Deficiency of platelet adhesion molecule CD226 causes megakaryocyte development and platelet hyperactivity.

This study used constitutive CD226 gene knockout (KO) mice as a model to investigate the functions and mechanisms of CD226 in megakaryocyte (MK) maturation and platelet activation. Although CD226 deficiency did not cause MK polyploidization or platelet granule abnormalities, increased MK counts were detected in the femora bone marrow (BM) and spleen of CD226 KO mice. Particularly, CD226 KO mice have a more extensive membrane system in MKs and platelets than wild-type (WT) mice. We also demonstrated that CD226 KO mice displayed increased platelet counts, shortened bleeding time, and enhanced platelet aggregation. CD226 KO platelets had an increased mature platelet ratio compared to the control platelets. In addition, the observed reduction in bleeding time may be due to decreased nitric oxide (NO) production in the platelets. Platelet-specific CD226-deficient mice showed similar increased MK counts, shortened bleeding time, enhanced platelet aggregation, and decreased NO production in platelets. Furthermore, we performed middle cerebral artery occlusion-reperfusion surgery on WT and CD226 KO mice to explore the potential effect of CD226 on acute ischemia-reperfusion injury; the results revealed that CD226 deficiency led to significantly increased infarct area. Thus, CD226 is a promising candidate for the treatment of thrombotic disorders.

A combined effect of anti-HPA-1a and anti-HLA Class I in pregnancy?

BACKGROUND: Maternal anti-human leukocyte antigen (HLA) Class I is commonly detected alongside anti-human platelet antigen (HPA)-1a in fetal and neonatal alloimmune thrombocytopenia (FNAIT). Little is known regarding whether the presence of anti-HLA Class I may exert an additive effect on the risk and severity of FNAIT. METHODS AND MATERIALS: We reanalyzed samples originally collected as part of a large Norwegian screening study on FNAIT during 1995-2004. This study identified and managed 170 pregnancies where the mother was HPA-1a negative and had detectable anti-HPA-1a during pregnancy. Maternal samples from 166 of these pregnancies were rescreened for anti-HLA Class I, revealing 111 (67%) that were antibody positive. Various regression models were used to assess if and how maternal anti-HLA Class I influenced the neonatal platelet count. RESULTS AND CONCLUSIONS: Unadjusted neonatal platelet counts and the frequency of neonatal thrombocytopenia was not significantly affected by the presence of anti-HLA Class I alongside anti-HPA-1a, but results from regression analyses revealed a possible increased risk when the mother was nulliparous. These results warrant further investigation.

Gain-of-Function Mutation in Filamin A Potentiates Platelet Integrin αIIbß3 Activation.

OBJECTIVE: Dominant mutations of the X-linked filamin A (FLNA) gene are responsible for filaminopathies A, which are rare disorders including brain periventricular nodular heterotopia, congenital intestinal pseudo-obstruction, cardiac valves or skeleton malformations, and often macrothrombocytopenia. APPROACH AND RESULTS: We studied a male patient with periventricular nodular heterotopia and congenital intestinal pseudo-obstruction, his unique X-linked FLNA allele carrying a stop codon mutation resulting in a 100-amino acid-long FLNa C-terminal extension (NP_001447.2: p.Ter2648SerextTer101). Platelet counts were normal, with few enlarged platelets. FLNa was detectable in all platelets but at 30% of control levels. Surprisingly, all platelet functions were significantly upregulated, including platelet aggregation and secretion, as induced by ADP, collagen, or von Willebrand factor in the presence of ristocetin, as well as thrombus formation in blood flow on a collagen or on a von Willebrand factor matrix. Most importantly, patient platelets stimulated with ADP exhibited a marked increase in αIIbß3 integrin activation and a parallel increase in talin recruitment to ß3, contrasting with normal Rap1 activation. These results are consistent with the mutant FLNa affecting the last step of αIIbß3 activation. Overexpression of mutant FLNa in the HEL megakaryocytic cell line correlated with an increase (compared with wild-type FLNa) in PMA-induced fibrinogen binding to and in talin and kindlin-3 recruitment by αIIbß3. CONCLUSIONS: Altogether, our results are consistent with a less binding of mutant FLNa to ß3 and the facilitated recruitment of talin by ß3 on platelet stimulation, explaining the increased αIIbß3 activation and the ensuing gain-of-platelet functions.

