Switching Ratio from Parenteral to Oral Methadone 1:1.2 Is Safer Compared with Ratio 1:2 in Patients with Controlled Cancer Pain: A Multicenter Randomized-Controlled Trial (RATIOMTD-010810).

Background: The most commonly used switching ratio from parenteral to oral methadone is 1:2. Methadone is highly bioavailable and a lower ratio might result in similar analgesia with less toxicity. Objective: To compare success and side effects with two ratios from parenteral to oral methadone: 1:2 versus 1:1.2 in hospitalized patients with cancer pain. Design: A multicenter double-blind randomized clinical trial. Settings/Particiants: Inpatients with well-controlled cancer pain with parenteral methadone requiring rotation to the oral route. Measurements: Outcomes included pain intensity (Brief Inventory Pain), opioid toxicity (Common Toxicology Criteria for Adverse Events), and methadone dose. Success was defined as no toxicity with good pain control at 72 hours. Results: Thirty-nine of forty-four randomized patients were evaluable: 21 in ratio 1:2 and 18 in ratio 1:1.2. Seventy-one percent male. Median age 65 years. No significant differences in basal clinical characteristics between both groups. Median methadone dose pre/post switching was 24.5 mg ±13.5 and 49 mg ±27.3 for ratio 1:2, versus 23.3 mg ±9.4 (p: not significant) and 28 mg ±11.3 (p < 0.01) for ratio 1:1.2. Pain was well controlled without differences between both ratios. Drowsiness at day +1 (p < 0.017) and myoclonus at day +3 (p < 0.019) were more prevalent in group 1:2. Success was observed in 12 patients in ratio 1:2 versus 18 in ratio 1:1.2 (p < 0.001). Methadone side effects were observed in 12 patients in ratio 1:2 (mainly neurotoxicity symptoms) versus 2 in ratio 1:1.2 (p < 0.005). Conclusion: Ratio 1:1.2 when changing from parenteral to oral methadone resulted in lower toxicity and no difference in analgesia. More conservative dose adjustment during methadone route change should be considered. European Clinical Trials Register (EudraCT No. 2010-024092-39).

Propuesta de un instrumento para la detección del optimismo estratégico en cáncer de mama / Proposal for an instrument for the detection of strategic optimism in breast cancer

Objetivo: Elaborar un instrumento que nos permita evaluar algunas estrategias cognitivas que formarían parte de las personas optimistas ante una dificultad de salud como el cáncer de mama. Método: 96 pacientes, de ellas 30 se encontraban en una fase de intervalo libre del cáncer de mama, 35 recibiendo tratamiento activo para el cáncer de mama y 31 tenían un diagnóstico de trastorno adaptativo sin historial médico oncológico. Se han comparado los mecanismos cognitivos del instrumento para valorar el optimismo estratégico entre los grupos de salud; se ha efectuado el análisis factorial del instrumento y se ha observado la relación de éste con variables como la calidad de vida y el optimismo disposicional en cada grupo. Resultados: el análisis factorial muestra cuatro componentes del total de ítems del optimismo estratégico: 1) Expectativas de resultado positivo y percepción de control ante la dificultad de salud; 2) Valoración positiva del pasado en el momento actual de salud; 3) Benefit finding a raíz de la enfermedad; y 4 ) Responsabilidad/culpabilidad por la situación de salud. La varianza total explicada por los factores es del 70,67%. Los factores del optimismo estratégico estarían relacionados con una mejor calidad de vida de las pacientes (p <0,01; r= 0,591), mostrando relaciones variables en función de la gravedad de la situación médica. Conclusión: el instrumento aportado para evaluar el optimismo estratégico permite identificar estilos cognitivos, relacionados con el mantenimiento de la calidad de vida en cáncer de mama, que pueden ser de utilidad para potenciar las terapias psicológicas de apoyo

