Figure 3.
Constitutive phosphorylation and nuclear translocation of STAT-5 in EBs generated ex vivo from PV patients. Levels of STAT-5 phosphorylation in cell extracts of EBs obtained in HEMA culture (Prol) from 1 ND and 3 PV patients and in cultures deprived of growth factor for 4 hours (GFD) and then treated with erythropoietin (EPO; 3 U/mL) and dexamethasone (DXM; 10−6M), alone or in combination, as indicated. STAT-5 phosphorylation was analyzed by WB of cell extracts immunoprecipitated (IP) with anti–STAT-5 antibody. The cell lysates were then analyzed by WB for β-actin and/or STAT-5 as quantitative control. The intensity of the signal was quantified by densitometry and expressed as a ratio (FI indicates fold increase) with the signal from cells in Prol. Similar data on STAT-5 phosphorylation of EBs from 10 additional NDs were reported previously.20