Evaluation of platelet surface glycoproteins in patients with Glanzmann thrombasthenia: Association with bleeding symptoms.

BACKGROUND & OBJECTIVES: Glanzmann thrombasthenia (GT) is a rare, inherited autosomal recessive disorder characterized by qualitative or quantitative deficiency of integrin αIIbß3 [glycoprotein IIb (GPIIb)/IIIa, CD41/CD61] diagnosed by absent or reduced platelet aggregation to physiological agonists, namely, collagen, adenosine-di-phosphate, epinephrine and arachidonic acid. The objective of this study was to quantitate platelet surface GPs, classify GT patients and relate the results with the severity of bleeding and platelet aggregation studies. METHODS: Fifty one patients of GT diagnosed by platelet aggregation studies were evaluated for the expression of CD41, CD61, CD42a and CD42b on platelet surface by flow cytometry. The association between the clinical phenotype based on bleeding score and GT subtype on flow cytometric evaluation was assessed. RESULTS: Twenty four (47%) patients of GT were classified as type I (as CD41/CD61 were virtually absent, <5%), six (11.8%) patients as type II (5-20% CD41/CD61) and 21 (41.2%) as type III or GT variants as they had near normal levels of CD41 and CD61. Type III GT patients had significantly lower numbers of severe bleeders (P=0.034), but the severity of bleeding did not vary significantly in type I and II GT patients. In all GT patients, mean CD41 expression was found to be lower than mean CD61 expression (P=0.002). INTERPRETATION & CONCLUSIONS: Type I GT was found most common in our patients and with lowered mean CD41 expression in comparison with CD61. Type III GT patients had significantly lower numbers of severe bleeders, but the severity of bleeding did not vary significantly in type I and II GT patients.

Associations of combined polymorphisms of the platelet membrane glycoproteins Ia and IIIa and the platelet-endothelial cell adhesion molecule-1 and P-Selectin genes with IVF implantation failures.

The aim of the study was to investigate the combined impact of the genetic heterogeneity of the glycoproteins Ia (GpIa) and IIIa (GpIIIa) and the platelet-endothelial cell adhesion molecule-1 (PECAM-1) and P-Selectin genes on IVF embryo transfer implantation failures (IVF-ET failures). Sixty nulligravida women with previous IVF-ET failures and 60 fertile controls were genotyped for the GpIa-C807T, GpIIIa-PlA1/PA2, PECAM-1-C373G (Leu125Val) and P-Selectin-A37674C (Thr715Pro) polymorphisms by pyrosequencing. Compared with wild-type combined homozygotes, carriers of combinations of risk alleles in two gene loci were at significantly increased risk for IVF-ET failure, whereas carriers of the combination of GpIa-807T, GpIIIa-PlA2 and PECAM-1-373G alleles had OR = 52.50 (95%CI: 4.05-680.95, p < .001). The area under the receiver-operating characteristic curve (AUC) based on the number of polymorphisms and the number of risk alleles per subject was 75.4% (95%CI: 66.7%-82.8%, p < .001) and 72.5% (95%CI: 63.6%-80.3%, p < .001), respectively. The OR per polymorphism and risk allele increase was 4.26 (95%CI: 2.15-8.41, p < .001) and 2.85 (95%CI: 1.71-4.76, p < .001), respectively. The above associations were more robust among younger women. The combined analysis of these polymorphisms revealed strong association of combined carriers with IVF-ET failures especially for younger women and provided a genetic risk score with good diagnostic accuracy in the prediction of IVF-ET failures.