Objective: To elaborate an instrument that allows us to evaluate some cognitive strategies that would be part of the optimistic people facing a health difficulty such as breast cancer. Method: 96 patients, 30 of them were in a breast cancer free interval phase, 35 were receiving active treatment for breast cancer and 31 patients had a diagnosis of adaptive disorder with no oncological medical history. The cognitive mechanisms of the instrument have been compared to evaluate the strategic optimism among the three health conditions. The factorial analysis of the instrument has been carried out and its relationship with variables such as quality of life and dispositional optimism in each group has been observed. Results: the factor analysis shows four components of the total items of strategic optimism: 1) Expectations of positive results and perception of control in front of health difficulties; 2) Positive evaluation of the past in the current health moment; 3) Benefit finding in the wake of the disease; and 4) Responsibility / guilt for the current health situation. The total variance explained by the factors is 70.67%. The factors of strategic optimism would be related to a better quality of life of patients (p<0.01; r = 0.591), showing distinct relationships depending on the severity of the medical situation. Conclusion: The provided instrument to assess strategic optimism allows the identification of specific thoughts related to the maintenance of the quality of life in breast cancer, which may be useful in supportive therapies

[Coping strategies in women victims of gender violence and their relation with PTSD]. / Estrategias de afrontamiento en mujeres víctimas de violencia de género y su relación con el trastorno por estrés postraumático.

In our country, it is estimated that about 40% of the female population are physically and/or mentally abused by their partners, and between 21% and 60% of those women develop post-traumatic stress disorder. In this population, the knowledge about coping strategies is limited. OBJECTIVES: Identify whether signifcant differences exist as regards coping behaviors used in victims of gender violence between those women who develop post-traumatic stress disorder and those who do not. METHODS: The study involved 30 women who received an informed consent, a sociodemographic questionnaire, the questionnaire of the coping responses inventory CRI-A and the SCID-I. RESULTS: There were statistically signifcant differences regarding the use of logical analysis, seeking guidance and support, cognitive avoidance, and acceptance-resignation, used more by the post-traumatic stress disorder group; and minor use of positive reappraisal, problem solving and emotional discharge. DISCUSSION: Coping strategies of positive revaluation, problem solving and emotional discharge have been related to the decrease in the severity of depressive and anxious symptomatology. On the other hand, the usage of Cognitive Avoidance has been related to the increase in anxious and depressive symptomatology.

The in vitro interference of synthetic progestogens with carp steroidogenic enzymes.

Synthetic progestogens represent a class of pharmaceuticals widely used in oral contraceptives and in hormone replacement therapies. They reach the aquatic environment through wastewater effluents; however, environmental concentrations and effects on non-target organisms are poorly known. Given the important role of progestogens regulating fish spawning processes, this study aimed at assessing the in vitro interference of four currently used progestogens-drospirenone (DRO), levonorgestrel (LNG), norethindrone (NOR) and cyproterone acetate (CPA) - with key enzymatic activities involved in the synthesis of active steroids in carp (Cyprinus carpio). The enzymatic pathways investigated were (a) CYP17 (C17,20-lyase) and CYP11ß involved in the synthesis of androgens, (b) CYP19 that catalyses the aromatization of androgens to estrogens, and (c) 20ß-hydroxysteroid dehydrogenase (20ß-HSD) responsible for the synthesis of maturation-inducing hormones. All tested progestogens significantly inhibited the synthesis of androgens: DRO (IC50: 3.8 µM) was the strongest inhibitor of CYP17 followed by CPA (IC50s: 183 µM). Moreover, NOR (IC50: 0.4 µM), DRO (IC50: 1.8 µM) and CPA (IC50s: 87 µM) inhibited CYP11ß. An inhibition by NOR of ovarian CYP19 activity, and by DRO and CPA of 20ß-HSD was also observed, but at rather high concentrations (500 µM). Overall, this study highlights the potential of synthetic progestogens, and particularly DRO and NOR, to interfere with the biosynthesis of androgens in carp gonads.

