BACKGROUND

Clinically chronic renal allograft dysfunction (CRAD) manifests as a progressive elevation of serum creatinine (Cr) with an associated deterioration in proteinuria and decrease in glomerular filtration rate (GFR) following the first year after transplantation. Currently, how to delay or reverse CRAD and improve the long-term survival of transplanted kidneys is an intensively researched topic in the transplantation field.

OBJECTIVE

The objective of this study was to examine the therapeutic efficacy and safety profile of inhibiting platelet activation on living related donor renal transplant recipients with CRAD.

METHODS

We performed a prospective cohort study to examine the effects of inhibiting platelet activation on the expression of platelet activation markers, namely platelet surface glycoprotein IIIa (CD61), lysosomal enzyme glycoprotein (CD63), and fibrinogen receptor monoclonal antibody (PAC-1); we also studied platelet-derived growth factor-BB (PDGF-BB) and transforming growth factor β(1) (TGF-β(1)) in the peripheral blood of renal graft recipients with CRAD. In addition, we correlated this metrics with the function of transplanted kidneys, peripheral blood platelet counts and trough concentrations of immunosuppressants.

RESULTS

Inhibiting platelet activation decreased the expression of platelet activation markers in peripheral blood: CD61 (P < .001), CD63 (P = .031), PAC-1 (P = .002), PDGF-BB (P = .013), and TGF-β(1) (P = .030). Cr and urea nitrogen remained stable compared with the control group, which showed progressive increases. The GFR of transplanted kidneys was improved (P = .033); however, there was no difference before versus after treatment in the platelet count (P = .991) or the trough concentration of immunosuppressants (cyclosporine, P = .071; tacrolimus, P = .984).

CONCLUSIONS

Inhibition of platelet activation, which was effective and safe, seemed to treat CRAD probably through improving the microcirculation of transplanted kidneys, down-regulating the expression of TGF-β, and thereby delaying fibrosis.

An increased platelet activation status is present in patients with VVI pacemakers. With platelet activation, there is modulation of platelet surface molecule expression. In the current study, the expression of platelet surface markers in VVI patients before and after ticlopidine treatment and control subjects was investigated by means of flow cytometry. The study group consisted of 25 patients with VVI pacemaker, and 15 control subjects. CD42b, CD61, and CD62p expression were significantly increased in VVI patients compared with control subjects (CD42b p < 0.001, CD61 p< 0.005 and CD62p p < 0.001). In addition, after ticlopidine treatment, platelets showed a significant fall in expression of all these markers in VVI patients (CD42b p < 0.001, CD61 p < 0.005 and CD62p p< 0.001). Our data suggest an increase of the surface expression of all these markers on platelets and demonstrate the efficacy of ticlopidine in reducing them.

EGFR inhibitors spur tumor cells to express CD61 and become more stem cell-like. A combination treatment that includes bortezomib might reverse these changes.

Following an immunomagnetic isolation and enrichment procedure, CD34+ cells were harvested from the peripheral blood of about 50 healthy donors. A battery of cytochemical staining reactions, monoclonal and carbohydrate-specific antibodies, proliferation markers and lectins was applied on smears and sections from paraffin-embedded pellets. Additionally, a morphometric analysis and ultrastructural investigation was carried out. More than 95% of the total yield of progenitor cells expressed CD34 and CD43 (MT1) and of these about 90% CD45 (LCA) and 25% CD45A (MT2). The CD34+/CD45RA-population was thought to represent very primitive, probably not lineage-restricted stem cells. On the other hand, reactivity with ANAE, CD11c, CD15, CD20, Ret40f, KiM1P, and CD61 (ranging between 1 to 20%) was considered to indicate a transition into more differentiated elements of hemopoiesis. The failure to detect any staining with proliferation markers (Ki-67/MIB 1, PCNA, KiS1) was in keeping with a quiescent status. Carbohydrate antigens revealed a pattern which underlines the fact that the CD34 and CD43 antigens belong to the family of heavily O-glycosylated sialomucins. Blood group antigens which are located at the peripheral regions of mucin-oligosaccharides (H type 2, Lewis, Lewis) could be demonstrated, but not A, B, Sialyl-Lewis and Lewis. Morphometric analysis revealed that CD34+ progenitors were larger than small lymphocytes. Electron microscopy showed a relatively primitive cytoplasmic organization and numerous tiny magnetic beads clustered at the plasma membrane.