[Modern methods of diagnosis of thrombophylic states and complex treatment of patients with thrombotic complications of severe forms of varicose disease].

Actual issues of surgical treatment of patients, suffering complications of severe forms of varicose disease of the lower extremities (VDLE) are discussed. The causes of unsatisfactory results of treatment in patients, suffering varicothrombophlebitis (VTHPH), the main of which--absence of the only one tactics for operative treatment and anticoagulant therapy, were analyzed. The results of patients examination, suffering thrombotic complications of severe forms of VDLE, while its recurrent course, in conjunction of VTHPH and thrombosis of deep veins of the lower extremities, using diagnostic complex "PLR genetics thrombophilia", are adduced. Differential tactics of treatment in patients, suffering severe forms of VDLE, while various localization of thrombotic process, concerning the presence of thrombophilic states, is proposed.

[Thrombotic complications of severe forms of varicose disease: modern approach to diagnosis and treatment of hereditary thrombophilia and immunohistochemical peculiarities of vascular wall].

Experience of treatment of 176 patients, suffering thrombotic complications of severe forms of the lower extremities varicose disease (VDLE), was analyzed. In 20 patients, suffering varicothrombophlebitis (VTHPH) in severe forms of VDLE, morphological and immunohistochemical changes in the venous wall and surrounding tissues were studied. There were examined 28 patients, in whom thrombotic complications of the VDLE have had occurred, using diagnostic complex "PLR genetics thrombophilia". Recurrent course of thrombotic complications and coexistence of VTHPH and thrombosis of deep veins have had constitute the main criterion of such patients selection. The groups of patients, suffering severe forms of VDLE, were delineated, depending on thrombotic process localization, differentiated tactics of their surgical treatment was proposed.

FlnA-null megakaryocytes prematurely release large and fragile platelets that circulate poorly.

Filamin A (FlnA) is a large cytoplasmic protein that crosslinks actin filaments and anchors membrane receptors and signaling intermediates. FlnA(loxP) PF4-Cre mice that lack FlnA in the megakaryocyte (MK) lineage have a severe macrothrombocytopenia because of accelerated platelet clearance. Macrophage ablation by injection of clodronate-encapsulated liposomes increases blood platelet counts in FlnA(loxP) PF4-Cre mice and reveals the desintegration of FlnA-null platelets into microvesicles, a process that occurs spontaneously during storage. FlnA(loxP) PF4-Cre bone marrows and spleens have a 2.5- to 5-fold increase in MK numbers, indicating increased thrombopoiesis in vivo. Analysis of platelet production in vitro reveals that FlnA-null MKs prematurely convert their cytoplasm into large CD61(+) platelet-sized particles, reminiscent of the large platelets observed in vivo. FlnA stabilizes the platelet von Willebrand factor receptor, as surface expression of von Willebrand factor receptor components is normal on FlnA-null MKs but decreased on FlnA-null platelets. Further, FlnA-null platelets contain multiple GPIbα degradation products and have increased expression of the ADAM17 and MMP9 metalloproteinases. Together, the findings indicate that FlnA-null MKs prematurely release large and fragile platelets that are removed rapidly from the circulation by macrophages.

Preserved thrombin-inducible platelet activation in thienopyridine-treated patients.

