Nitrogen monoxide (NO) and glucose - Unexpected links between energy metabolism and NO-mediated iron mobilization from cells

Nitrogen monoxide (NO) affects cellular iron metabolism due to its high aff inity for this metal ion. Indeed, NO has been shown to increase the mRNA bi nding activity of the iron-regulatory protein 1, which is a major regulator of iron homeostasis, Recently, we have shown that NO generators increase F e-59 efflux from cells pre-labeled with Fe-59-transferrin (Wardrop, S. L., Watts, R. N., and Richardson, D. R. (2000) Biochemistry 39, 2748-2758), The mechanism involved in this process remains unknown, and in this investigat ion we demonstrate that it is potentiated upon adding D-glucose (D-Glc) to the reincubation medium. In D-Glc-free or D-Glc-containing media, 5.6 and 1 6.5% of cellular 59Fe was released, respectively, in the presence of S-nitr osoglutathione. This difference in 59Fe release was observed with a variety of NO generators and cell types and was not due to a change in cell viabil ity. Kinetic studies showed that D-Glc had no effect on the rate of NO prod uction by NO generators. Moreover, only the metabolizable monosaccharides D -Glc and D-mannose could stimulate NO-mediated Fe-59 mobilization, whereas other sugars not easily metabolized by fibroblasts had no effect. Hence, me tabolism of the monosaccharides was essential to increase NO-mediated Fe-59 release. Incubation of cells with the citric acid cycle intermediates, cit rate and pyruvate, did not enhance NO-mediated Fe-59 release. Significantly , preincubation with the GSH-depleting agents, L-buthionine-[S,R]-sulfoximi ne or diethyl maleate, prevented NO-mediated Fe-59 mobilization, This effec t was reversed by incubating cells with N-acetyl-L-cysteine that reconstitu tes GSH. These results indicate that GSH levels are essential for NO-mediat ed Fe-59 efflux. Hence, D-Glc metabolism via the hexose monophosphate shunt resulting in the generation of GSH may be essential for NO-mediated Fe-59 release. These results have important implications for intracellular signal ing by NO and also NO-mediated cytotoxicity of activated macrophages that i s due, in part, to iron release from tumor target cells.