Abstract
Mitochondrial glutathione pool is vital in protecting cells against oxidative stress as the majority of the cellular reactive oxygen species are generated in mitochondria. Oxidative stress is implicated as a causative factor in neuronal death in neurodegenerative disorders. We hypothesized that depletion of mitochondrial glutathione leads to mitochondrial dysfunction and apoptotic death of SK-N-SH (human neuroblastoma) cells and investigated the neuroprotective strategies against GSH depletion. SK-N-SH cells were treated with two distinct inhibitors of glutathione metabolism: L-buthionine-(S, R)-sulfoximine (BSO) and ethacrynic acid (EA). EA treatment caused depletion of both the total and mitochondrial glutathione (while BSO had no effect on mitochondrial glutathione), enhanced rotenone-induced ROS production, and reduced the viability of SK-N-SH cells. Glutathione depletion by BSO or EA demonstrated positive features of mitochondria-mediated apoptosis in neuroblastoma cell death. Prevention of apoptosis by Bcl2 overexpression or use of antioxidant ebselen did not confer neuroprotection. Co-culture with U-87 (human glioblastoma) cells protected SK-N-SH cells from the cell death. Our data suggest that depletion of mitochondrial glutathione leads to mitochondrial dysfunction and apoptosis. The study indicates that preventing mitochondrial glutathione depletion could become a novel strategy for the development of neuroprotective therapeutics in neurodegenerative disorders.
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Abbreviations
- BSO:
-
L-Buthionine-(S,R)-sulfoximine
- EA:
-
Ethacrynic acid
- ETC:
-
Electron transport chain
- GSH:
-
Glutathione
- PI:
-
Propidium iodide
- ROS:
-
Reactive oxygen species
- SK-N-SH:
-
A human neuroblastoma cell line
- U-87:
-
A human glioblastoma cell line
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Acknowledgments
This study was supported by a grant from Idaho Biomedical Research Infrastructure Network (NIH NCRR BRINIP20RR016454) to JCKL, Idaho State University GSRSC pre-doctoral grant to VVD and NIH RO1 DK077910 to ALMB. We are thankful for the important suggestions from Drs. Christopher Daniels, Alok Bhushan and Leslie Devaud. VVD thanks the Idaho INBRE NIH program (grant # P20RR016454) for a research fellowship.
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Dukhande, V.V., Kawikova, I., Bothwell, A.L. et al. Neuroprotection against neuroblastoma cell death induced by depletion of mitochondrial glutathione. Apoptosis 18, 702–712 (2013). https://doi.org/10.1007/s10495-013-0836-4
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DOI: https://doi.org/10.1007/s10495-013-0836-4