antimycin and Alzheimer-Disease

Mitochondrial dysfunction has been widely associated with the pathology of Alzheimer's disease, but there is no consensus on whether it is a cause or consequence of disease, nor on the precise mechanism(s). We addressed these issues by testing the effects of expressing the alternative oxidase AOX from Ciona intestinalis, in different models of AD pathology. AOX can restore respiratory electron flow when the cytochrome segment of the mitochondrial respiratory chain is inhibited, supporting ATP synthesis, maintaining cellular redox homeostasis and mitigating excess superoxide production at respiratory complexes I and III. In human HEK293-derived cells, AOX expression decreased the production of beta-amyloid peptide resulting from antimycin inhibition of respiratory complex III. Because hydrogen peroxide was neither a direct product nor substrate of AOX, the ability of AOX to mimic antioxidants in this assay must be indirect. In addition, AOX expression was able to partially alleviate the short lifespan of Drosophila models neuronally expressing human beta-amyloid peptides, whilst abrogating the induction of markers of oxidative stress. Our findings support the idea of respiratory chain dysfunction and excess ROS production as both an early step and as a pathologically meaningful target in Alzheimer's disease pathogenesis, supporting the concept of a mitochondrial vicious cycle underlying the disease.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Antimycin A; Antioxidants; Ciona intestinalis; Disease Models, Animal; Electron Transport Complex I; Electron Transport Complex IV; Gene Expression Regulation; HEK293 Cells; Humans; Hydrogen Peroxide; Mitochondria; Mitochondrial Proteins; Oxidative Stress; Oxidoreductases; Plant Proteins; Reactive Oxygen Species; Superoxides

Intracellular amyloid beta (Aβ) oligomers and extracellular Aβ plaques are key players in the progression of sporadic Alzheimer's disease (AD). Still, the molecular signals triggering Aβ production are largely unclear. We asked whether mitochondrion-derived reactive oxygen species (ROS) are sufficient to increase Aβ generation and thereby initiate a vicious cycle further impairing mitochondrial function.. Complex I and III dysfunction was induced in a cell model using the respiratory inhibitors rotenone and antimycin, resulting in mitochondrial dysfunction and enhanced ROS levels. Both treatments lead to elevated levels of Aβ. Presence of an antioxidant rescued mitochondrial function and reduced formation of Aβ, demonstrating that the observed effects depended on ROS. Conversely, cells overproducing Aβ showed impairment of mitochondrial function such as comprised mitochondrial respiration, strongly altered morphology, and reduced intracellular mobility of mitochondria. Again, the capability of these cells to generate Aβ was partly reduced by an antioxidant, indicating that Aβ formation was also ROS dependent. Moreover, mice with a genetic defect in complex I, or AD mice treated with a complex I inhibitor, showed enhanced Aβ levels in vivo.. We show for the first time that mitochondrion-derived ROS are sufficient to trigger Aβ production in vitro and in vivo.. Several lines of evidence show that mitochondrion-derived ROS result in enhanced amyloidogenic amyloid precursor protein processing, and that Aβ itself leads to mitochondrial dysfunction and increased ROS levels. We propose that starting from mitochondrial dysfunction a vicious cycle is triggered that contributes to the pathogenesis of sporadic AD.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Amyloid Precursor Protein Secretases; Animals; Antimycin A; Aspartic Acid Endopeptidases; Cell Line; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Humans; Mice; Mice, Mutant Strains; Microscopy, Confocal; Mitochondria; Reactive Oxygen Species; Rotenone

