title: Neuronal nitric oxide synthase is a key factor in doxorubicin-induced toxicity to rat-isolated cortical neurons.
abstract: Doxorubicin (DOX) is neurotoxic to serum-free cultures of rat cortical neurons in a biphasic concentration manner. For concentrations up to 0.5 μM, cell death follows an apoptotic pattern, while for higher concentrations apoptosis is inhibited and necrosis becomes dominant. Considering the potential toxic effects of DOX resulting from its redox-cycling, in this study we investigated the generation of reactive species and subsequent oxidative stress effects, formation of quinoproteins, activation of NF-kB, depletion of energy levels and consequent cell death in cultures of primary rat cortical cells challenged with this antitumour drug. The influence of neuronal nitric oxide synthase (nNOS) on DOX-induced neuronal cell damage was subsequently evaluated. The exposure of rat cortical primary cell cultures to DOX resulted in a significant generation of ROS/RNS, activation of NF-kB, depletion of GSH levels, depletion of ATP, and cell death, in a concentration biphasic manner. Doxorubicin also significantly increased protein-bound quinone products in neurons in a concentration-dependent manner. Inhibition of nNOS decreased neuronal cell death induced by DOX in a significant way, at the first phase of the biphasic curve. In conclusion, this study shows, for the first time, a clear involvement of nNOS and subsequent ROS/RNS generation as crucial signalling mediators of DOX-induced neurotoxicity on isolated cortical neurons. Inhibition of ROS/RNS formation, modulation of NOS isoforms and modulation of NF-kB activation could be of beneficial in preventing damage in the CNS caused by DOX.
authors.affiliation: REQUIMTE, Faculty of Pharmacy, Toxicology Department, University of Porto, Rua Aníbal Cunha 164, 4050-047 Porto, Portugal. miguelaclopes@yahoo.de
authors.forename: Miguel Angelo
authors.initials: MA
authors.lastname: Lopes
authors.forename: Andreas
authors.initials: A
authors.lastname: Meisel
authors.forename: Félix Dias
authors.initials: FD
authors.lastname: Carvalho
authors.forename: Maria de Lourdes
authors.initials: Mde L
authors.lastname: Bastos
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects / Q000201:enzymology*; D004317:Doxorubicin / Q000633:toxicity*; D009474:Neurons / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D052248:Nitric Oxide Synthase Type I / Q000502:physiology*; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology
chemical_list: D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D004317:Doxorubicin; D052248:Nitric Oxide Synthase Type I