OBJECTIVE

OLT is the best alternative for patients with end-stage liver diseases. However, as the need for organs surpasses donor availability, alternatives to OLT are required. LCT could be a useful option versus OLT in several patients even though its low cell-engraftment hampers its efficiency. Endothelial cell barrier is the main obstacle for the implantation of cells into the parenchyma. Our study has focused on the modification of the endothelial barrier with monoclonal antibodies against adhesion molecules in order to increase cell engraftment in a mouse model of liver cell transplantation.

METHODS

Anti-mouse CD54 and anti-mouse CD61 antibodies were administered intrasplenically to healthy mice within 60 min prior to stem cell transplantation. Animals were sacrificed either short term at 2h or middle term seven days after transplantation. Immunohistochemical techniques to detect alkaline phosphatase activity were used to identify the transplanted cells within the liver parenchyma.

RESULTS

Anti-CD54 and anti-CD61 administration increases vascular patency and cell engraftment. This represents a 32% and 45% increase, respectively, of engrafted cells compared to the control (p<0.05).

CONCLUSIONS

Modification of the vascular wall with monoclonal antibodies against endothelial adhesion molecules before cell transplantation enhances cell engraftment into the mouse liver.

OBJECTIVE

To investigate the influence of depsides salts from salvia miltiorrhiza (DSM) on the functions of platelet and vascular endothelial cell in severe chronic obstructive pulmonary diseases (COPD) patients with acute exacerbation.

METHODS

Forty patients with severe COPD in acute exacerbation stage were randomly divided into two groups: conventional treatment (CT) group and DSM treatment (DSM) group, each consisting of 20 cases, and 20 healthy adults served as control group. All COPD patients were given conventional treatment, while for the patients in DSM group 0.2 g depsides DSM was given through intravenous drip everyday in addition for 2 weeks. The levels of the plasma platelet membrane glycoprotein 140 (GMP140) and von Willebrand factor (vMF) in the blood samples were determined with enzyme linked immunosorbent assay (ELISA) and the level of CD62P/CD61 with flow cytometry (FCM).

RESULTS

The level of GMP140 [CT group: (17.51+/-2.75) microg/L, DSM group: (16.94+/-2.57) microg/L], vMF [CT group: (13.64+/-2.37) microg/L, DSM group: (14.14+/-2.17) microg/L] and CD62P/CD61 [CT group: (20.24+/-2.64)% , DSM group: (19.54+/-2.69)%] were elevated significantly in severe COPD patients with acute exacerbation compared to the control group before treatment [(11.21+/-1.11)%, (9.25+/-1.80) microg/L, (6.13+/-1.17) microg/L, all P<0.01]. After intervention, the levels of the above three indexes in both treatment groups were significantly decreased, and the decrease in DSM group [GMP140: (3.91+/- 0.57) microg/L , vWF: (3.86+/-0.71) microg/L, CD62P/CD61: (3.69+/-0.62)%] was more prominent than the CT group [GMP140: (2.30+/-0.33) microg/L, vWF: (2.72+/- 0.45) microg/L , CD62P/CD61: (2.24+/-0.45)%, all P<0.01], but they were higher than normal levels.

CONCLUSIONS

DSM has the effect of inhibiting the activation of vascular endothelial cells and platelet. The medicine may be used to prevent thrombotic diseases.

