Changes in signal transduction downstream from the granulocyte-macrophage colony-stimulating factor receptor during differentiation of primary hemopoietic cells

Granulocyte-macrophage colony-stimulating factor (GMCSF) is a multifuctiona l cytokine, having different effects on primitive hemopoietic cells and ter minally differentiated end-cells of the myeloid lineage, Human primitive he mopoietic cells (CD34(+)) were obtained from the peripheral blood after mob ilization and induced to proliferate and then differentiate with a combinat ion of cytokines in vitro. Cells at different time points were then used to analyze the expression of the GM-CSF receptor and GM-CSF mediated activati on of the JAK 2-STAT 5 and MAP kinase pathways. Scatchard analysis as measu red by radioligand binding revealed that freshly purified CD34(+) cells exp ressed 36 +/- 1 high affinity receptors per cell (mean +/- SE, n = 3) and t he level of expression was not significantly different after 3 days in cult ure, but rose five- to tenfold by day 8, The day 0 CD34(+) cells were hypor esponsive to GM-CSF, but by 3 days in culture the cells were still morpholo gically immature but were actively proliferating and exhibited maximal GM-C SF induced JAK 2-STAT 5 and MAP kinase activation at the optimal time point . Further culture of the CD34(+) cells resulted in myeloid differentiation associated with prolongation of MAP kinase activation but not JAK 2-STAT 5 activation. These data indicate that the JAK 2-STAT 5 and MAP kinase pathwa ys are independently regulated and that changes in these signaling pathways occur with differentiation. (C) 1999 International Society for Experimenta l Hematology, Published by Elsevier Science Inc.