Browsing by Author "Ocqueteau, M"

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    Circulating platelet-derived microparticles in systemic lupus erythematosus - Association with increased thrombin generation and procoagulant state
    (GEORG THIEME VERLAG KG, 2006) Pereira, J; Alfaro, G; Goycoolea, M; Quiroga, T; Ocqueteau, M; Massardo, L; Perez, C; Saez, C; Panes, O; Matus, V; Mezzano, D
    The risk for thrombosis is significantly increased in systemic lupus erythematosus (SLE), affecting both venous and arterial vessels. Activated platelets are known to participate in thrombus formation and growth. A general feature of activated cells is the shedding of microparticles (MP) which support coagulation by exposure of negatively charged phospholipids and possibly tissue factor (TF). In this work we characterized circulating MP in patients with SLE and their relationship with a procoagulant state. Thirty patients with SLE (aged 15-72 years, mean age 38 years) and 20 healthy controls (aged 22-54 years, mean age 34 years) were studied; patients fulfilled 4 revised criteria for SLE. The number and cellular source of circulating MP were determined by flow cytometry using double labeling with specific monoclonal antibodies and annexin V. Thrombin generation was measured as the endogenous thrombin potential (ETP) without the addition of exogenous phospholipids and TF; under these conditions the generation of thrombin depended directly on the number of MP present in plasma. Thrombin anti-thrombin (TAT) and plasmin-antiplasmin (PAP) complexes were measured by ELISA. Compared to the controls, circulating MP were significantly elevated in the patient group (1218 +/- 136 vs 653 +/- 74 x 10(3)/ml plasma, p: 0.0007). In both groups, most of these MP were of platelet origin (927 +/- 131 vs 517 +/- 72 x 10(3)/ml plasma, p:0.009). ETP was higher among patients as compared to the controls (804 +/- 64 vs 631 +/- 37 nM thrombin, p: 0.025). Plasma levels of TAT in patients and controls were 3.4 +/- 0.8 and 1.4 +/- 0.5 mu g/L, respectively (p:0.04), and of PAP complexes were 62.5 +/- 14 and 24.05 +/- 2.5 mu g/ml, respectively (p:0.014). The number of platelet-derived MP correlated significantly with thrombin generation (r: 0.42; p: 0.038) and TAT levels (r: 0.40; p: 0.035). We did not find an association of circulating MP with disease activity nor with the presence of antiphospholipid antibodies. The increased number of circulating platelet-derived microparticles and their association with high ETP and activation of the coagulation system suggest that these microparticles play an important role in the pathogenesis of the prothrombotic state in SLE patients.
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    Glycoprotein Ib/IX complex is the target in rifampicin-induced immune thrombocytopenia
    (BLACKWELL SCIENCE LTD, 2000) Pereira, J; Hidalgo, P; Ocqueteau, M; Blacutt, M; Marchesse, M; Nien, Y; Letelier, L; Mezzano, D
    Thrombocytopenia is a major adverse effect of several drug treatments. Rifampicin has been recognized as a cause of immune thrombocytopenia during intermittent high-dose therapy. We characterized the antibody of a patient who presented with purpura and thrombocytopenia during treatment of tuberculosis with rifampicin. Drug-dependent binding of the antibody to platelets was demonstrated by flow cytometry. In a glycoprotein-specific immunoassay, the binding epitope of the IgG antibody was found in the glycoprotein Ib/IX complex, using four different monoclonal antibodies (mAbs) against various epitopes on the GPIb/IX complex, as well as mAbs against GPIIb/IIIa, GPIa/IIa and GPIV. By immunoprecipitation of biotin-labelled platelets, reactivity of the antibody with GPIb/IX was found only in the presence of the drug. These findings clearly demonstrate that rifampicin induces the formation of drug-dependent antibodies capable of causing thrombocytopenia. The binding site of the rifampicin-dependent antibody, located in the GPIb/IX complex, seems to be a favoured target for antibodies induced by different drugs.
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    Natural killer cell leukemia. Case report
    (SOC MEDICA SANTIAGO, 2005) Guerrero, M; Lira, P; Bertin, P; Galleguillos, M; Ocqueteau, M
    Natural killer leukemia is a rare and highly aggressive neoplasm, is more common in young male patients and has a very poor prognosis, with a median survival of few weeks. We report a 17 years old male patient who developed, after an acute upper respiratory disease, a rapidly multiorganic failure with pancytopenia. Bone marrow aspiration and trephine biopsy showed an acute lymphoblastic leukemia. The immunophenotypic and immunohistochemistry revealed a natural killer acute leukemia. The disease progressed rapidly and the patient died shortly after the diagnosis.