BACKGROUND: Abundant thrombin generation may be a major reason for subsequent thromboembolic events in patients with cardiovascular disease receiving dual antiplatelet therapy. We therefore investigated the susceptibility of thienopyridine responders and nonresponders to thrombin receptor-activating peptide (TRAP)-6- and adenosine diphosphate (ADP)-inducible platelet activation. MATERIALS AND METHODS: Response to clopidogrel or prasugrel was determined by the vasodilator-stimulated phosphoprotein (VASP) phosphorylation assay and multiple electrode aggregometry (MEA) in 317 patients undergoing angioplasty and stenting for cardiovascular disease. Baseline, TRAP-6-, and ADP-inducible P-selectin expression, activated glycoprotein IIb/IIIa (GPIIb/IIIa) and monocyte-platelet aggregate (MPA) formation were measured as sensitive parameters of platelet activation. RESULTS: In patients with high on-treatment residual ADP-inducible platelet reactivity (HRPR), baseline P-selectin expression, GPIIb/IIIa and MPA formation were similar to those in patients without HRPR (all P > 0.05). After platelet activation with TRAP-6 or ADP, patients with HRPR by both assays exhibited significantly higher levels of P-selectin expression, GPIIb/IIIa and MPA formation than patients with an adequate thienopyridine-mediated platelet inhibition (all P ≤ 0.02). However, high levels of TRAP-6-inducible P-selectin, GPIIb/IIIa and MPA formation also occurred in 20.4%, 19.1% and 20.1% of the good responders by the VASP assay, and in 19.6%, 16.6% and 20.6% of the good responders by MEA, respectively. CONCLUSIONS: Thienopyridine nonresponders are more susceptible to thrombin- and ADP-inducible platelet activation than patients with good platelet inhibition. However, even patients with adequate thienopyridine-mediated platelet inhibition often show a preserved responsiveness to thrombin. These patients may benefit from additional thrombin receptor blockage or inhibition of thrombin generation.

Sensitivity of assays for the detection of HPA-1a antibodies: results of an international workshop demonstrating the impact of cation chelation from integrin αIIbß3 on three widely used assays.

BACKGROUND AND OBJECTIVES: HPA-1a antibodies account for 70-80% of cases of fetal-neonatal alloimmune thrombocytopenia (FNAIT) in Caucasians. However, numerous workshops have demonstrated variability in their detection. We recently showed that exposure of αIIbß3 to ethylene diamine tetraacetic acid (EDTA) affected binding of many anti-αIIbß3 monoclonal, and HPA-1a allo-, antibodies; this adversely affected sensitivity of the monoclonal antibody-specific immobilization of platelet antigens (MAIPA) assay and indirect platelet immunofluorescence test (PIFT). This study presents results from an international workshop studying the impact of cation chelation on HPA-1a antibody detection in routine diagnostic laboratories. MATERIALS AND METHODS: Serum and EDTA-anticoagulated plasma samples containing anti-HPA-1a were distributed to 39 laboratories. Participants were asked to detect and identify any HPA antibodies present. RESULTS: 2/39 (5.1%) participants were able to detect and identify anti-HPA-1a in the serum, but not in the plasma sample. EDTA plasma reduced MAIPA assay sensitivity by ≥ 20% in 17/24 (70.8%) laboratories and by ≥ 50% in 9/24 (37.5%) when using HPA-1a1a platelets (mean: 27.7%, range 0-85.1%); when using HPA-1a1b platelets 3/4 (75%), participants reported ≥ 50% loss of sensitivity (mean 65.6%, range 0-96.6%). A small but significant increase in optical densities was observed in antigen capture ELISA assays when using plasma (mean difference: 0.081, P < 0.01). Insufficient PIFT data were returned to draw firm conclusions. CONCLUSION: Use of EDTA plasma significantly affects the sensitivity of the MAIPA assay and can affect detection of even potent, FNAIT-causing examples of anti-HPA-1a. These data highlight the importance of use of αIIbß3 in an appropriate conformation for the sensitive detection of anti-HPA-1a.

The GNB3 C825T polymorphism influences platelet aggregation in human whole blood.

