuz; Ilaria Nichele; Lucia D. Notarangelo; Tommaso Pippucci; Gian Marco Podda; Catherine Pouymayou; Agnes Rigouzzo; Bruno Royer; Pierre Sie; Virginie Siguret; Catherine Trichet; Alessandra Tucci; Béatrice Saposnik; Dino VeneriPatrizia, Noris; Nicole, Schlegel; Catherine, Klersy; Paula G., Heller; Elisa, Civaschi; Nuria Pujol, Moix; Fabrizio, Fabris; Remi, Favier; Paolo, Gresele; Véronique Latger, Cannard; Adam, Cuker; Paquita, Nurden; Andreas, Greinacher; Marco, Cattaneo; Erica De, Candia; Alessandro, Pecci; Marie Françoise Hurtaud, Roux; Ana C., Glembotsky; Eduardo Muñiz, Diaz; Maria Luigia, Randi; Nathalie, Trillot; Loredana, Bury; Thomas, Lecompte; Caterina, Marconi; Savoia, Anna; Carlo L., Balduini; Sophie, Bayart; Anne, Bauters; Schéhérazade Benabdallah, Guedira; Françoise, Boehlen; Jeanne Yvonne, Borg; Bottega, Roberta; James, Bussel; DE ROCCO, Daniela; Emmanuel de, Maistre; Faleschini, Michela; Emanuela, Falcinelli; Silvia, Ferrari; Alina, Ferster; Tiziana, Fierro; Dominique, Fleury; Pierre, Fontana; Chloé, James; Francois, Lanza; Véronique Le Cam, Duchez; Giuseppe, Loffredo; Pamela, Magini; Dominique Martin, Coignard; Fanny, Menard; Sandra, Mercier; Annamaria, Mezzasoma; Pietro, Minuz; Ilaria, Nichele; Lucia D., Notarangelo; Tommaso, Pippucci; Gian Marco, Podda; Catherine, Pouymayou; Agnes, Rigouzzo; Bruno, Royer; Pierre, Sie; Virginie, Siguret; Catherine, Trichet; Alessandra, Tucci; Béatrice, Saposnik; Dino, Vener

Cell Surface Platelet Tissue Factor Expression: Regulation by P2Y12 and Link to Residual Platelet Reactivity

BACKGROUND: ADP-induced platelet activation leads to cell surface expression of several proteins, including TF (tissue factor). The role of ADP receptors in platelet TF modulation is still unknown. We aimed to assess the (1) involvement of P2Y(1) and P2Y(12) receptors in ADP-induced TF exposure; (2) modulation of TFpos-platelets in anti-P2Y(12)-treated patients with coronary artery disease. Based on the obtained results, we revisited the intracellular localization of TF in platelets.METHODS: The effects of P2Y(1) or P2Y(12) antagonists on ADP-induced TF expression and activity were analyzed in vitro by flow cytometry and thrombin generation assay in blood from healthy subjects, P2Y(12)(-/-), and patients with gray platelet syndrome. Ex vivo, P2Y(12) inhibition of TF expression by clopidogrel/prasugrel/ticagrelor, assessed by VASP (vasodilator-stimulated phosphoprotein) platelet reactivity index, was investigated in coronary artery disease (n=238). Inhibition of open canalicular system externalization and electron microscopy (TEM) were used for TF localization.RESULTS: In blood from healthy subjects, stimulated in vitro by ADP, the percentage of TFpos-platelets (17.3 +/- 5.5%) was significantly reduced in a concentration-dependent manner by P2Y(12) inhibition only (-81.7 +/- 9.5% with 100 nM AR-C69931MX). In coronary artery disease, inhibition of P2Y(12) is paralleled by reduction of ADP-induced platelet TF expression (VASP platelet reactivity index: 17.9 +/- 11%, 20.9 +/- 11.3%, 40.3 +/- 13%; TFpos-platelets: 10.5 +/- 4.8%, 9.8 +/- 5.9%, 13.6 +/- 6.3%, in prasugrel/ticagrelor/clopidogrel-treated patients, respectively). Despite this, 15% of clopidogrel good responders had a level of TFpos-platelets similar to the poor-responder group. Indeed, a stronger P2Y(12) inhibition (130-fold) is required to inhibit TF than VASP. Thus, a VASP platelet reactivity index <20% (as in prasugrel/ticagrelor-treated patients) identifies patients with TFpos-platelets <20% (92% sensitivity). Finally, colchicine impaired in vitro ADP-induced TF expression but not alpha-granule release, suggesting that TF is open canalicular system stored as confirmed by TEM and platelet analysis of patients with gray platelet syndrome.CONCLUSIONS: Data show that TF expression is regulated by P2Y(12) and not P2Y(1); P2Y(12) antagonists downregulate the percentage of TFpos-platelets. In clopidogrel good-responder patients, assessment of TFpos-platelets highlights those with residual platelet reactivity. TF is stored in open canalicular system, and its membrane exposure upon activation is prevented by colchicine