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3-Nitropropionic Acid Enhances Ferroptotic Cell Death via NOX2-Mediated ROS Generation in STHdhQ111 Striatal Cells
Noria Okada1, Shinsuke Nakamura1, Masamitsu Shimazawa1
1Molecular Pharmacology, Department of Biofunctional Evaluation, Gifu Pharmaceutical University.
Insights
Huntington's disease (HD) cell death is triggered by mitochondrial dysfunction and ferroptosis, not apoptosis. Inhibiting NADPH oxidase 2 (NOX2) significantly reduces this cell death, implicating NOX2 in HD pathogenesis.
Area of Science:
- Neuroscience
- Cell Biology
- Genetics
Background:
- Huntington's disease (HD) is a hereditary neurodegenerative disorder linked to CAG repeat expansion in the Huntingtin (HTT) gene.
- While CAG repeat length is a factor, disease onset and progression vary, suggesting other mechanisms contribute to pathogenesis.
- Mitochondrial complex II dysfunction is implicated in HD pathogenesis.
Purpose of the Study:
- To investigate the mechanism of cell death induced by 3-nitropropionic acid (3-NP), a mitochondrial complex II inhibitor, in Huntington's disease (HD) striatal cells.
- To determine if cell death is mediated by apoptosis, ferroptosis, or other pathways.
- To identify the role of reactive oxygen species (ROS) and NADPH oxidase 2 (NOX2) in 3-NP-induced cell death.
Main Methods:
- Utilized STHdhQ111 striatal cells (HD model) and STHdhQ7 cells (normal HTT).
- Treated cells with 3-nitropropionic acid (3-NP) to inhibit mitochondrial complex II.
- Assessed cell death using inhibitors of ferroptosis (Ferrostatin-1), iron chelation (deferoxamine), apoptosis (Z-VAD-FMK), NOX2 (GSK2795039), and mitochondrial ROS (Mito-TEMPO).
Main Results:
- 3-NP significantly increased cell death in HD striatal cells (STHdhQ111) compared to normal cells (STHdhQ7).
- Ferrostatin-1 and deferoxamine markedly inhibited 3-NP-induced cell death, suggesting ferroptosis.
- Cell death was caspase-independent and not abrogated by a caspase inhibitor, ruling out apoptosis.
- Inhibition of NOX2 significantly reduced 3-NP-induced cell death, indicating a major role for NOX2-derived superoxide anion (O2-).
- Mitochondrial-targeted superoxide dismutase mimetic partially inhibited cell death, suggesting partial mitochondrial involvement.
Conclusions:
- 3-NP-induced cell death in HD striatal cells is caspase-independent and non-apoptotic.
- Ferroptotic cell death is the primary mechanism, significantly mediated by NOX2 activation and subsequent superoxide production.
- These findings highlight NOX2 as a potential therapeutic target for Huntington's disease.
Abstract:
Huntington's disease (HD) is a hereditary neurodegenerative disease that involves an expansion of the CAG repeats of the Huntingtin (HTT) gene, but the disease onset and progression do not necessarily correspond to the extent of CAG repeats. Decreased mitochondrial complex II activity has also been reported to be closely associated with disease pathogenesis. Here, we examined the mechanism of cell death induced by 3-nitropropionic acid (3-NP), a mitochondrial complex II inhibitor, using striatal cells (STHdhQ111 cells) derived from HD model mice with mutant HTT carrying the CAG repeat extended. Treatment with 3-NP (5 mM) enhanced cell death in STHdhQ111 compared to STHdhQ7 cells with normal HTT. Ferrostatin-1, an inhibitor of ferroptosis, and deferoxamine, an iron chelator, markedly inhibited 3-NP-induced cell death in both the STHdh cell lines. On the other hands, cell death was not abrogated by a broad-spectrum caspase inhibitor, Z-VAD-FMK, indicating that this cell death was caspase-independent. Cell death caused by 3-NP is suggested to be due to ferroptosis. Furthermore, 3-NP-induced cell death was markedly inhibited by GSK2795039, a reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2) inhibitor, suggesting that cell death is mainly mediated by intracellular superoxide anion (O2-) production through NOX2. Furthermore, a mitochondria-targeted superoxide dismutase mimetic (Mito-TEMPO), partially inhibited 3-NP-induced cell death, suggesting that O2- production in the mitochondria is partially responsible for cell death. These results indicate that 3-NP-induced cell death in the STHdhQ111 cells is caspase-independent, non-apoptotic, and that ferroptotic cell death is mainly induced via NOX2 activation.