Cyclodextrins (CDs), a class of cyclic oligosaccharides formed by α-(1,4) linked glucopyranose units, were functionalized with (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO) radicals to prepare water soluble supramolecular organic radical contrast agents (ORCAs) for the in vivo detection of glioma tumor in animal models. A first set of molecules (CDn1, n=6,7,8 is the number of both TEMPO and glucopyranose units) was studied by superconducting quantum interference devices (SQUID) magnetometry in order to define the role of the CD macrocycle on the effective magnetic moment (μ_eff ). The μ_eff value increased from 3.982 μ_B (CD61) to 4.522 μ_B (CD81) but was limited by intramolecular antiferromagnetic (AF) interactions. A set of water-soluble ORCAs (CDn8, n=6,7,8) was prepared by a sequence of thiol-ene and Cu(I)-catalyzed alkyne-azide "click" reactions. Their ¹ H water relaxivities r₁ of these ORCAs were between 0.739 mM^-1 s^-1 (CD68) to 1.047 mM^-1 s^-1 (CD88) in D₂ O/H₂ O 9 : 1 (v : v) at 300 K. One of them (CD78) was tested on glioma-bearing rats with reduced side effects and good relaxivity in vivo.

Keywords: contrast agents; cyclodextrins; gliomas; magnetic resonance imaging; polynitroxides.

An 11-month-old boy was transferred to our hospital because of fever and bleeding tendency on March 13, 1998. Laboratory studies showed a white blood cell count of 43,360/microliter with 75% blasts, a hemoglobin concentration of 8.4 g/dl, and a platelet count of 23 x 10(3)/microliter. Surface marker analysis with a flow cytometer revealed that only 21% and 11% of the blasts, respectively, were positive for CD41 and CD42b. Treatment with a permeabilizing agent apparently increased the reactivity of the blasts with anti-CD41 monoclonal antibody (MoAb), which can recognize IIb independently of IIIa. However no significant differences were observed in reactivity with anti-CD41 MoAb (which recognizes the IIb/IIIa complex) anti-CD61 MoAb and anti-CD42b MoAb before or after fixation. Blasts positive for platelet peroxidase were observed by electron microscopy, thus confirming the diagnosis of acute megakaryoblastic leukemia. We concluded that the detection of intracellular antigens is useful for the quick diagnosis of acute megakaryoblastic leukemia characterized by low surface expression of megakaryocytic lineage antigens.

BACKGROUND

Biological studies on megakaryopoiesis have been hampered by the scarcity of megakaryocytes in normal bone marrow and difficulty in long term culture. Alternatively, leukemic cell lines with megakaryocytic differentiation potential may provide good models to counter these problems.

METHODS

Leukemic cells from a patient with acute megakaryocytic leukemia were put into long-term culture and established into a cell line which was designated as VGH-MK1. The VGH-MK1 cells were challenged with differentiation agents and/or cytokines, and the differentiation of these cells was examined using morphological, immunocytochemical and surface-marker studies.

RESULTS

Morphologically, VGH-MK1 cells had prominent nucleoli and basophilic cytoplasm with some protrusions, but large cells were occasionally seen. Under regular culture condition, the cells had a doubling time of 36-48 hours. The cloned cell line exhibited markers characteristic of megakaryoblasts after differentiation induction. Specifically, when stimulated with 12-o-tetradecanoylphorbol-13-acetate (TPA), cells became larger and had large or multinuclei. They were induced to express platelet glycoproteins GPIb (CD42b), GPIIb/IIIa (CD41), and GPIIIa (CD61) antigens, but not erythroid nor lymphoid markers. Platelet peroxidase (PPO) activity was also induced. Retinoic acid did not exhibit similar differentiation-inducing effects. In contrast, it counteracted the effects induced by TPA.

CONCLUSIONS

An unique human leukemic cell line, VGH-MK1, has been established here. It could be induced to exhibit some characteristics of megakaryocytic lineage, and may be an useful model for the biological studies of megakaryopoiesis.

