Protective effect of norepinephrine, dopamine and N-methyldopamine against chemically-induced oxidative ferroptosis in HT22 neuronal cells: Protein disulfide isomerase as a mechanistic target for protection.

Under pathogenic conditions, the endogenous neurotransmitters dopamine and norepinephrine can readily undergo auto-oxidation to generate reactive oxygen species (ROS), culminating in glutathione depletion and oxidative neuronal injury. Recently, we have revealed that protein disulfide isomerase (PDI) is a mediator of glutathione depletion-associated oxidative ferroptosis, which is also a novel target for ferroptosis protection. In this study, we identify that three chemicals of the endogenous catecholamine family, i.e., norepinephrine, dopamine and N-methyldopamine, are capable of inhibiting PDI and can effectively protect against oxidative ferroptosis in cultured HT22 hippocampal neurons after challenged with different ferroptosis inducers, including erastin, RSL3, glutamate, sulfasalazine and l-buthionine-(S,R)-sulfoximine. Evidence is presented to show that norepinephrine, dopamine and N-methyldopamine can directly bind to PDI in live HT22 cells, and covalently modify the free thiol groups in PDI's catalytic sites, likely through electrophilic attacks. PDI knockdown attenuates the protective effect of norepinephrine, dopamine and N-methyldopamine against chemically-induced ferroptosis in these cells. Mechanistically, inhibition of PDI by norepinephrine, dopamine and N-methyldopamine or PDI knockdown by siRNAs each markedly reduces iNOS and nNOS activation (dimerization) and NO accumulation, and these changes are associated with reduced accumulation of cellular reactive oxygen species (ROS) and lipid-ROS and alleviation of chemically-induced ferroptotic neuronal death. Collectively, the findings of this study reveal that certain oxidative derivatives of catecholamine neurotransmitters, which may be highly cytotoxic, also possess the unique ability to rescue neuronal cells from glutathione depletion-associated oxidative ferroptosis, and PDI serves as a key mechanistic target which mediates their cytoprotective actions.