Optimismo disposicional y calidad de vida en mujeres con cáncer de mama / Dispositional optimism and qualaity of life in women wih breast cancer

Objetivo: Analizar la posible relación entre el optimismo disposicional y la calidad de vida en una muestra de mujeres con cáncer de mama. Se trata de conocer qué aspectos de la calidad de vida estarían relacionados con una disposición optimista y cuáles no, y entender mejor el funcionamiento del optimismo para poder potenciarlo en aquellos casos en que sea conveniente para adaptarse mejor a la situación médica. Método: 50 mujeres con cáncer de mama fueron entrevistadas individualmente y completaron medidas de calidad de vida y optimismo disposicional. Resultados: El optimismo disposicional está significativamente relacionado con diversos aspectos de la calidad de vida. Las mujeres con cáncer de mama más optimistas sienten menos ira, menos dolor, manifiestan tener menos dificultades en sus movimientos, se sienten menos cansadas, duermen mejor, continuan con sus quehaceres diarios en casa, sienten más ilusión y menos miedo. Sin embargo, en algunos aspectos como las relaciones sociales y el cansancio, parece ser más importante no ser pesimista que ser optimista.Conclusión: En congruencia con estudios previos, el optimismo disposicional es un predictor de la preservación de la calidad de vida de las mujeres con cáncer de mama

Objective: To assess the relationship between dispositional optimism and quality of life in a sample of women with breast cancer. Our purpose is to know which aspects of quality of life are influenced by dispositional optimism and which are not, to improve the understanding of optimism and improve it when it’s necessary for the adaptation to the medical situation. Method: 50 women with breast cancer answered individually quality of life and dispositional optimism measures. Results: Dispositional optimism is significantly related to many aspects of quality of life. Women with breast cancer with high dispositional optimistic feel less anger, less pain, have less difficulties in their movements, feel less tiredness, sleep better, go on with theirs home tasks, feel more illusion and less fear. But in some aspects, like social function or fatigue, it seems more important not being pessimist than being optimistic. Conclusion: In congruence with previous reports, dispositional optimism is a predictor of quality of life preservation in women with breast cancer

Characterization of the environmental quality of sediments from two estuarine systems based on different in-vitro bioassays.

This study characterizes the environmental quality of sediments from the Arade and Guadiana estuaries using different in-vitro bioassays: a) fish hepatoma cell line (PLHC-1) to determine cytotoxicity and presence of CYP1A and oxidative stress inducing agents; b) gonad subcellular fractions from sea bass (Dicentrarchus labrax) to detect compounds that are likely to act as endocrine disrupters by interfering with the synthesis of androgens (CYP17, CYP11ß) and estrogens (CYP19). Approximately 60% of extracts from the Arade estuary were cytotoxic when tested at 60 mg eQsed/mL, while only one sample from Guadiana showed cytotoxicity. Sediments from Arade collected close to harbours and waste water effluents were enriched with CYP1A inducing agents, while those from the upper Guadiana induced oxidative stress in PLHC-1 cells. On the other hand, several extracts from both estuaries were able to significantly inhibit CYP17, CYP11ß and CYP19 activities in gonad subcellular fractions of sea bass, which indicates the presence of endocrine disrupters, particularly in several sites from the Arade estuary. Overall, the study highlights the usefulness of in-vitro bioassays to identify those sediments that could pose risk to aquatic organisms and that require further action to improve their environmental quality.

Factor v leiden and inflammation.

Factor V Leiden, is a variant of human factor V (FV), also known as proaccelerin, which leads to a hypercoagulable state. Along these years, factor V Leiden (FVL) has been studied from the pathophysiologic point of view, and research has been focused on finding clinical approaches for the management of the FVL associated to a trombophilic state. Less attention has been paid about the possible role of FVL in inflammatory conditions known to be present in different disorders such as uremia, cirrhosis, liver transplantation, depression as well as sepsis, infection or, inflammatory bowel disease (IBD). Whether platelet FVL will increase the activation of coagulation and/or in which proportion is able to determine the final outcome in the previously mentioned inflammatory conditions is a subject that remains uncertain. This paper will review the association of FVL with inflammation. Specifically, it will analyze the important role of the endothelium and the contribution of other inflammatory components involved at both the immune and vascular levels. This paper will also try to emphasize the importance of being a FVL carrier in associations to diseases where a chronic inflammation occurs, and how this condition may be determinant in the progression and outcome of a specific clinic situation.