Platelet hyperaggregation in ischaemic stroke patients is a proven finding, and associated with increased expression of the platelet surface GPIIb/IIIa receptor. The polymorphism occurs at nucleotide position 1565 of the GPIIIa gene resulting a 33Leu-Pro change. Data are conflicting regarding the abnormal function of the PlA1/A2 receptor in stroke. The aim of the study was to address the difference of platelet receptor function in ischemic stroke patients with the wild PlA1/A1 and heterozygous PlA1/A2 genotype. A total of 51 patients with PA1/A1 and 54 patients with PlA1/A2 genotypes were enrolled. Polymerase chain reaction was used for genotyping of platelets. Platelet aggregation was measured in whole blood and in platelet rich plasma (PRP). Flow cytometry was used for measuring surface molecule expression (CD42b, CD41a, CD61, CD62P) and fibrinogen binding capacity of cells with phosphate buffer solution (PBS) in comparison with activation by ristocetin in whole blood as well as by adenosine diphosphate (ADP) in PRP. In comparison with wild types, platelets carrying the PlA1/A2 genotypes showed hyperaggregation measured in whole blood and induced by ristocetin (p< 0.05). Using whole blood flow cytometry with ristocetin induction, the CD62P+/FIB- (P selectin) and the CD62P+/FIB+ were more expressed in heterozygous platelets as compared to wild types (p< 0.01 and p< 0.05), respectively. According to mean fluorescence intensity with ADP induction, an increased expression of CD61+, CD61+/CD41+ and CD62P+ in PlA1/A2 platelets were detected as compared to the group carrying the wild type (p< 0.0001, p= 0.006, p= 0.0001), respectively. These findings support the possibility that in ischaemic stroke patients, platelets carrying PlA1/A2 genotypes can be activated by different inductors in a way, which leads to permanent hyperfunction of platelet surface receptor GPIIIa.

BACKGROUND

Thrombosis is a dangerous complication of paroxysmal nocturnal hemoglobinuria (PNH) and has a high mortality rate. However, the mechanism underlying the development of thrombosis in PNH remains unclear. To explore this, platelet function and serum complement activity were investigated in 14 patients with classical PNH, 11 with PNH aplastic anemia (AA) and 30 healthy controls.

METHODS

Serum concentrations of the terminal complement complex (sC5b-9) were determined by enzyme-linked immunofluorescence assay (ELISA), and the levels of C5b-9, CD61 and CD62p on platelet membranes were determined by flow cytometry. Clinical parameters were assessed, including D-dimer and platelet aggregation induced by adenosine diphosphate (ADP) and arachidonic acid (ARA).

RESULTS

Serum sC5b-9 concentrations were significantly lower in the PNH/PNH-AA than in the control group (P<0.01). C5b-9 deposition was significantly higher on CD59- platelets than on CD59+ platelets in PNH/PNH-AA patients and healthy controls (P<0.01 for each). D-dimer concentration was significantly higher in PNH/PNH-AA patients - especially those with lactate dehydrogenase (LDH) concentrations>1000U/L - than in controls (P<0.05). CD61 (P<0.05) expression was lower on CD59+ platelets in PNH than in controls and CD5- platelets in PNH. Expression of CD62p (P<0.01) was lower on CD59- and CD59+ platelets (P<0.01) in PNH cases than in controls. Platelet aggregation stimulated by the agonists ADP and ARA in the PNH/PNH-AA patients was significantly lower than that in controls (P<0.05).

CONCLUSIONS

The adhesion and aggregation of platelets, especially of CD59+ platelets, were compensatively decreased in PNH/PNH-AA patients without active thrombosis.

OBJECTIVE

To evaluate whether markers of platelet activation, including P-selectin expression, phosphatidylserine exposure, platelet-leukocyte aggregates, and microparticle formation, could be measured in nonstimulated and stimulated canine blood samples and develop a standardized protocol for detection of activated platelet markers in canine blood.

METHODS

Blood samples from 10 dogs.

METHODS

Platelet activation was determined by flow cytometric measurement of platelets with P-selectin expression, platelet-leukocyte aggregates, platelet microparticles, and platelets with phosphatidylserine exposure. Changes in specific markers of platelet activation in nonstimulated versus stimulated samples were assessed by use of varying concentrations of 2 platelet agonists, platelet-activating factor (PAF) and adenosine diphosphate. Flow cytometry was used to detect platelet CD61 (glycoprotein IIIa), CD62P (P-selectin), and the leukocyte marker CD45. Annexin V was used to identify exposed phosphatidylserine.

