Mitochondrial dysfunction due to mutant copper/zinc superoxide dismutase associated with amyotrophic lateral
Simone Beretta1, Gessica Sala, Laura Mattavelli
1Department of Neuroscience and Biomedical Technologies, University of Milano-Bicocca, San Gerardo Hospital, via Donizetti, 106, 20052, Monza (MI), Italy.
Abstract:
We report that the expression of mutant G93A copper/zinc superoxide dismutase (SOD1), associated with familial amyotrophic lateral sclerosis, specifically causes a decrease in MTT reduction rate and ATP levels and an increase in both cytosolic and mitochondrial reactive oxygen species (ROS) production in human neuroblastoma SH-SY5Y cells compared to cells overexpressing wild-type SOD1 and untransfected cells. Exposure to N-acetylcysteine lowers ROS production and returns mitochondrial functional assays to control levels. No large aggregates of human SOD1 are detectable under basal growth conditions in any of the investigated cell lines. After proteasome activity inhibition, SOD1 aggregates can be detected exclusively in G93A-SOD1 cells, even though they do not per se enhance cell death compared to control cell lines. Our findings indicate that mitochondrial homeostasis is affected by mutant SOD1-generated ROS independently from the formation of aggregates and that this alteration is reversed by antioxidants.
Insights
Mutant G93A SOD1 expression in neuroblastoma cells disrupts mitochondrial function and increases reactive oxygen species (ROS). Antioxidant treatment with N-acetylcysteine reverses these effects, independent of SOD1 aggregate formation.
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- Familial amyotrophic lateral sclerosis (ALS) is linked to mutations in copper/zinc superoxide dismutase (SOD1).
- Understanding the cellular mechanisms of mutant SOD1 toxicity is crucial for ALS research.
Purpose of the Study:
- To investigate the impact of mutant G93A SOD1 on mitochondrial function and reactive oxygen species (ROS) production in human neuroblastoma cells.
- To determine the role of SOD1 aggregation in mutant SOD1-induced cellular dysfunction.
- To assess the therapeutic potential of antioxidants in mitigating mutant SOD1 toxicity.
Main Methods:
- Overexpression of wild-type and mutant G93A SOD1 in human neuroblastoma SH-SY5Y cells.
- Assessment of MTT reduction rate and ATP levels as indicators of mitochondrial function.
- Measurement of cytosolic and mitochondrial ROS production.
- Analysis of SOD1 aggregation following proteasome inhibition.
- Treatment with N-acetylcysteine (antioxidant) to evaluate its effects on ROS and mitochondrial function.
Main Results:
- Mutant G93A SOD1 expression significantly decreased MTT reduction rate and ATP levels, indicating impaired mitochondrial function.
- Increased cytosolic and mitochondrial ROS production was observed in cells expressing G93A SOD1 compared to controls.
- N-acetylcysteine treatment reduced ROS levels and restored mitochondrial function to control levels.
- SOD1 aggregates were detected only in G93A-SOD1 cells after proteasome inhibition, but did not directly correlate with cell death.
- Mitochondrial dysfunction was linked to ROS generated by mutant SOD1, irrespective of aggregate formation.
Conclusions:
- Mutant G93A SOD1 disrupts mitochondrial homeostasis through ROS production, independent of aggregate formation.
- Antioxidant intervention can ameliorate the detrimental effects of mutant SOD1 on mitochondrial function.
- These findings highlight the critical role of oxidative stress in ALS pathogenesis and suggest antioxidant strategies for therapeutic intervention.
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