External calcium facilitates signalling, contractile and secretory mechanisms induced after activation of platelets by collagen.

Platelet activation leads to the initiation of intracellular signalling processes, many of which are triggered by Ca2+. We have studied the involvement of exogenous Ca2+ in platelet response to collagen activation. Platelet suspensions were prepared with and without adding external calcium in the suspension buffers. Activation with collagen (Col-I) was carried out, before and after incubation with cytochalasin B (Cyt-B) to block the actin assembly and the cytoskeletal reorganization. We evaluated changes in (i) tyrosine phosphorylation of proteins, in platelet lysates and associated with the cytoskeletal fraction, (ii) the association of contractile proteins to the cytoskeleton, (iii) expression of intraplatelet substances at the surface, and (iv) cytosolic Ca2+ levels ([Ca2+]i). Ultrastructural evaluation of platelets by electron microscopy was also performed. Platelet activation by Col-I in the absence of added Ca2+ was followed by mild association of actin and other contractile proteins, low phosphorylation of proteins at tyrosine residues, lack of expression of intraplatelet substances at the membrane, and absence of aggregation. In the presence of millimolar Ca2+, Col-I induced intense actin filament formation with association of contractile proteins with the cytoskeleton, resulting in profound morphological changes. Under these conditions, Col-I induced signalling through tyrosine phosphorylation, with increases in the [Ca2+]i, release of intragranule content and aggregation. Inhibiting actin polymerization with Cyt-B prevented all these events. Our data indicates that platelet activation by collagen requires external Ca2+. Studies with Cyt-B indicate that assembly of new actin and cytoskeleton-mediated contraction, both dependent on exogenous Ca2+, are key events for platelet activation by collagen. In addition, our results confirm that entrance of exogenous Ca2+ depends on a functional cytoskeleton.

Inhibition of tyrosine kinase activity prevents the adhesive and cohesive properties of platelets and the expression of procoagulant activity in response to collagen.

INTRODUCTION: Platelet activation leads to signal transduction mechanisms, in which phosphotyrosine proteins play a relevant role. MATERIAL AND METHODS: Platelet suspensions were independently activated by collagen and thrombin in the absence and in the presence of two tyrosine kinase inhibitors, tyrphostin 47 and genistein. Samples were processed to visualize morphological changes by electron microscopy, to evaluate changes in cytoskeletal assembly, to analyze modifications in the expression of activation dependent antigens, and the procoagulant activity at the surface level by flow cytometry. Additional experiments applying flow conditions were performed to assess the effect of inhibiting tyrosine phosphorylation on primary platelet adhesion and fibrin formation. RESULTS: Inhibition of tyrosine phosphorylation blocked shape change and cytoskeletal assembly induced by collagen, and inhibited, though partially, those effects due to thrombin. Both activating agents induced the expression of the intraplatelet antigens CD62P and CD63 at the surface, although only collagen promoted expression of anionic phospholipids. Both tyrphostin 47 and genistein prevented those effects. The extent of platelet adhesion on both collagen-coated and subendothelial surfaces was significantly diminished by the presence of the tyrosine kinase inhibitors assayed. Fibrin formation was also significantly reduced. CONCLUSIONS: Platelet shape change and secretion during platelet activation depends on tyrosine phosphorylation. In addition, primary adhesion of platelets induces signaling through tyrosine kinases to achieve full spreading, and results in the exposure of a procoagulant surface on platelets.

Platelet microparticles are heterogeneous and highly dependent on the activation mechanism: studies using a new digital flow cytometer.