RESULTS

A significant difference was detected in the percentages of platelets with P-selectin, plateletleukocyte aggregates, microparticles, and platelets with annexin V exposure (phosphatidylserine) in samples stimulated with 10nM PAF versus the nonstimulated samples, with platelet-leukocyte aggregates having the greatest difference.

CONCLUSIONS

Platelet activation is essential for thrombus formation and hemostasis and may be potentially useful for evaluation of dogs with suspected thromboembolic disease. Prior to development of a thrombotic state, a prothrombotic state may exist in which only a small number of platelets is activated. Identification of a prothrombotic state by use of activated platelets may help direct medical intervention to prevent a thromboembolic episode.

In August, 1992, we established a leukemic cell line (NS-Meg) from a patient in megakaryoblastic transformation of Philadelphia chromosome-positive chronic myeloid leukemia. The NS-Meg cells were positive for alpha-naphthyl acetate esterase and periodic acid-Schiff (PAS) staining and for surface CD4, CD7, CD13, CD34, CD41a, and glycophorin A antigens. Ultrastructurally, the cells had alpha-granules, demarcation membranes, and platelet peroxidase activity. The NS-Meg cells spontaneously produced platelet-like particles which contained alpha-granules, mitochondria and dense bodies, strongly suggesting platelet production. Erythropoietin (Epo), granulocyte/macrophage colony stimulating factor(GM-CSF), and interleukin 3 (IL-3) promoted the growth of NS-Meg cells. Phorbol-12-myristate-13-acetate increased the expression of both CD41a and CD61 antigens. Ten-day exposure to Epo induced mature erythroblasts and red cells. These benzidine-positive cells were positive for hemoglobin F staining. Untreated NS-Meg cells expressed mRNA for the Epo receptor (EpoR), for GATA-1, and for alpha 1, alpha 2 and gamma globin genes. These results indicate that NS-Meg cells undergo terminal differentiation of both megakaryocytic and erythroid lineages. This cell line should be a very useful tool for the investigation of both megakaryocytic and erythroid maturation.

In October 1992, a 36-year-old man was diagnosed as having mediastinum mixed germ cell tumor (stage II), and was treated with surgical operation and combination chemotherapy including VP16 (total VP16 dose; 1,500 mg/m2). After that, remission had been sustained, but leukocytosis (15,700/microliter) with 37% of peroxidase-negative blasts and thrombocytopenia developed in September, 1995. Bone marrow showed remarkable reticulin fibrosis and increase of atypical immature cells that were immunophenotypically factor VIII+/CD42+/CD61+. Thus, we diagnosed acute megakaryoblastic leukemia (M7). Based on no abnormality of chromosome 11q23 and no rearrangements of MLL gene, we diagnosed the syndrome of mediastinal germ-cell tumors associated with hematologic neoplasia. Furthermore, the neuron-specific enolase level was elevated (95.9 ng/ml). Soon after complete remission was reached by combination chemotherapy, the leukemia was relapsed, and the he died 3 months after the onset of leukemia. To our knowledge, this is the third case report of this syndrome in Japan and the first one of leukemia with high level of serum neuron-specific enolase.

We have established a new hematopoietic cell line from a patient with myelodysplastic syndrome (MDS), which was refractory anemia with excess blasts (RAEB). This cell line, designated TER-3, depends on several cytokines for long-term survival and growth, and requires interleukin-3 (IL-3) for continuous growth. Cytochemical analysis revealed that TER-3 cells are weakly dianisidine positive and nonspecific esterase positive, but peroxidase negative. The surface marker profile shows that the TER-3 cells are strongly positive for myeloid, lymphoid, and megakaryocytic antigens such as CD15, CD19, and CD61, and negative for some common multilineage antigens such as CD13, CD33, and CD34. Thus, this cell line has a multilineage phenotype, suggesting that the transformation event occurred in multipotent stem cells. Dianisidine- and nonspecific esterase-positive TER-3 cells increase with granulocyte-colony stimulating factor (G-CSF) rather than with IL-3. These results suggest that the cell line is useful for understanding the mechanism underlying G-CSF-associated hematopoietic cell differentiation and activation in the patient with MDS.

