Evaluation of Mitochondrial Oxidative Stress in the Brain of a Transgenic Mouse Model of Alzheimer's Disease by in vitro Electron Paramagnetic Resonance Spectroscopy

Alzheimer's disease (AD) is one of the most common neurodegenerative diseases responsible for progressive dementia. Deposition of amyloid-beta (A beta) in the brain is the most important pathophysiological hallmark of AD. In addition, recent evidence indicates that reactive oxygen species (ROS) derived from mitochondria contribute to progression of AD pathology. We thus hypothesized that A beta accumulates and oxidative stress increases in the brain mitochondria of a transgenic mouse model of AD (APdE9). We measured the quantity of A beta and the activity of the antioxidant enzyme superoxide dismutase (SOD) in brain mitochondrial fractions prepared from APdE9 and wild-type (WT) mice aged 6, 9, 15, and 18 months. We also quantified the age-related changes in redox status in the mitochondrial fractions obtained from both APdE9 and WT mouse brains by electron paramagnetic resonance (EPR) spectrometry using a paramagnetic nitroxide Mito-Tempo [(2-(2,2,6,6-Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl) triphenylphosphonium chloride monohydrate] as a mitochondria-targeted redox-sensitive probe. In APdE9 mice, A beta accumulated in brain mitochondria earlier than in the non-mitochondrial fraction of the brain. Furthermore, increased oxidative stress was demonstrated in brain mitochondria of APdE9 mice by in vitro SOD assay as well as EPR spectroscopy. EPR combined with a mitochondria-targeted redox-sensitive nitroxide probe is a potentially powerful tool to elucidate the etiology of AD and facilitate the development of new therapeutic strategies for AD.