Increased megakaryocytopoiesis in Lyn-deficient mice

Incomplete restoration of Mpl expression in the mpl−/− mouse produces partial correction of the stem cell–repopulating defect and paradoxical thrombocytosis

“…31 Simultaneously, unstained cells were harvested by low-speed centrifugation (400 rpm for 10 minutes) and purified by unit gravity sedimentation on a discontinuous bovine serum albumin (BSA) density gradient for RNA isolation and cell lysis for Western blot analysis. 30 …”

“…Liquid culture megakaryocyte progenitors were generated by culturing bone marrow mononuclear cells in IMDM supplemented with 1% Nutridoma-SP (Roche Molecular Biochemicals), 2 mM L-glutamine, and 37.5 ng/mL mTPO as previously described. 30 After 72 hours in culture (37°C; 5% CO 2 , humidified incubator) megakaryocyte lineage differentiation and cell ploidy was evaluated by flow cytometry by staining with anti-CD41-FITC (BD Pharmingen) and propidium iodide (Sigma-Aldrich, St Louis, MO) as described. 31 Simultaneously, unstained cells were harvested by low-speed centrifugation (400 rpm for 10 minutes) and purified by unit gravity sedimentation on a discontinuous bovine serum albumin (BSA) density gradient for RNA isolation and cell lysis for Western blot analysis.…”

Deficiency of Src family kinases compromises the repopulating ability of hematopoietic stem cells

“…Examples of molecules that are activated in response to hematopoietic growth factors include, but are not limited to, nonreceptor tyrosine kinases, such as Src family kinases (SFKs). SFKs function in signal transduction from growth factor receptors for granulocyte macrophage colony-stimulating factor (GM-CSF), interleukin (IL)-3, IL-5, stem cell factor (SCF), erythropoietin, M-CSF, G-CSF, thrombopoietin, and Flt3 [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16]. Although diverse cytokine receptors activate SFKs in response to ligand binding, most of the data thus far has been derived by studying cell-line models using dominant negative and activating approaches.…”

Opposite expression pattern of Src kinase Lyn in acute and chronic haematological malignancies

“…deficiency or abundance, might interfere with normal cellular growth and function and potentially leads to distinct haematological phenotypes. Recent data on aberrant megakaryopoiesis in Lyn deficient cells and mice [6,8] attracted our intention, because abnormal megakaryocyte proliferation is also the predominant feature of Ph − MPN. The entirety of Ph − MPN under investigation showed a constitutive Lyn mRNA expression in total cellularity compared to control cases independent of an underlying mutation in JAK2 or MPL.…”

“…The animals showed a massive splenomegaly, myeloid lineage neoplasia and, with rising age, an increase of myeloid and erythroid progenitors. In a related model, Lyn −/− mice showed a prominent expansion of megakaryopoiesis in the bone marrow, an increase in platelet counts, and a severe splenomegaly [6]. Moreover, Lyn −/− megakaryocytes strikingly resembled features demonstrable in Philadelphia-chromosome-negative chronic myeloproliferative neoplasms (Ph − MPN) such as primary myelofibrosis (PMF) [7] including high ploidy (>8N), a significant increase of colony-forming units-megakaryocyte, and large colonies of clustered megakaryocytes phenotypically similar to bone marrows in Ph − MPN [6,8].…”