BACKGROUND: Platelet-derived microparticles (MPs) are believed to play an important role in coagulation and inflammatory disorders. Unfortunately, MP size renders them difficult to study and analyze by conventional flow cytometry. METHODS: We analyzed and characterized platelet-derived MPs, using antibodies against the major surface glycoproteins (GP), the platelet activation antigen P-selectin (CD62P), and a marker of procoagulant activity (phosphatidylserine exposure). MPs were generated by exposure of platelets to thrombin receptor activating peptide (TRAP) or ionophore. Both agonists induced significant microvesiculation of platelets, and the resulting MPs were analyzed by a new digital flow cytometer: Becton-Dickinson FACSAria. RESULTS: Membrane GPs were equally well represented in MPs generated by either reagent. In contrast, P-selectin was more intensely expressed in TRAP-MPs, while phosphatidylserine (PS) expression was markedly increased in ionophore-MPs. Two distinct populations of TRAP-MPs (one PS-positive and another PS-negative) were apparent. The latter characteristic facilitated sorting of MPs according to their PS exposure. CONCLUSIONS: The data presented herein show a significant improvement in the methodology applied until now to the characterization of MPs. The ability to characterize and sort MP subpopulations may help to resolve their contributions to normal and pathological functions.

Effects of etamsylate on equine platelets: in vitro and in vivo studies.

The aim of this study was to investigate whether etamsylate produces equine platelet activation. In vitro and in vivo studies were designed in which seven and eight adult healthy horses were included, respectively. In the in vitro study, citrated blood was incubated with different concentrations of etamsylate, and P-selectin expression and annexin V binding were determined by flow cytometry. In the in vivo study, blood was collected before and 1 and 2h after IV administration of etamsylate, and P-selectin expression was evaluated. In the in vitro study, a significant increase in P-selectin expression, leukocyte-platelet aggregate formation and annexin V binding were observed. In the in vivo study, a marked increase in P-selectin expression and heterotypic aggregate formation was seen in two and five horses, respectively, although no significant differences were detected when analyzing results from all the animals together. The results of the in vitro study indicate that etamsylate produces a pre-activation state in equine platelets, but this fact could be confirmed by the in vivo study.

Assessment of platelet function in horses: ultrastructure, flow cytometry, and perfusion techniques.

We studied equine platelet function and activation using ultrastructural examination, flow cytometry, and perfusion. The main aim of the study was to evaluate hemostatic mechanisms in horses using these techniques. Ultrastructural observations were done on resting and activated platelets. Flow cytometry was used to evaluate binding of antibodies to major platelet glycoproteins (GPIIb-IIIa, GPIV, and GPIb) and activation-dependent antigens (P-selectin and lysosomal integral membrane protein [LIMP]). Perfusion techniques were used to evaluate the interaction between platelets and damaged subendothelium. Aggregation experiments were done to identify the best agonists for flow cytometry. Ultrastructural observations confirmed that equine platelets lack a developed open canalicular system and that release of granule contents occurs by fusion of adjacent granule membranes that ultimately connect with external membranes. Flow cytometry identified a 2-fold increase in binding of antibodies against GPIIb-IIIa and GPIV after activation. Binding of antibodies against P-selectin and LIMP increased from 2.12 and 1.74% to 15.5 and 11.6%, respectively, in response to thrombin and to 21.86 and 10.50%, respectively, in response to collagen. Annexin V binding increased moderately after activation. Perfusion experiments with citrated blood indicated that equine platelets react more strongly to subendothelium than do human platelets. When blood was anticoagulated with low molecular weight heparin, a marked impairment of platelet interactions was observed. In conclusion, although some differences were observed between human and equine platelet function, some techniques currently used to assess human platelet function may be useful to assess equine platelets.

Stored platelets contain residual amounts of tissue factor: evidence from studies on platelet concentrates stored for prolonged periods.

