title: Inhibition of autophagy by 3-methyladenine protects 1321N1 astrocytoma cells against pyocyanin- and 1-hydroxyphenazine-induced toxicity.
abstract: Central nervous system (CNS) infections due to Pseudomonas aeruginosa are difficult to treat and have a high mortality rate. Pyocyanin, a virulence factor produced by P. aeruginosa, has been shown to be responsible for the majority of P. aeruginosa's pathogenicity in mammalian cells. Several lines of evidence in respiratory cells suggest that this damage is primarily mediated by pyocyanin's ability to generate ROS and deplete host antioxidant defense mechanisms. However, it has yet to be established whether pyocyanin or 1-hydroxyphenazine have potential toxicity to the CNS. Therefore, the aim of this study was to compare the CNS toxicity of pyocyanin and 1-hydroxyphenazine in vitro and to provide insight into mechanisms that underlie this toxicity using 1321N1 astrocytoma cells. To achieve this, we investigated the contribution of oxidative stress and other mediators of cell death including autophagy, senescence and apoptosis. We show that oxidative stress is not a primary mediator of pyocyanin (0-100 μM) and 1-hydroxyphenazine (0-100 μM) induced toxicity in 1321N1 cells. Instead, our results suggest that autophagy may play a central role. The autophagy inhibitor 3-methyladenine (5 mM) protected 1321N1 astrocytoma cells against both pyocyanin and 1-hydroxyphenazine-induced cell injury and increased accumulation of acidic vesicular organelles, a hallmark of autophagy. Furthermore, apoptosis and senescence events may be secondary to autophagy in pyocyanin and 1-hydroxyphenazine-mediated cell injury. In conclusion, this study provides the first evidence on mechanisms underlying the toxicity of both pyocyanin and 1-hydroxyphenazine to astrocytoma cells and provides novel evidence suggesting that this toxicity may be mediated by the formation of acidic vesicular organelles, a hallmark of autophagic cell death.
authors.affiliation: Griffith Health Institute, Griffith University, Gold Coast, QLD, Australia.
authors.forename: Amelia J
authors.initials: AJ
authors.lastname: McFarland
authors.forename: Shailendra
authors.initials: S
authors.lastname: Anoopkumar-Dukie
authors.forename: Anthony V
authors.initials: AV
authors.lastname: Perkins
authors.forename: Andrew K
authors.initials: AK
authors.lastname: Davey
authors.forename: Gary D
authors.initials: GD
authors.lastname: Grant
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000975:Antioxidants / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D001254:Astrocytoma; D001343:Autophagy / Q000187:drug effects; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000187:drug effects; D004347:Drug Interactions; D006801:Humans; D018384:Oxidative Stress / Q000187:drug effects; D010619:Phenazines / Q000633:toxicity*; D011710:Pyocyanine / Q000633:toxicity*; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000975:Antioxidants; D010619:Phenazines; D017382:Reactive Oxygen Species; C025946:3-methyladenine; C050093:1-hydroxyphenazine; D011710:Pyocyanine; D000225:Adenine