An autopsy case (85-year-old Japanese male) of myeloperoxidase- (MPO-) positive acute myeloid leukemia with maturation (M1) accompanying megakaryocytic differentiation is presented. The patient manifested acute coronary syndrome. Even after emergent percutaneous coronary intervention, his performance status remained poor, so no chemotherapy against leukemia was given. The final white blood cell count reached 291,700/μL, and the platelet count was elevated to 510,000/μL. No cytogenetic studies were performed. He died at the 25^th day of hospitalization. Autopsy revealed marked leukemic infiltration to the endocardium and subendocardial myocardium. Subendocardial myonecrosis was surrounded or replaced by the leukemic blasts, and neither granulation tissue reaction nor fibrosis was observed. In the cardiovascular lumen, lard-like blood clots were formed and microscopically consisted of leukemic blasts and platelets (leukemic thrombi). Infiltration of leukemic blasts was seen in the body cavities and systemic organs including the lung. The MPO-positive blasts lacked azurophilic granules and expressed the stem cell markers, CD34 and CD117 (c-kit). No features of myelofibrosis were seen in the 100% cellular marrow. In the endocardium, liver, lymph nodes, and bone marrow, megakaryocytic cells (CD42b/CD61+, MPO-) were distributed, while the small-sized blastic cells in the blood and tissues predominantly expressed MPO. The blasts lacked expression of CD42b/CD61. Megakaryocytic differentiation might be stimulated by certain tissue factors. AML accompanying megakaryocytic differentiation in certain tissues and organs should be distinguished from acute megakaryoblastic leukemia. The mechanisms provoking acute coronary syndrome in acute myeloid leukemia are discussed.

Background: Brief non-harmful ischemia, remote ischemic preconditioning (RIPC) has been proposed to confer benefit to patients with coronary artery disease (CAD) via unknown mechanisms.

Objectives: We aimed to investigate the effect of RIPC on circulating levels of extracellular vesicles (EV) and global coagulation and fibrinolytic factors in patients with coronary disease.

Patients/methods: Blood samples were taken from 60 patients presenting for coronary angiography enrolled in a randomized, controlled trial before and after RIPC (3×5 min administration of 200 mmHg sphygmomanometer on the arm, n=31) or sham (n=29) treatment. Most patients (n=48) had significant coronary artery disease and all were taking at least one antiplatelet agent.

Results: RIPC significantly decreased circulating levels of EV expressing platelet markers CD41 and CD61 detected by flow cytometry in plasma, while no such effect was found on EV expressing phosphatidylserine, CD62P, CD45, CD11b, CD144, CD31+/CD41-, or CD235a. RIPC had no effect on the overall hemostatic potential (OHP) assay or circulating antigen levels of tissue plasminogen activator, urokinase, plasminogen activator inhibitor-1, or plasminogen. Sham treatment had no effect on any studied parameter. Statin use inhibited the effect of RIPC on CD61+ EV, diabetes modified the effect of RIPC on CD45+ and CD11b+ EV, and hypertension modified the effect of RIPC on CD235a+ EV.

Conclusions: RIPC decreased circulating levels of platelet-derived EV in patients with coronary disease taking conventional antiplatelet therapy. This may reflect increased EV clearance/uptake or change in production. Clinical variables may alter the effectiveness of RIPC.

Keywords: blood coagulation; coronary artery disease; extracellular vesicles; fibrinolysis; ischemic preconditioning.