BACKGROUND: The advent of new strategies for pathogen reduction has raised the question of whether platelet (PLT) concentrates (PCs) exposed to longer periods of storage retain adequate hemostatic function. STUDY DESIGN AND METHODS: The effects of prolonged storage on adhesive and procoagulant functions of PLTs in PCs have been analyzed. The ability of PLTs to interact with surfaces was assessed by en face electron microscopy. Exposure of anionic phospholipids or tissue factor (TF) antigen on PLTs was assessed by flow cytometry and by immunocytochemical methods at the ultrastructural level. Studies were performed in six different PCs followed 0, 3, 5, 7, and 11 days of storage. RESULTS: A progressive impairment of PLT-adhesive functions was observed after 5 days of storage. A progressive increase in expression of anionic phospholipids and development of procoagulant activity (PCA) measured by a modified prothrombin time (mPT) was observed along the storage. Incubation of PLTs with a specific anti-TF resulted in prolongation of the mPT by approximately 10 to 15 percent. Flow cytometry revealed minimal TF expression at later storage times. Immunocytochemical studies showed minimal TF labeling when studies were performed on PLT whole mounts. Labeling was markedly improved when PLTs were previously exposed to sonication. CONCLUSION: Prolonged storage of PCs was associated with decreased PLT-adhesive capacities and enhanced PCA. Current preparation procedures and storage media have important limitations for preserving PCs for longer than 1 week. PLTs in PCs retain residual amounts of TF as assessed by immunocytochemical and functional assays. The origin and hemostatic significance of this TF should be investigated further.

Effects of Helicobacter pylori eradication on platelet activation and disease recurrence in patients with acute coronary syndromes.

BACKGROUND: Platelet activation is consistently observed in animal models of Helicobacter pylori infection and could help to explain the alleged epidemiological association between H. pylori and coronary heart disease. MATERIALS AND METHODS: Ninety-two patients with recent acute coronary syndromes were enrolled. Helicobacter pylori-positive patients were randomized to receive a 7-day course of omeprazole, amoxycillin and metronidazole or placebos. Two months later, H. pylori status was reassessed and baseline parameters, including soluble P-selectin and platelet surface expression of CD62P, CD63 and CD41, were measured again. Patients were followed-up for 1 year or until death or readmission. RESULTS: No baseline differences were observed between H. pylori-positive and -negative cases. Among H. pylori-positive patients, 18 received placebo and 31 received active medication resulting in eradication in 21 cases. No differences were observed in inflammatory parameters or platelet activation markers between patients with persistent or resolved H. pylori infection. However, coronary events recurred at 6 and 12 months, respectively, in 35% and 55% of patients with persisting H. pylori infection compared with 10% and 25% of patients in whom H. pylori was either absent or eradicated (p = .01). Only final H. pylori status [RR 3.07 (95% CI 1.35-98)] and number of coronary risk factors [RR 2.58 (95% CI 1.51-4.41)] were independent predictors of recurrence. CONCLUSIONS: Infection with H. pylori does not induce significant platelet activation in patients treated for coronary disease. Helicobacter pylori-infected patients, however, may have an increased risk of recurrence of coronary events.

Effect of holding buffy coats 4 or 18 hours before preparing pooled filtered PLT concentrates in plasma.

