Nitric Oxide Enhances Cytotoxicity of Lead by Modulating the Generation of Reactive Oxygen Species and Is Involved in the Regulation of Pb(2+) and Ca(2+) Fluxes in Tobacco BY-2 Cells

Lead is a heavy metal known to be toxic to both animals and plants. Nitric oxide (NO) was reported to participate in plant responses to different heavy metal stresses. In this study, we analyzed the function of exogenous and endogenous NO in Pb-induced toxicity in tobacco BY-2 cells, focusing on the role of NO in the generation of reactive oxygen species (ROS) as well as Pb(2+) and Ca(2+) fluxes using non-invasive micro-test technology (NMT). Pb treatment induced BY-2 cell death and rapid NO and ROS generation, while NO burst occurred earlier than ROS accumulation. The elimination of NO by 2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO) resulted in a decrease of ROS, and the supplementation of NO by sodium nitroprusside (SNP) caused an increased accumulation of ROS. Furthermore, the addition of exogenous NO stimulated Pb(2+) influx, thus promoting Pb uptake in cells and aggravating Pb-induced toxicity in cells, whereas the removal of endogenous NO produced the opposite effect. Moreover, we also found that both exogenous and endogenous NO enhanced Pb-induced Ca(2+) effluxes and calcium homeostasis disorder. These results suggest that exogenous and endogenous NO played a critical regulatory role in BY-2 cell death induced by Pb stress by promoting Pb(2+) influx and accumulation and disturbing calcium homeostasis.