BACKGROUND: Platelet aggregation varies among individuals; and genetic factors may alter platelet activation through G-protein-coupled receptors, thus influencing results of point-of-care platelet aggregometry in whole blood. We tested the hypothesis that the C825T polymorphism of the gene GNB3 encoding the G-protein ß-3 subunit and the platelet GPIIIa Pl(A1)/(A2) polymorphism of the glycoprotein IIIa influence platelet aggregation. METHODS: Evoked [thrombin receptor activating peptide (TRAP), ADP, TXA(2) agonist U46619, epinephrine, and collagen] platelet aggregation in whole blood was measured using impedance aggregometry (Multiplate) in 143 healthy individuals (age: 40.2 years ±11.7 SD). Genotypes were determined using pyrosequencing and restriction analysis. Data were analyzed by linear one-way analysis of variance and Student's t-test, linear and multiple regression, and the χ(2)-test, as appropriate. RESULTS: Homozygous carriers of the GNB3 825C-allele showed significantly (P≤0.022) increased maximum aggregation for EC(75) dosages compared with CT and TT genotypes [e.g. ADP: CC 150±36 vs. TT 126±33 aggregation unit (AU); thrombin receptor activating peptide: CC 175±46 vs. TT 150±38 AU; U46619: CC 164±33 vs. 149±32 AU; epinephrine: CC 66±41 vs. TT 48±34 AU]. In contrast, genotypes of glycoprotein IIb/IIIa PI(A)-polymorphism had no effect. Regression analysis revealed the GNB3 C825T polymorphism as an independent factor for enhanced platelet aggregation, besides factors such as female sex and blood cell values. CONCLUSION: In human whole blood, the GNB3 825CC genotype is associated with enhanced platelet aggregation.

Novel function of tenascin-C, a matrix protein relevant to atherosclerosis, in platelet recruitment and activation under flow.

OBJECTIVE: The identification of platelet-reactive proteins exclusively present in atherosclerotic plaques could provide interesting targets for effective and safe antithrombotic strategies. In this context, we explored platelet adhesion and activation to tenascin-C (TN-C), a matrix protein preferentially found within atheroma. METHODS AND RESULTS: We show that platelets efficiently adhere to TN-C under both static and flow conditions. Videomicroscopy revealed a unique behavior under flow, with platelets exhibiting stationary adhesion to TN-C; in contrast, platelets rolled over von Willebrand factor and detached from fibrinogen. Platelet interaction with TN-C was predominantly supported by integrin α(2)ß(1) under static conditions, whereas under high shear, it was dependent on both the α(2)ß(1) integrin and the glycoprotein Ib-IX complex. Integrin α(IIb)ß(3) appeared to play a secondary role but only at low shear rates. The glycoprotein Ib-IX-dependent interaction was indirect, relying on von Willebrand factor, and increased as a function of wall shear rate. Von Willebrand factor bound directly to TN-C, as shown by ELISA and coimmunoprecipitation, suggesting that it acts as a bridge between TN-C and platelets. The adhesion of platelets to TN-C triggered their activation, as demonstrated by a shape change and increases in intracellular calcium level. CONCLUSIONS: This study provides evidence that TN-C serves as a novel adhesive matrix for platelets in a context that is relevant to atherothrombosis.

Properties of procoagulant platelets: defining and characterizing the subpopulation binding a functional prothrombinase.

OBJECTIVE: The goal of this study was to define and characterize the subpopulation of platelets capable of regulating the functional interactions of factors Va (FVa) and Xa (FXa) on the thrombin-activated platelet surface. METHODS AND RESULTS: Flow cytometric analyses were used to define and characterize platelet subpopulations. At a concentration of thrombin known to elicit maximal platelet activation, platelet-derived FVa release, and prothrombinase assembly/function, only a subpopulation of platelets was positive for FVa and FXa binding. An additional subpopulation bound lower levels of FVa but little, if any, FXa. Fluorescence microscopy analyses confirmed these data. Phenotypically, platelets capable of binding FXa were more highly reticulated and demonstrated significantly increased expression of several key adhesion molecules, including P-selectin, glycoprotein Ibα, and integrins α(IIb) and ß(3). This platelet subpopulation was also defined by the expression of a nondissociable, membrane-bound pool of functional platelet-derived FVa, which made up ≈35% to 50% of the total membrane-bound cofactor. CONCLUSIONS: The ability of activated platelets to support thrombin generation is defined by a subpopulation of platelets expressing a nondissociable pool of platelet-derived FVa and increased adhesive receptor density. This subpopulation is hypothesized to play a significant role in regulating both normal hemostasis and pathological thrombus formation because the adherent properties of platelets and their ability to mount and sustain a procoagulant response are crucial steps in both of these processes.

