Regulation of calcium signaling prevents neuronal death mediated by NIST DEP in xenoferroptotic cell death conditions.

Increased calcium levels are associated with the ferroptosis pathway in neurodegenerative conditions. Recent evidence showed that exposure to particulate matter (PM) could accelerate the pathology of neurodegenerative diseases. However, the molecular mechanisms of how PM could affect brain cell pathology is not fully understood. We hypothesized that diesel exhaust particles (NIST DEP) could alter the ferroptosis pathway through calcium signaling, and therefore accelerate the cell death pathway. In this study, we used mouse hippocampal neuronal-like HT22 cells to evaluate whether exposure to NIST DEP could accelerate RSL-3-induced ferroptosis by increasing calcium deregulation, mitochondrial dysfunction and reactive oxygen species (ROS). MTT assay results showed that NIST DEP (25, 50, 75, and 100 μg/mL) did not significantly reduce the survival rate of HT22 cells, while NIST DEP significantly increased RSL-3-induced ferroptotic cell death in a concentration-dependent manner. Based on fluorescence image analysis, co-exposure to NIST DEP and RSL-3 disrupted HT22 cell mitochondrial morphology, intracellular and mitochondrial calcium levels. Combined exposure resulted in an increase in ER-mitochondria contact sites measured by proximity ligation assay (PLA) compared to control solvent group. Additionally, lipid peroxidation, mitochondrial ROS and malondialdehyde content, were increased significantly by combined exposure to NIST DEP and RSL-3. Interestingly, the calcium regulators of the mitochondrial calcium uniporter MCUi4 and positive modulation of small conductance calcium-activated potassium channels by CyPPA significantly preserved cellular metabolic activity, restored calcium homeostasis, and alleviated fragmentation of mitochondria. Consequently, targeting calcium signaling may be promising therapeutic option for xenoferroptotic conditions in which PM affect cell survival.