BACKGROUND: Filtered PLT concentrates (PCs) were prepared in plasma pooling three (for children) or six buffy coats (BCs; for adults) after holding them a maximum of 4 hours (blood bags collected in the afternoon) or 18 hours (blood bags collected in the morning). STUDY DESIGN AND METHODS: With flow cytometry, PCs prepared after holding BCs 4 or 18 hours were compared. BCs removed from whole-blood donations in quadruple bag packs ("top-top") were held 4 or 18 hours before pooling them with a sterile connecting device. After the BCs were centrifuged, the supernatant was transferred through a BC filter (Autostop, Pall Medical) to a CLX bag. Samples for analysis were collected from the whole-blood bag, BCs, and PCs immediately after preparation and after 1, 3, 5, and 7 days of storage on a flat-bed agitator at 22 +/- 2 degrees C. The main PLT membrane glycoproteins (GPs, IIb-IIIa, IV, and Ibalpha), some of their ligands (fibrinogen, fibronectin, and VWF), activation-dependent antigens (CD62P and CD63), and procoagulant activity markers (annexin V and bound coagulation FV-Va) have been studied. RESULTS: In the 12 PCs (six pools of 3 units each group) studied, a minor increase in activation markers during preparation was observed. During the storage, a significant increase in the expression of GPIIb-IIIa, CD62P, CD63, annexin V, and FVa was measured. After 5 days of storage, only the percentage of PLTs with bound fibrinogen was significantly greater in PCs prepared after holding BCs for 4 hours. CONCLUSION: In PCs prepared after holding BCs 4 or 18 hours before pooling and filtering, only a minor significant difference in the percentage of PLTs with bound fibrinogen was found after 5 days of storage. This difference is probably of little, if any, transfusional significance.

Glycoproteins expression on platelet membrane in inherited macrothrombocytopenias.

INTRODUCTION: Inherited giant platelet disorders (IGPD) are a heterogeneous group of rare diseases characterized by thrombocytopenia, large platelets and variable bleeding symptoms. Glycoprotein (GP) expression on platelet surface in these conditions is poorly characterized. We have investigated the expression of constitutively expressed platelet membrane GP and the response to TRAP activation in a group of patients with different forms of inherited macrothrombocytopenias. MATERIALS AND METHODS: Two patients diagnosed Epstein syndrome (ES), two with Bernard-Soulier syndrome (BSS) and eight with Mediterranean macrothrombocytopenia (MM) were studied using flow cytometry combined to monoclonal antibodies (MoAbs). Mean platelet volume was also measured. RESULTS: In general, there was an increase in the binding of MoAbs directed against platelet membrane GPs in the patients studied (except, obviously, in BSS patients). The observed increase ranged between 0.9 and 3.36 times the value of the control for GPIbalpha, between 1.36 and 7.2 times for GPIIb-IIIa and 0.94 and 8.07 times for GPIV. However, when the observed value was divided by the measured platelet volume, the estimated density was similar to controls in the case of GPIbalpha but were increased for GPIIb-IIIa and GPIV. TRAP activation provoked significant increments in the number of copies of GPIIb-IIIa complexes present on platelet surface in controls (91%), MM patients (49%), but almost no changes in BSS platelets (6%). CONCLUSION: In the group of macrothrombocytopenia studied, an increase in the expression of platelet membrane GP was observed, although a wide variability exists even in patients with the same disorder.

Inhibition of cytoskeletal assembly by cytochalasin B prevents signaling through tyrosine phosphorylation and secretion triggered by collagen but not by thrombin.

Activation of platelets leads to cytoskeletal assembly that is responsible for platelet motility and internal contraction. We have evaluated the involvement of the cytoskeleton in platelet activation by two strong agonists, collagen and thrombin. Activation was assessed by measuring changes in cytoskeletal assembly, externalization of activation-dependent markers and expression of procoagulant activity, and tyrosine phosphorylation of proteins, in both the absence and the presence of cytochalasin B. Activation of platelets with collagen and thrombin induced morphological changes and increased the expression of CD62P, CD63, glycoprotein IV, and binding of annexin V to platelets. Moreover, both activating agents induced actin polymerization, increased the association of other contractile proteins, and promoted tyrosine phosphorylation of multiple proteins, some of which were associated with the cytoskeleton. The presence of cytochalasin B blocked the previous events when collagen was used as the activating agent, although binding of annexin V still occurred. In contrast, platelet response to thrombin was not completely prevented by the presence of cytochalasin B. Thus, activation by collagen requires a functional cytoskeleton to trigger signaling through tyrosine phosphorylation and secretion. This is not the case for thrombin, which is capable of activating signaling mechanisms in the presence of strong inhibitors of cytoskeletal assembly. Moreover, the expression of a procoagulant surface in platelets still occurs even when platelet motility has been inhibited.