In Alzheimer's disease (AD), rod-like cofilin aggregates (cofilin-actin rods) and thread-like inclusions containing phosphorylated microtubule-associated protein (pMAP) tau form in the brain (neuropil threads), and the extent of their presence correlates with cognitive decline and disease progression. The assembly mechanism of these respective pathological lesions and the relationship between them is poorly understood, yet vital to understanding the causes of sporadic AD. We demonstrate that, during mitochondrial inhibition, activated actin-depolymerizing factor (ADF)/cofilin assemble into rods along processes of cultured primary neurons that recruit pMAP/tau and mimic neuropil threads. Fluorescence resonance energy transfer analysis revealed colocalization of cofilin-GFP (green fluorescent protein) and pMAP in rods, suggesting their close proximity within a cytoskeletal inclusion complex. The relationship between pMAP and cofilin-actin rods was further investigated using actin-modifying drugs and small interfering RNA knockdown of ADF/cofilin in primary neurons. The results suggest that activation of ADF/cofilin and generation of cofilin-actin rods is required for the subsequent recruitment of pMAP into the inclusions. Additionally, we were able to induce the formation of pMAP-positive ADF/cofilin rods by exposing cells to exogenous amyloid-beta (Abeta) peptides. These results reveal a common pathway for pMAP and cofilin accumulation in neuronal processes. The requirement of activated ADF/cofilin for the sequestration of pMAP suggests that neuropil thread structures in the AD brain may be initiated by elevated cofilin activation and F-actin bundling that can be caused by oxidative stress, mitochondrial dysfunction, or Abeta peptides, all suspected initiators of synaptic loss and neurodegeneration in AD.

Topics: Actin Depolymerizing Factors; Actins; Adenosine Triphosphate; Alzheimer Disease; Amino Acid Motifs; Amyloid beta-Peptides; Animals; Animals, Newborn; Antimycin A; Brain; Bridged Bicyclo Compounds, Heterocyclic; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Cells, Cultured; Chick Embryo; Depsipeptides; Enzyme Inhibitors; Fluorescence Resonance Energy Transfer; Green Fluorescent Proteins; Humans; Hydrogen Peroxide; Ionophores; Neurites; Neurons; Organ Culture Techniques; Oxidants; p21-Activated Kinases; Peptide Fragments; Phosphorylation; Rats; RNA, Small Interfering; Serine; tau Proteins; Thiazolidines; Transfection

There is a large body of literature indicating that aggregated amyloid-beta peptide (Abeta) is toxic to neurons and suggesting that this neurotoxicity represents the final common pathway for neuronal degeneration in Alzheimer's disease. Previous studies have shown the outgrowth of a subclone of the rat neuronal cell line PC12 that is resistant to the toxic effect of aggregated Abeta peptide if the parent cell line is grown in the presence of aggregated Abeta peptide for a number of passages [Behl, Davis, Lesley and Schubert (1994) Cell 77, 817-827; Boland, Behrens, Choi, Manias and Perlmutter (1996) J. Biol. Chem. 271, 18032-18044]. To begin to characterize the mechanism by which PC12 cells become resistant to the apoptotic effect of Abeta peptide, in the present study we examined whether the resistance was specific to aggregated peptides, specific to an apoptotic form of cell death, and specific in cell type or was a general resistance to cell death that could be elicited in diverse cell types. The results show that the resistance is specific to compounds that have apoptotic effects through the generation of hydroxyl radical or H2O2, including aggregated Abeta-(25-35), Abeta-(1-40), Abeta-(1-42), Abeta-(1-43), amylin, 6-hydroxydopamine and H2O2 itself. The resistant subclones of PC12 were not resistant to other forms of apoptotic cell death or to necrotic cell death. The resistant state was also identified in a human hepatoma cell line, HepG2, when it was grown in the presence of aggregated Abeta-(25-35) for several passages, indicating that the mechanism(s) or molecule(s) responsible for this resistance are not restricted to neuronal cells and may be relevant to the pathobiology of oxidative injury in other cell types.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Antimycin A; Apoptosis; Cell Line; Cytotoxins; DNA Fragmentation; Free Radical Scavengers; Humans; Hydrogen Peroxide; Hydroxyl Radical; Nerve Growth Factors; Oxidative Stress; Oxidopamine; Peptide Fragments; Rats