Nrf2 maintains reactive oxygen species at an optimal level during endoplasmic reticulum stress in HT22 cells

The endoplasmic reticulum (ER) is a central organelle for protein folding and redox regulation. Disulfide bond formation in the ER inevitably generates reactive oxygen species (ROS), creating a more oxidative environment than the cytosol. The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) is a master regulator of antioxidant defense, yet its role in intracellular redox regulation and antioxidant pathway selectivity during ER stress remains incompletely characterized. Here, we investigated how Nrf2 modulates ROS levels under basal and ER stress conditions in HT22 mouse hippocampal cells. We found that Nrf2 attenuates hydrogen peroxide and superoxide accumulation under ER stress induced by thapsigargin or tunicamycin. Mechanistically, Nrf2 maintains redox balance through multiple pathways, including transcriptional induction of heme oxygenase-1 and preservation of glutathione and peroxiredoxin-4 levels. Furthermore, our results suggest that hydrogen peroxide functions not only as a cytotoxic molecule but also as a signaling mediator that sustains basal Keap1-Nrf2 pathway activity, thereby reinforcing redox homeostasis under both physiological and stress conditions. Among Keap1-Nrf2 target proteins, heme oxygenase-1 contributes to the regulation of mitochondrial superoxide levels. Together, these findings highlight the dual roles of hydrogen peroxide and Nrf2: limiting ROS-induced damage and orchestrating adaptive redox signaling in the ER. This study demonstrates the importance of Nrf2 in maintaining ER redox homeostasis and suggests its potential as a therapeutic target for diseases characterized by chronic ER stress, such as neurodegeneration.