title: Kaempferol blocks oxidative stress in cerebellar granule cells and reveals a key role for reactive oxygen species production at the plasma membrane in the commitment to apoptosis.
abstract: Micromolar concentrations of the flavonoid kaempferol were found to efficiently block cerebellar granule cell (CGC) death through low K+-induced apoptosis, as demonstrated by prevention of the activation of caspase-3, internucleosomal DNA fragmentation, and chromatin condensation, without a significant rise in intracellular free Ca2+ concentration. Half of the maximum protection against CGC apoptosis was attained with 8 +/- 2 microM kaempferol. Reactive oxygen species (ROS) were monitored with 2',7'-dichlorodihydrofluorescein diacetate. Quantitative analysis of intracellularly and extracellularly oriented ROS production up to 3 h from the onset of low K+-induced CGC apoptosis was carried out with acquired digital fluorescence microscopy images of CGC in culture plates using a CCD camera, and also with fluorescence measurements of resuspended CGCs. In both cases, nearly 90% of ROS production by CGCs during the early stages (up to 3 h) after induction of low-K+ apoptosis occurs at the plasma membrane. Kaempferol, at concentrations that blocked CGC apoptosis, has been found to be a particularly potent blocker of extracellularly oriented ROS production by CGCs, and to inhibit the ascorbate-dependent NADH oxidase and superoxide anion production activities of the neuronal plasma membrane redox chain.
authors.affiliation: Departamento de Bioquímica y Biología Molecular y Genética, Facultad de Ciencias and Escuela de Ingenierías Agrarias, Universidad de Extremadura, Badajoz, Spain.
authors.forename: Alejandro Khalil
authors.initials: AK
authors.lastname: Samhan-Arias
authors.forename: Francisco Javier
authors.initials: FJ
authors.lastname: Martín-Romero
authors.forename: Carlos
authors.initials: C
authors.lastname: Gutiérrez-Merino
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002118:Calcium / Q000502:physiology; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D002470:Cell Survival / Q000187:drug effects; D002531:Cerebellum / Q000502:physiology*; D044949:Kaempferols / Q000494:pharmacology*; D007700:Kinetics; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D011189:Potassium Chloride / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism*
chemical_list: D044949:Kaempferols; D017382:Reactive Oxygen Species; D011189:Potassium Chloride; C006552:kaempferol; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; D002118:Calcium