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Toxicity in rat primary neurons through the cellular oxidative stress induced by the turn formation at positions 22 and 23 of A?42.

ABSTRACT: The 42-mer amyloid ?-protein (A?42) aggregates to form soluble oligomers that cause memory loss and synaptotoxicity in Alzheimer's disease (AD). Oxidative stress is closely related to the pathogenesis of AD. We previously identified the toxic conformer of A?42 with a turn at positions 22 and 23 ("toxic turn") by solid-state NMR and demonstrated that a monoclonal antibody (11A1) against the toxic turn in A?42 mainly detected the oligomer in the brains of AD patients. Our recent study suggested that oxidative stress is a key factor of the oligomerization and cognitive impairment induced by A? overproduction in vivo. However, the involvement of the toxic conformer in A?42-induced oxidative damage remains unclear. To investigate this mechanism, we examined the levels of intracellular reactive oxygen species (ROS) and neurotoxicity in rat primary neurons using E22P-A?42, a mutant that induces a turn at positions 22 and 23, and E22V-A?42, a turn-preventing mutant. E22P-A?42, but not E22V-A?42, induced greater ROS production than Wt-A?42 in addition to potent neurotoxicity. Interestingly, the formation of the toxic conformer in both E22P-A?42 and Wt-A?42 probed by the 11A1 antibody preceded A?42-induced neurotoxicity. Trolox (a radical scavenger) and Congo red (an aggregation inhibitor) significantly prevented the neurotoxicity and intracellular ROS induced by E22P-A?42 and Wt-A?42, respectively. These results suggest that A?42-mediated toxicity is caused by the turn that favors toxic oligomers, which increase generation of ROS.

SUBMITTER: Izuo N

PROVIDER: S-EPMC3447395 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature

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Toxicity in rat primary neurons through the cellular oxidative stress induced by the turn formation at positions 22 and 23 of Aβ42.

Izuo Naotaka N Kume Toshiaki T Sato Mizuho M Murakami Kazuma K Irie Kazuhiro K Izumi Yasuhiko Y Akaike Akinori A

ACS chemical neuroscience 20120606 9

The 42-mer amyloid β-protein (Aβ42) aggregates to form soluble oligomers that cause memory loss and synaptotoxicity in Alzheimer's disease (AD). Oxidative stress is closely related to the pathogenesis of AD. We previously identified the toxic conformer of Aβ42 with a turn at positions 22 and 23 ("toxic turn") by solid-state NMR and demonstrated that a monoclonal antibody (11A1) against the toxic turn in Aβ42 mainly detected the oligomer in the brains of AD patients. Our recent study suggested th ...[more]

PMID: 23019494

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Project description:Ferroptosis is a type of programmed cell death dependent on iron. It is different from other forms of cell death such as apoptosis, classic necrosis and autophagy. Ferroptosis is involved in many neurodegenerative diseases. The role of ferroptosis in glutamate-induced neuronal toxicity is not fully understood. To test its toxicity, glutamate (1.25-20 mM) was applied to HT-22 cells for 12 to 48 hours. The optimal experimental conditions occurred at 12 hours after incubation with 5 mM glutamate. Cells were cultured with 3-12 ?M ferrostatin-1, an inhibitor of ferroptosis, for 12 hours before exposure to glutamate. The cell viability was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Autophagy was determined by monodansylcadaverine staining and apoptosis by caspase 3 activity. Damage to cell structures was observed under light and by transmission electron microscopy. The release of lactate dehydrogenase was detected by the commercial kit. Reactive oxygen species were measured by flow cytometry. Glutathione peroxidase activity, superoxide dismutase activity and malondialdehyde level were detected by the appropriate commercial kit. Prostaglandin peroxidase synthase 2 and glutathione peroxidase 4 gene expression was detected by real-time quantitative polymerase chain reaction. Glutathione peroxidase 4 and nuclear factor erythroid-derived-like 2 protein expression was detected by western blot analysis. Results showed that ferrostatin-1 can significantly counter the effects of glutamate on HT-22 cells, improving the survival rate, reducing the release of lactate dehydrogenase and reducing the damage to mitochondrial ultrastructure. However, it did not affect the caspase-3 expression and monodansylcadaverine-positive staining in glutamate-injured HT-22 cells. Ferrostatin-1 reduced the levels of reactive oxygen species and malondialdehyde and enhanced superoxide dismutase activity. It decreased gene expression of prostaglandin peroxidase synthase 2 and increased gene expression of glutathione peroxidase 4 and protein expressions of glutathione peroxidase 4 and nuclear factor (erythroid-derived)-like 2 in glutamate-injured HT-22 cells. Treatment of cultured cells with the apoptosis inhibitor Z-Val-Ala-Asp (OMe)-fluoromethyl ketone (2-8 ?M), autophagy inhibitor 3-methyladenine (100-400 ?M) or necrosis inhibitor necrostatin-1 (10-40 ?M) had no effect on glutamate induced cell damage. However, the iron chelator deferoxamine mesylate salt inhibited glutamate induced cell death. Thus, the results suggested that ferroptosis is caused by glutamate-induced toxicity and that ferrostatin-1 protects HT-22 cells from glutamate-induced oxidative toxicity by inhibiting the oxidative stress.

Dataset Information

Toxicity in rat primary neurons through the cellular oxidative stress induced by the turn formation at positions 22 and 23 of A?42.

Publications

Toxicity in rat primary neurons through the cellular oxidative stress induced by the turn formation at positions 22 and 23 of Aβ42.

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