Biomarkers involved in evaluation of platelets function in South-Eastern Romanian patients with hematological malignancies subtypes.

ABSTRACT: At present, various researches presented how subtypes of hematological malignancies are related to stages of the immune response, because the activated immune system represents a promising form in cancer treatment. This study explores the relationship between the adaptive immune system (T cells), and the coagulation system (platelets, platelet membrane glycoproteins, platelets derivate microparticles) which seems to play an important role in host immune defense of patients with acute myeloblastic leukemia (AML) or B cell lymphoma (BCL), 2 of the most common hematological malignancies subtypes.Blood samples (n = 114) obtained from patients with AML or BCL were analyzed for platelet membrane glycoproteins (CD42b, CD61), glycoprotein found on the surface of the T helper cells (CD4+), protein complex-specific antigen for T cells (CD3+), platelet-derived microparticles (CD61 PMP) biomarkers by flow cytometry, and hematological parameters were quantified by usual methods.In patients with AML, the means of the percentage of the expressions of the molecules on platelet surfaces (CD61 and CD42b, P < .01; paired T test) were lower as compared to both control subgroups. The expression of cytoplasmic granules content (CD61 PMP) had a significantly higher value in patients with AML reported to controlling subgroups (P < .01; paired T test), which is suggesting an intravascular activation of platelets.The platelet activation status was presented in patients with low stage BCL because CD61 and CD42b expressions were significantly higher than control subgroups, but the expression of CD 61 PMP had a significantly decreased value reported to control subgroups (all P < .01; paired T test). T helper/inducer lineage CD4+ and T lymphoid lineage CD3+ expressions presented significant differences between patients with AML or low stage BCL reported to control subgroups (all P < .01; paired T test).Platelet-lymphocyte interactions are involved in malignant disorders, and CD61, CD42b present on platelet membranes, as functionally active surface receptors mediate the adhesion of active platelets to lymphocytes, endothelial cells, and cancer cells.

Evaluation of low platelet counts by optical, impedance, and CD61-immunoplatelet methods: estimation of possible inappropriate platelet transfusion.

BACKGROUND: The Cell-Dyn Sapphire (Abbott Diagnostics) detects platelets (PLTs) with a CD61 monoclonal antibody directed against glycoprotein IIIa as well as impedance (IMP) and optical (OPT) technology. We decided to evaluate low PLT counts produced by IMP and OPT methods and to compare them with the CD61 method. We also examined the possibility of inappropriate PLT transfusion resulting from an inaccurate PLT count. STUDY DESIGN AND METHODS: We analyzed consecutive blood samples with OPT PLT counts of less than 50 x 10(9)/L. We performed the PLT count with the OPT, IMP, and CD61 methods and we compared the number of prophylactic PLT transfusion indications according to the PLT counts determined by the OPT and IMP methods with the number of prophylactic PLT transfusion indications according to our reference CD61 method. RESULTS: We collected 135 samples. In the bias analysis, the OPT method and the IMP method showed higher PLT counts when compared with the CD61 method (mean of difference 1.69 x 10(9) and 19.1 x 10(9)/L, respectively). We saw overtransfusion in 1.5% of cases and undertransfusion in 15.2% of cases (p = 0.01; McNemar's test) when we selected a threshold of 10 x 10(9)/L with the OPT method. We saw undertransfusion in 22.2% of cases (p = 0.03; McNemar's test) when we selected a threshold of 5 x 10(9)/L with the OPT method. CONCLUSIONS: Low PLT counts determined by the OPT and IMP methods showed some disagreement when compared with the CD61 method. This disagreement caused both PLT undertransfusion and overtransfusion.

Neonatal alloimmune thrombocytopenia caused by an antibody specific for a newly identified allele of human platelet antigen-7.

