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Published on: March 11, 2020
Glutathione redox cycle dysregulation in Huntington's disease knock-in striatal cells
Márcio Ribeiro1, Tatiana R Rosenstock, Teresa Cunha-Oliveira
1CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal.
Insights
Huntington's disease striatal cells show increased oxidative stress and altered glutathione metabolism. Mutant cells have impaired glutathione export, contributing to neurodegeneration despite boosted antioxidant defenses.
Area of Science:
- Neuroscience
- Biochemistry
- Genetics
Background:
- Huntington's disease (HD) is a neurodegenerative disorder linked to CAG repeat expansion in the HD gene.
- Oxidative stress is implicated in HD neurodegeneration, but the glutathione system's role remains unclear.
- Full-length mutant huntingtin (FL-mHtt) expression impacts striatal cells, the primary affected region in HD.
Purpose of the Study:
- To investigate the glutathione redox cycle in striatal cells expressing FL-mHtt.
- To understand how FL-mHtt affects glutathione synthesis, levels, and export.
- To correlate glutathione system alterations with oxidative stress and apoptosis in HD models.
Main Methods:
- Analysis of glutathione system components and activities in striatal cells from HD knock-in mice.
- Measurement of intracellular reactive oxygen species (ROS) and caspase-3 activity.
- Assessment of glutathione ethyl ester treatment and multidrug resistance protein 1 (Mrp1) inhibition.
Main Results:
- Mutant cells exhibited increased ROS and caspase-3 activity, reduced by glutathione ethyl ester.
- Elevated intracellular glutathione levels and activities of glutathione peroxidase (GPx), glutathione reductase (GRed), glutathione-S-transferase (GST), and γ-glutamyl transpeptidase (γ-GT) were observed.
- Decreased de novo glutathione synthesis (reduced glutamate-cysteine ligase (GCL) and glutathione synthetase (GS) activities) and impaired glutathione export (decreased Mrp1 expression and activity) were found.
Conclusions:
- FL-mHtt expression in striatal cells dysregulates the glutathione system, leading to decreased synthesis and impaired export.
- Reduced Mrp1-mediated glutathione export contributes to altered intracellular glutathione levels in HD cells.
- Enhanced glutathione antioxidant defenses are insufficient to overcome increased ROS and apoptosis in HD striatal cells.
Abstract:
Huntington's disease (HD) is a CAG repeat disorder affecting the HD gene, which encodes for huntingtin (Htt) and is characterized by prominent cell death in the striatum. Oxidative stress was previously implicated in HD neurodegeneration, but the role of the major endogenous antioxidant system, the glutathione redox cycle, has been less studied following expression of full-length mutant Htt (FL-mHtt). Thus, in this work we analyzed the glutathione system in striatal cells derived from HD knock-in mice expressing mutant Htt versus wild-type cells. Mutant cells showed increased intracellular reactive oxygen species (ROS) and caspase-3 activity, which were significantly prevented following treatment with glutathione ethyl ester. Interestingly, mutant cells exhibited an increase in intracellular levels of both reduced and oxidized forms of glutathione, and enhanced activities of glutathione peroxidase (GPx) and glutathione reductase (GRed). Furthermore, glutathione-S-transferase (GST) and γ-glutamyl transpeptidase (γ-GT) activities were also increased in mutant cells. Nevertheless, glutamate-cysteine ligase (GCL) and glutathione synthetase (GS) activities and levels of GCL catalytic subunit were decreased in cells expressing FL-mHtt, highly suggesting decreased de novo synthesis of glutathione. Enhanced intracellular total glutathione, despite decreased synthesis, could be explained by decreased extracellular glutathione in mutant cells. This occurred concomitantly with decreased mRNA expression levels and activity of the multidrug resistance protein 1 (Mrp1), a transport protein that mediates cellular export of glutathione disulfide and glutathione conjugates. Additionally, inhibition of Mrp1 enhanced intracellular GSH in wild-type cells only. These data suggest that FL-mHtt affects the export of glutathione by decreasing the expression of Mrp1. Data further suggest that boosting of GSH-related antioxidant defense mechanisms induced by FL-mHtt is insufficient to counterbalance increased ROS formation and emergent apoptotic features in HD striatal cells.
