Abstract

Introduction. Ferroptosis, an iron-dependent form of nonapoptotic cell death, has been identified as a type of death that occurs in conditions of brain damage, such as hemorrhagic infarction. Ferroptosis is characterized by increased lipid reactive oxygen species (ROS) due to a Fenton reaction. Ascorbic acid (AA) is the main antioxidant of the central nervous system, and under pathophysiological conditions, it is oxidized to dehydroascorbic acid (DHA), favoring the Fenton reaction and increasing ROS. To date, the initiating mechanisms of neural cancer cells (NCC) ferroptosis are unknown. Here, we show that NCC treated with vitamin C are sensitized to death by ferroptosis. Materials and methods. NCC (N2a, U87, C3, HN33.11, and SH-SY-5Y) were treated with a single dose of 10μM Erastin (inducer of ferroptosis) with AA or DHA every 12 h for 48 h. Additionally, ferrostain was used as an inhibitor of ferroptosis. The cell viability was monitored by the incorporation of Sitox green with real-time live-cell microscopy over 48 h using an IncuCyte S3 system. Total (CellRox) and lipid (Image-iT lipid perdoxidation) ROS were determined by FACS. To determine if the effect was induced by AA or DHA, we generated SVCT2-/- N2a cells with CRISPR/Cas9. As a control of necroptosis, we used MLKL-/- N2a cells. Using SIM-3D microscopy and mitotracker, we analyzed mitochondrial fragmentation. In addition, we used Mdivi-1 as a Drp-1 inhibitor. Results. Vitamin C (AA/DHA) supplementation favors ferroptosis in NCC. SVCT2-/- N2a cells are sensitive to ferroptosis induced by DHA. Drp-1 does not participate in the ferroptotic process because its inhibition does not prevent cell death. Discussion. AA/DHA can induce neuronal ferroptosis under pathophysiological conditions and Erastin treatment.