BACKGROUND: Neonatal alloimmune thrombocytopenia (NAIT) is a neonatal disorder characterized by maternal alloimmunization against fetal platelet (PLT) antigens inherited from the father. A healthy 30-year-old Japanese woman (Hit) gave birth to her second child after an uneventful pregnancy. Nine hours after birth, the infant presented with severe petechiae and a PLT count of 6 x 10(9)/L. STUDY DESIGN AND METHODS: To elucidate the maternal cause of NAIT in the infant, serologic and genetic studies, including PLT genotyping and sequence-based analysis, were conducted. Additionally, serologic screening for the new PLT antigen was performed. RESULTS: Serum from the NAIT infant's mother contained antibodies directed against a human PLT antigen (HPA) of the newborn. Using five-cell-lineage flow cytometry, we localized the antigen to a PLT glycoprotein (GP). Subsequent monoclonal antibody immobilization of PLT antigen assay and PLT immunofluorescence inhibition experiments localized the antigen to the GPIIIa subunit of the GPIIb/IIIa complex. GPIIIa localization was confirmed by sequence-based typing studies, which identified a 1297C>T (407proline>serine substitution) mutation on the ninth exon of the GPIIIa gene. This mutation identified the third allele of HPA-7. Anti-Hit(a) reacted with mutated GPIIIa-transfected cells but not with stable transfectants expressing wild-type GPIIIa. Serologic screening for Hit(a) in the Japanese population revealed a phenotypic frequency of approximately 0.0015. CONCLUSIONS: We identified a new third allele of HPA-7, which is characterized by a 1297C>T mutation in the GPIIIa gene. This 1297C>T allele was found in 0.15% of the Japanese population. An antibody against this antigen could be the cause of severe NAIT.

Acute megakaryoblastic leukemia in dogs: a report of three cases and review of the literature.

Three dogs of different breeds, ages, and genders were presented with pale mucous membranes, depression, anorexia, and splenomegaly. Observed were severe normocytic, nor-mochromic, nonregenerative anemia, thrombocytopenia, and leukopenia. Blood smears contained large, atypical cells with blue vacuolated cytoplasm, cytoplasmic blebs, round to oval central nuclei, and elevated numbers of cytoplasmic fragment resembling macroplatelets. Bi- and multinucleated atypical cells were found mainly in spleen, lymph nodes, and bone marrow. A final diagnosis of acute megakaryoblastic leukemia (AMegL) was made based on morphology and positivity to the megakaryocyte-derived cell-specific markers von Willebrand factor and CD61. In case nos. 1 and 2, no treatment was initiated, and the dogs died on days 4 and 3, respectively. Case no. 3 received supportive therapy with prednisone, and after a brief improvement the dog died spontaneously 35 days after initial presentation. Only 11 cases of AMegL have been reported in dogs, and the specific diagnostic criteria have not been well established. The presence of vacuolization, cytoplasmic blebs, central round nuclei, cytoplasmic fragments, and multinucleated cells in these three cases were considered useful to differentiate AMegL from other hematopoietic neoplasms.

[The influence of GPIIIA gene polymorphism on the variability of standard electrocardiogram in patients with acute coronary syndrome].

The authors analyse effect of GPIIIA gene (PI a allele) polymorphism on the frequency of complicated coronary heart disease in patients with dyslipidemia and hypertensive disease. Specific features of ventricular repolarization (T-wave variability) in patients with acute coronary syndrome are described.

[Platelet structure and functions (a review of literature). Part 1].

The paper reviews literature updates on the organization and specific features of megakaryocytopoiesis, the mechanisms of its regulation, possible ways for platelets to occur from megakaryocytes. It characterizes thrombocytopoiesis, by describing the detailed structural organization of platelets and gives data on the relationship of the morphofunctional features of platelets to the ploidy of megakaryocytes that give rise to the latter. The structure and clinical value of a number of platelet activation markers, such as CD41/CD61, CD42, CD62p, platelet-leukocyte aggregates, microvesicles, are described.