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    Platelet aging in vivo is associated with activation of apoptotic pathways: Studies in a model of suppressed thrombopoiesis in dogs
    (SCHATTAUER GMBH-VERLAG MEDIZIN NATURWISSENSCHAFTEN, 2002) Pereira, J; Soto, M; Palomo, I; Ocqueteau, M; Coetzee, LM; Astudillo, S; Aranda, E; Mezzano, D
    The mechanism(s) involved in the clearance of senescent platelets are largely unknown. We have recently demonstrated that platelet aging in vivo is associated with loss of membrane phospholipid asymmetry, a universal phenomenon in cells undergoing apoptosis. Thus, We postulated that senescent platelets may exhibit programmed cell death changes, which may trigger their removal from circulation. Since platelets contain the apoptosis machinery as well as mitochondria, a key organelle in the regulation of apoptosis, we studied the appearance of apoptotic-like changes during platelet aging in vivo. To investigate this, we assessed changes in mitochondrial membrane potential in circulating canine platelets during decline in platelet Count after suppression of thrombopoiesis by estradiol injection. a validated model to obtain circulating platelets of increasing mean ace. Phosphatidyl-serine (PS) exposure was determined by flow cytometry by binding of FITC-labeled annexin V. Mitochondrial Deltapsi was studied with the cationic lipophilic dye DIOC6 (3) and the J-aggregate-forming cation JC-1 and analysis by flow cytometry. The proportion of platelets with exposed PS rose significantly with age, from 2.88% before to 6.7%. 8 days after estradiol injection. By flow, cytometry it was demonstrated a significant decreased in DIOC6 (3) fluorescence (median fluorescence intensity 791 98 vs 567 1021 day 0 vs day 8 post injection of estradiol, respectively n:11; p<0.01), consistent with mitochondrial &UDelta;ψ collapse. JC-1 has the unique property of forming J-aggregates tinder high mitochondrial &UDelta;ψ (red fluorescence, FL2) whereas the monomeric form fluoresces in green (FL1). Aged platelets in vivo, loaded with JC-1, exhibited a significant increase in FL1/FL2 ratio (2.5&PLUSMN;1.7 vs 4.7&PLUSMN;1.6, day 0 vs day 8 post injection of estradiol, respectively n:13; p<0.05). confirming the mitochondrial Deltapsi alteration.
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    Platelet aging in vivo is associated with loss of membrane phospholipid asymmetry
    (F K SCHATTAUER VERLAG GMBH, 1999) Pereira, J; Palomo, I; Ocqueteau, M; Soto, M; Aranda, E; Mezzano, D
    The mechanism(s) involved in the clearance of senescent platelets are largely unknown. The loss of membrane phospholipid (PL) asymmetry, with phosphatidylserine (PS) exposure appears to be an important signal for the ingestion by macrophages of apoptotic nucleated cells and it has also been suggested as a signal for the removal of aged erythrocytes. Accordingly, it seems possible that the clearance of normal aged platelets from circulation might be triggered by PS exposure. To investigate this, we determined PS exposure in human aging platelets taking advantage of the relationship between platelet density and platelet age and in dog platelets in a model of platelet aging in vivo. PS exposure was determined in two experimental conditions: 1) human platelet density subpopulations obtained by centrifugation in arabinogalactan gradients; 2) circulating canine platelets during decline in platelet count after suppression of thrombopoiesis following estradiol injection. PS exposure was determined by flow cytometry after labeling the cells with FITC-conjugated annexin V. The proportion of human platelets with exposed PS was significantly higher in high density (HD) platelets compared to low density (LD) platelets (11.3 +/- 8.0% vs 5.2 +/- 3.7%; p < 0.05, respectively). In dogs, the proportion of cells with exposed PS rises dramatically with age, from 3.1 +/- 0.4% before to 17.7 +/- 12.3% ten days after estradiol injection. These findings suggest that platelet aging is associated with loss of phospholipid asymmetry and PS exposure on the outer leaflet of cell membrane, which may play an important role in the recognition and subsequent removal of senescent platelets.