An adult, mixed-breed, feline leukaemia virus (FeLV-) positive female cat was presented with mucosal jaundice and a history of anorexia and constipation for three days. Physical examination revealed splenomegaly, cachexia, and dehydration. Humane euthanasia was conducted, followed by postmortem examination. Grossly, the cat was icteric, and presented hepatomegaly with multifocal white spots and splenomegaly. Histologically, the bone marrow was nearly completely replaced by a proliferation of megakaryocytes and megakaryoblasts, and there was a proliferation of fibrous connective tissue. Similar neoplastic proliferation was observed infiltrating the liver, lymph nodes, spleen, kidney, skeletal muscle, and lungs. Immunohistochemistry was performed for von Willebrand Factor (VWF), CD79α, CD3, feline immunodeficiency virus, FeLV, and CD61. Marked cytoplasmic labelling was observed in the neoplastic cells for FeLV, VWF and CD61, corroborating the diagnosis of acute megakaryoblastic leukaemia.

Keywords: CD61; FeLV; acute myeloid leukaemia; feline; myelofibrosis.

Cell motility and changes in cell shape are largely powered by actin polymerization and depolymerization. Eight to ten second periodic changes in human polymorphonuclear neutrophil (PMN) shape were detected by video-image analysis of PMN crawling on a surface and by right angle light scattering (RALS) in suspended PMN. However, sustained RALS oscillations in suspended PMN requires pre-treatment with an inhibitor of phosphatidylinositol 3-kinase or an activator of protein kinase C. Here, we show that cross-linking of the beta(2) (CD18) or beta(3) (CD61), but not beta(1) (CD 29) integrins in the presence of a low dose of formyl-Methionyl-Leucyl-Phenylalanine (fMLP) enables similar 8-s periodic RALS oscillations in suspended PMN in response to stimulation with two consecutive doses of chemoattractants. This effect did not appear to be due to increased surface expression of CD18 or CD61. RALS oscillations occurred in phase with 8-s oscillations in the stable F-actin pool and peaks in F-actin correlated with predominance of cells exhibiting a nascent lamella. Thus, simulation of surface attachment by CD18 and CD61 cross-linking after exposure to fMLP in suspended cells supports shape oscillations that are the result of actin-driven cyclic extension/retraction of nascent lamellae at the same frequency as the shape changes previously observed in crawling PMN.

Purpose: Imatinib mesylate transformed the treatment and paradigms of chronic myeloid leukemia. European LeukemiaNet (ELN) group has defined specific treatment milestones with an optimal outcome to be achieved in patients.

Methods: In a retrospective cohort study, we evaluated the impact of clinical and biological variables on achieving an optimal response at 6 and 12 months according to ELN recommendations. We included 106 patients with chronic phase chronic myeloid leukemia (CML) with appropriate bone marrow aspirate and biopsy for immunohistochemistry.

Results: The number of white blood cells (WBC), the percentage of peripheral blast, the values of Sokal and ELTS scores and the percentage of Ki-67+ cells in the bone marrow predicted a complete cytogenetic response (CCyR) at 6 months, but only WBC and EUTOS score predicts CCyR at 12 months. We found that Sokal score below 0.775 was very sensitive to achieve of CCyR at 6 months (m) and that all patients with a value <0.550 achieved CCyR-6m. Patients with a low percentage of blast in the peripheral blood (≤1.5%) or in the bone marrow (≤5%) together with lower WBC (≤100×109/L) were likely to have significantly higher CCyR rates at 6 and 12 months respectively. Also, patients with a higher number of Ki67+ cells in the leukemic areas of the bone marrow had a significantly better outcome. Unfortunately, our investigation did not reveal that bone marrow fibrosis (MF grade), microvascular density, percentage of CD34+, CD61+ or PTCH1+ cells could have any effect on achievement of CCyR at 6 or 12 months.

Conclusion: Our investigation has shown that only a few biological characteristics in patients with CML can predict the optimal treatment outcome after imatinib.

Background: Accidental hypothermia results in various dysfunctions in the human body. Additionally, coagulation disorder can lead to a life-threatening condition. We previously demonstrated that platelets stored in the spleen were activated and thus triggered coagulation disorder in a mouse model of hypothermia. In the present study, we wanted to investigate if this phenomenon in mice also occurs in humans as a reaction to hypothermia.

