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The pathophysiological mechanisms contributing to the decreased platelet count in immune thrombocytopenia (ITP) are not entirely understood. Here, we investigated the key step of proplatelet formation (PPF) by studying the effect of ITP plasma in thrombopoiesis. Normal cord blood-derived mature megakaryocytes were cultured in the presence of recalcified plasma from ITP patients, and PPF was evaluated by microscopic analysis. Patient samples induced a dose-dependent inhibition in PPF, as well as decreased complexity of proplatelet architecture. Although slightly increased, plasma-induced megakaryocyte apoptosis was not related to PPF impairment. Purified IgG reproduced the inhibitory effect, while platelet-adsorbed plasma induced its reversion, suggesting the involvement of auto-antibodies in the inhibition of thrombopoiesis. Impaired PPF, induced by ITP plasmas bearing anti-GPIIb-IIIa antibodies, was related to their ability to interfere with the normal function of this integrin, as assessed by megakaryocyte PAC-1 binding and β3 integrin phosphorylation while the presence of anti-glycoprotein Ia-IIa auto-antibodies was associated with loss of normal inhibition of PPF induced by type I collagen. In conclusion, abnormal thrombopoiesis comprising decreased PPF and morphological changes in proplatelet structure are induced by patient samples, unveiling new mechanisms contributing to decreased platelet count in ITP., (© 2014 John Wiley & Sons Ltd.)

This study investigated the involvement of chemokines including stromal derived factor 1 (SDF-1), interleukin 8 (IL-8), growth-related oncogene alpha (GRO-alpha) and their receptors, CXCR4, CXCR2 and CXCR1 in essential thrombocythemia (ET), a chronic myeloproliferative disease characterized by megakaryocytic hyperplasia and high platelet count. Fifty-three ET patients were studied. Plasma levels of SDF-1, IL-8 and GRO-alpha, evaluated by enzyme-linked immunosorbent assay, and flow cytometric analysis of CXCR1 and CXCR2 on the platelet membrane, were found to be normal in ET patients. CXCR4 expression on platelet surface as well as platelet CXCR4 mRNA detected by real-time reverse transcription polymerase chain reaction, were decreased. Platelet CXCR4 internalization rate was normal while SDF-1-induced platelet aggregation was delayed, decreased or absent. Immunohistochemical staining revealed that megakaryocytes were also affected. CXCR4 decrease was not observed either in peripheral white blood cells or in circulating CD34(+) precursors. These results show that CXCR4 is decreased in the megakaryocytic lineage in ET, mainly due to a reduced CXCR4 production, and an abnormal platelet response to SDF-1. This report is the first to describe platelet and megakaryocytic CXCR4 deficiency in a human disease and the presence of this abnormality in a megakaryocytic-related illness highlights the important role of SDF-1/CXCR4 axis in platelet development.

Plasmatic levels of PDGF-AB, TGFbeta1, and bFGF are increased in patients with essential thrombocythemia (ET) while intraplatelet levels are low for PDGF, normal for TGFbeta, and elevated for bFGF. To evaluate the contribution of gene expression to the dysregulated cytokine levels, we studied platelet PDGF-A, PDGF-B, TGFbeta1, and bFGF mRNA in ET patients before and during anagrelide treatment. We found decreased PDGF-A and PDGF-B, increased TGFbeta1, and normal bFGF mRNA levels. During treatment, mRNA levels remained decreased for PDGF-A, were increased for PDGF-B and normal for TGFbeta1. In untreated patients, protein expression of PDGF paralleled its mRNA levels while different patterns of RNA and protein were found for TGFbeta1 and bFGF., (Copyright 2005 Wiley-Liss, Inc.)

We studied 15 patients with essential thrombocythemia (ET) before treatment and after normalization of platelet count by anagrelide. Significantly increased plasma levels of PDGF, TGFbeta, and bFGF were found. Patients with mild reticulin fibrosis in bone marrow had higher PDGF levels. During treatment, plasma TGFbeta and bFGF levels remained elevated in most patients (P < 0.0001 and P < 0.01, respectively). Intraplatelet PDGF levels were low before treatment (P < 0.006) and normal on hematological remission, without relation with the presence or absence of reticulin fibrosis in bone marrow. Intraplatelet TGFbeta levels were normal regardless of the platelet count. Intraplatelet bFGF levels were raised before (P < 0.001) and during treatment (P < 0.01). By immunostaining, TGFbeta and bFGF were seen in megakaryocytes and lymphocytes with a similar pattern of intensity in patients and controls, suggesting that other cells might also contribute to the raised plasma values. We believe that the plasma increment of these cytokines suggests that they play a role in the pathogenesis of ET. The normal PDGF plasma level found during treatment may be in relation with the platelet count. However, the persistent increase of TGF-beta in plasma and bFGF both in plasma and platelets may indicate dysregulation of cytokine synthesis in TE., (Copyright 2002 Wiley-Liss, Inc.)

Argentine hemorrhagic fever (AHF) is a viral disease caused by Junin virus and characterized by hematologic and neurological involvement. The main hematologic features are leukopenia, thrombocytopenia, and bone marrow hypoplasia. Hematopoietic growth factors serum levels were measured by ELISA technique in forty-eight patients with confirmed diagnosis of AHF. Patients were classified according to the clinical picture in 15 severe (SCF), 17 moderate (MoCF), and 16 mild (MiCF) cases. Erythropoietin levels were decreased in 28 of 45 patients and raised in 4 SCF patients. Twenty-four of 38 patients had high G-CSF levels at admittance in accordance with clinical picture severity, while IL-3, GM-CSF, and TGF-beta were normal in most cases. A direct correlation was found between G-CSF and TNF-alpha levels. Thrombopoietin levels were found to be raised in 19 of 21 patients. In conclusion, the low levels of Epo may contribute to the severe bone marrow erythroblastopenia described in AHF patients, while G-CSF seems to be a marker of illness severity.