In Vitro Megakaryocyte Differentiation and Proplatelet Formation in Ph-Negative Classical Myeloproliferative Neoplasms: Distinct Patterns in the Different Clinical Phenotypes

Background: Ph-negative myeloproliferative neoplasms (MPNs) are clonal disorders that include primary myelofibrosis (PMF), polycythemia vera (PV) and essential thrombocythemia (ET). Although the pathogenesis of MPNs is still incompletely understood, an involvement of the megakaryocyte lineage is a distinctive feature. Methodology/Principal Findings: We analyzed the in vitro megakaryocyte differentiation and proplatelet formation in 30 PMF, 8 ET, 8 PV patients, and 17 healthy controls (CTRL). Megakaryocytes were differentiated from peripheral blood CD34+ or CD45+ cells in the presence of thrombopoietin. Megakaryocyte output was higher in MPN patients than in CTRL with no correlation with the JAK2 V617F mutation. PMF-derived megakaryocytes displayed nuclei with a bulbous appearance, were smaller than ET- or PV-derived megakaryocytes and formed proplatelets that presented several structural alterations. In contrast, ET- and PV-derived megakaryocytes produced more proplatelets with a striking increase in bifurcations and tips compared to both control and PMF. Proplatelets formation was correlated with platelet counts in patient peripheral blood. Patients with pre-fibrotic PMF had a pattern of megakaryocyte proliferation and proplatelet formation that was similar to that of fibrotic PMF and different from that of ET. Conclusions/Significance: In conclusion, MPNs are associated with high megakaryocyte proliferative potential. Profound differences in megakaryocyte morphology and proplatelet formation distinguish PMF, both fibrotic and prefibrotic, from ET and PV

Recent advances in the bcr-abl negative chronic myeloproliferative diseases

The chronic myeloproliferative disorders are clonal hematopoietic stem cell disorders of unknown etiology. In one of these (chronic myeloid leukemia), there is an associated pathognomonic chromosomal abnormality known as the Philadelphia chromosome. This leads to constitutive tyrosine kinase activity which is responsible for the disease and is used as a target for effective therapy. This review concentrates on the search in the other conditions (polycythemia vera, essential thrombocythemia and idiopathic mylofibrosis) for a similar biological marker with therapeutic potential. There is no obvious chromosomal marker in these conditions and yet evidence of clonality can be obtained in females by the use of X-inactivation patterns. PRV-1mRNA over expression, raised vitamin B(12 )levels and raised neutrophil alkaline phosphatase scores are evidence that cells in these conditions have received excessive signals for proliferation, maturation and reduced apoptosis. The ability of erythroid colonies to grow spontaneously without added external erythropoietin in some cases, provided a useful marker and a clue to this abnormal signaling. In the past year several important discoveries have been made which go a long way in elucidating the involved pathways. The recently discovered JAK2 V617F mutation which occurs in the majority of cases of polycythemia vera and in about half of the cases with the two other conditions, enables constitutive tyrosine kinase activity without the need for ligand binding to hematopoietic receptors. This mutation has become the biological marker for these conditions and has spurred the development of a specific therapy to neutralize its effects. The realization that inherited mutations in the thrombopoietin receptor (c-Mpl) can cause a phenotype of thrombocytosis such as in Mpl Baltimore (K39N) and in a Japanese family with S505A, has prompted the search for acquired mutations in this receptor in chronic myeloproliferative disease. Recently, two mutations have been found; W515L and W515K. These mutations have been evident in patients with essential thrombocythemia and idiopathic myelofibrosis but not in polycythemia vera. They presumably act by causing constitutional, activating conformational changes in the receptor. The discovery of JAK2 and Mpl mutations is leading to rapid advancements in understanding the pathophysiology and in the treatment of these diseases