Methods: We analyzed splenic tissue collected from 22 deceased subjects who have died from hypothermia. These samples were compared with 22 control cases not exposed to cold environment. We performed immunohistochemical staining for CD61 (a marker of all platelets) and CD62P (a marker of activated platelets). We also evaluated the morphology of platelets in the spleen with scanning electron microscopy.

Results: Immunohistochemical analysis revealed no significant changes in the amounts of CD61-positive platelets between the hypothermia and control cases. However, the hypothermia cases contained abundant CD62P-positive platelets compared with those of the control cases. Immunohistochemical analysis also revealed that the activated platelets formed aggregates and adhered to splenic sinusoidal endothelial cells in the hypothermia cases. However, we observed no significant fibrin formation around the activated platelets.

Conclusions: Hypothermia resulted in splenic platelet activation, which may be used as a postmortem marker of hypothermia. The release of activated platelets from the spleen into to circulation upon rewarming may promote coagulation disturbances.

Keywords: Fibrin; Hypothermia; Platelet; Spleen; Splenic sinusoidal endothelial cell.

Objective: To explore the phenotypic and genetic characteristics of acute megakaryoblastic leukemia (AMKL) in young children accompany by WT1, MLL-PTD and EVI1, in order to improve the diagnosis level of AMKL.

Methods: EDTA-K₂ anticoagulation venous blood was collected for blood routine and morphological analysis of blood cells; bone marrow was extracted for cell morphology, immunophenotype, chromosome karyotype and fusion gene analysis.

Results: White blood cell count was 12.3× 10⁹/L, hemoglobin was 73 g/L, and platelet count was 13× 10⁹/L. The morphological analysis of blood cells showed that the size of immature cells was like that of primitive immature lymphocytes, which was circular or irregular and part of them with obvious pseudopodia. The cytoplasm is basophilic with heterogeneous coloration and granules. Nuclear chromatin is fine and even, 1-3 nucleoli can be seen, these immature cells account for about 40%; the morphology of bone marrow cells was consistent with acute leukemia, negative for peroxidase staining, negative for AS-DNCE staining and alpha-NBE staining. Flow cytometry results showed that the protocells account for about 52% and significant expression of megakaryocytes related markers (cCD41+, CD61+, CD36+). Chromosome karyotype is 46, XX, der(3) add(3)(p21)add(3)(q25), add (9)(q22), -13, +mar [4]/46, XX, del(13)(q12q22) [3]/46, XX[3]. The fusion gene WT1 was overexpressed, MLL-PTD and EVI1 were positive.

Conclusion: Acute megakaryocytic leukemia has unique and complex phenotypic and genetics characteristics.

Studies on platelet function in children older than neonatal period are few and their results are controversial. The pediatric platelets were alternatively reported to be more active or less active than adults' ones. We compared platelet function in the several age groups of children to adults and evaluated the age when platelet function reaches the adults' status. The study included 76 healthy children and 49 healthy adult volunteers. Types of platelet activation used included: collagen-related peptide (CRP) and PAR-1 activating peptide SFLLRN; SFLLRN, PAR-4 activating peptide AYPGKF and adenosine diphosphate (ADP); ADP. The parameters determined included forward (FSC) and side scatter (SSC), CD42b, CD61, CD62P, PAC-1, annexin V binding and mepacrine release levels. Resting pediatric platelets were similar to adults' platelets except for 1.2-fold decreased FSC and dense granules volume in youngest children, and 2.5-fold increased annexin V level in children aged 1-10 years. After CRP+SFLLRN stimulation, pediatric platelets had a 1.2-fold lower alpha- and 1.1-fold lower dense granule release than adults. For SFLLRN+AYPGKF+ADP stimulation, this was observed only for youngest children. The response to ADP stimulation was identical for pediatric platelets and adults. Pediatric platelets have lower granular release than adults' platelets, which persists until the age of 18.

Keywords: Flow cytometry; healthy children; platelet function.