ascorbic-acid and diphenyliodonium

Plants exhibit rapid, systemic signaling systems that allow them to coordinate physiological and developmental responses throughout the plant body, even to highly localized and quickly changing environmental stresses. The propagation of these signals is thought to include processes ranging from electrical and hydraulic networks to waves of reactive oxygen species (ROS) and cytoplasmic Ca(2+) traveling throughout the plant. For the Ca(2+) wave system, the involvement of the vacuolar ion channel TWO PORE CHANNEL1 (TPC1) has been reported. However, the precise role of this channel and the mechanism of cell-to-cell propagation of the wave have remained largely undefined. Here, we use the fire-diffuse-fire model to analyze the behavior of a Ca(2+) wave originating from Ca(2+) release involving the TPC1 channel in Arabidopsis (Arabidopsis thaliana). We conclude that a Ca(2+) diffusion-dominated calcium-induced calcium-release mechanism is insufficient to explain the observed wave transmission speeds. The addition of a ROS-triggered element, however, is able to quantitatively reproduce the observed transmission characteristics. The treatment of roots with the ROS scavenger ascorbate and the NADPH oxidase inhibitor diphenyliodonium and analysis of Ca(2+) wave propagation in the Arabidopsis respiratory burst oxidase homolog D (AtrbohD) knockout background all led to reductions in Ca(2+) wave transmission speeds consistent with this model. Furthermore, imaging of extracellular ROS production revealed a systemic spread of ROS release that is dependent on both AtRBOHD and TPC1 These results suggest that, in the root, plant systemic signaling is supported by a ROS-assisted calcium-induced calcium-release mechanism intimately involving ROS production by AtRBOHD and Ca(2+) release dependent on the vacuolar channel TPC1.

Topics: Arabidopsis; Arabidopsis Proteins; Ascorbic Acid; Biphenyl Compounds; Calcium Channels; Calcium Signaling; Gene Knockout Techniques; Models, Biological; Molecular Imaging; NADPH Oxidases; Onium Compounds; Plant Roots; Reactive Oxygen Species; Salt Tolerance; Serum Albumin, Bovine; Stress, Physiological

Glutathione (GSH) depletion is widely used to sensitize cells to anticancer treatment inducing the progression of programmed cell death and overcoming chemoresistance. It has been reported that neuroblastoma cells with MYCN amplification are unable to start TRAIL-dependent death and MYCN, in concert with cytotoxic drugs, efficiently induces the mitochondrial pathway of apoptosis through oxidative mechanisms. In this study, we show that GSH loss induced by L-buthionine-S,R-sulfoximine (BSO), an inhibitor of GSH biosynthesis, leads to overproduction of reactive oxygen species (ROS) and triggers apoptosis of MYCN-amplified neuroblastoma cells. BSO susceptibility of SK-N-BE-2C, a representative example of MYCN-amplified cells, has been attributed to stimulation of total SOD activity in the absence of changes in the level and the activity of catalase. Therefore, the unbalanced intracellular redox milieu has been demonstrated to be critical for the progression of neuroblastoma cell death that was efficiently prevented by antioxidants and rottlerin. These results describe a novel pathway of apoptosis dependent on ROS formation and PKC-delta activation and independent of p53, bcl-2, and bax levels; the selective redox modulation of PKC-delta might be suggested as a potential strategy for sensitizing MYCN-amplified cells to therapeutic approaches.

Topics: Acetophenones; Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Ascorbic Acid; Benzopyrans; Biphenyl Compounds; Buthionine Sulfoximine; Calpain; Caspases; Catalase; Cell Line, Tumor; DNA Damage; Glutathione; Humans; N-Myc Proto-Oncogene Protein; Neuroblastoma; Nuclear Proteins; Oncogene Proteins; Onium Compounds; Protein Kinase C; Reactive Oxygen Species; Superoxide Dismutase; Superoxide Dismutase-1

Expression of the multidrug resistance (MDR) transporter P-glycoprotein (P-gp) has been demonstrated to be regulated by hypoxia-inducible factor-1alpha (HIF-1alpha) and inhibited by intracellular reactive oxygen species (ROS). Herein, P-gp and HIF-1alpha expression were investigated in multicellular prostate tumor spheroids overexpressing the ROS-generating enzyme Nox-1 in comparison to the mother cell line DU-145. In Nox-1-overexpressing tumor spheroids (DU-145Nox1) generation of ROS as well as expression of Nox-1 was significantly increased as compared to DU-145 tumor spheroids. ROS generation was significantly inhibited in the presence of the NADPH-oxidase antagonists diphenylen-iodonium chloride (DPI) and 4-(2-aminoethyl)benzenesulfonyl fluoride (AEBSF). Albeit growth kinetic of DU-145Nox1 tumor spheroids was decreased as compared to DU-145 spheroids, elevated expression of Ki-67 was observed indicating increased cell cycle activity. In DU-145Nox1 tumor spheroids, expression of HIF-1alpha as well as P-gp was significantly decreased as compared to DU-145 spheroids, which resulted in an increased retention of the anticancer agent doxorubicin. Pretreatment with the free radical scavengers vitamin E and vitamin C increased the expression of P-gp as well as HIF-1alpha in Nox-1-overexpressing cells, whereas no effect of free radical scavengers was observed on mdr-1 mRNA expression. In summary, the data of the present study demonstrate that the development of P-gp-mediated MDR is abolished under conditions of elevated ROS levels, suggesting that the MDR phenotype can be circumvented by modest increase of intracellular ROS generation.

Topics: Ascorbic Acid; ATP Binding Cassette Transporter, Subfamily B, Member 1; Biphenyl Compounds; Cell Line, Tumor; Drug Resistance, Multiple; Free Radical Scavengers; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Multidrug Resistance-Associated Proteins; NADPH Oxidase 1; NADPH Oxidases; Onium Compounds; Prostatic Neoplasms; Reactive Oxygen Species; Spheroids, Cellular; Sulfones; Transcription Factors; Vitamin E