Signal pathways in ouabain-induced proliferation of leukemia cells

Abstract

Cardiotonic steroids (CTSs) can bind to Na(+)/K(+)-ATPase and activate protein kinase cascades, resulting in changes in cell proliferation, differentiation or apoptosis in a cell-specific manner. We explored the participation of ouabain-activated signaling pathways in growth regulation of leukemia cells. Lymphocytic leukemia Jhhan cells and megakaryocytic leukemia M07e cells were incubated at different concentrations of ouabain (0, 1 and 10 nmol) for 24 hours. Cell proliferation was measured by methyl thiazolyl tetrazolium (MTT) assay. To probe the role of ouabain-induced signaling in control of cell growth, we employed Src kinase inhibitor PP2 and the MEK inhibitor PD98059, respectively. The expression of Na(+)/K(+)-ATPase alpha1 subunit of leukemia cells was evaluated by RT-PCR and Western blotting. One nmol and 10 nmol ouabain promoted proliferation of both Jhhan and M07e cells. Ouabain also up-regulated the expression of Na(+)/K(+)-ATPase alpha1 subunit. Addition of either PP2 or PD98059 blocked the effects of ouabain on cell proliferation. Ouabain activates Src and ERK1/2 pathways and regulates the proliferation of leukemia cells.