Protective Effect of SESN2 on Glutamate Neurotoxicity via Keap1‐Nrf2 Pathway‐Mediated Mitophagy

Glutamate (Glu) possesses functional significance concerning neurological disorders by producing neurotoxicity as a major excitatory amino acid neurotransmitter. Sestrin2 (SESN2) has been affirmed to elicit wide neuroprotective properties as a highly conserved stress-responsive protein. Therefore, this project sets out to ascertain the impacts of SESN2 on Glu neurotoxicity and the concealed operating mechanism. Cell counting kit-8 (CCK-8) assay, lactate dehydrogenase (LDH) assay kit, and Western blot estimated cell viability, cytotoxicity, and SESN2 expression. Commercial kits and fluorescence probes were employed to assess the degree of oxidative stress. The apoptotic changes were evaluated by terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) and Western blot. Mitochondrial function was measured by 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine iodide (JC-1) staining, MitoSOX staining, and Western blot analysis of mitophagy-related proteins and immunofluorescence. Kelch-like ECH-associated protein 1 (Keap1)-nuclear factor erythroid 2-related factor 2 (Nrf2) pathway-related proteins were also examined with Western blot. SESN2 expression was elevated in HT-22 cells upon stimulation with Glu. SESN2 upregulation reduced the viability loss, LDH release, oxidative stress, and apoptosis in HT-22 cells imposed by the Glu challenge, while a contrary trend was observed when SESN2 was downregulated. Moreover, hyperexpressed SESN2 activated the Keap1-Nrf2 pathway to promote mitophagy in Glu-exposed HT-22 cells. Deletion of Nrf2 partly abolished the effects of SESN2 elevation on the mitophagy, and mitophagy blocker Mdivi-1 partly reverted the influences of SESN2 overexpression on the viability, LDH release, oxidative stress, and apoptosis in Glu-stimulated HT-22 cells. SESN2 might mediate mitophagy via the Keap1-Nrf2 pathway to confer neuroprotection toward Glu-provoked toxicity.