THE INFLUENCE OF NICKEL AND COBALT ON PUTATIVE MEMBERS OF THE OXYGEN-SENSING PATHWAY OF ERYTHROPOIETIN-PRODUCING HEPG2 CELLS

Cobalt and nickel stimulate, as does hypoxia, the production of erythr opoietin (EPO) in HepG2 cells. Under hypoxic conditions, a decrease in the level of intracellular reactive oxygen species (ROS) is thought t o stimulate EPO expression. Cobalt and nickel may interact with the pu tative oxygen sensor by changing the redox state of the central iron a tom of heme proteins, similar to the effects of hypoxia. It was invest igated, therefore, whether cobalt and nickel interact with hemeprotein s or ROS scavenging systems in the control of intracellular ROS level. Cobalt chloride (100 mu M, 24 h) oxidized non respiratory as well res piratory hemeproteins and increased the oxygen consumption. In contras t, nickel chloride (300 mu M, 24 h) primarily reduced respiratory heme proteins and decreased the oxygen consumption. In HepG2 cells treated with CoCl2, iron and cobalt were localized in cytosolic granules close to the cell nucleus and in mitochondria at concentrations up to 22 mM or 41 mM, respectively. Intracellular nickel was not measurable. Thre e-dimensional reconstruction of confocal laser microscopy images revea led hot spots of hy droxyl radical generation by a Fenton reaction at the sites of cytosolic iron accumulation. The (OH)-O-. levels decrease d in cobalt-treated (to 81%) as well as in nickel-treated (to 67%) Hep G2 cells, accompanied by an increase of EPO expression to 167% and 150 %, respectively. Our results underline the importance of (OH)-O-. form ed by a Fenton reaction for triggerimg EPO production. Identification of the primary hemeprotein being the oxygen sensor was not possible du e to the antagonistic effects of cobalt and nickel on the redox state of detectable hemeproteins.