Protective effect and mechanism of Saikosaponin A against oxidative damage in cochlear hair cells in vitro

This study aims to investigate the protective effect and underlying mechanism of Saikosaponin A (SSA) against oxidative stress-induced damage in mouse cochlear hair cells (HEI-OC1). An oxidative stress model was established by treating HEI-OC1 cells with 50 mU/mL glucose oxidase (GO). Cells were divided into four groups: Control, GO, Control + SSA, and GO + SSA (1 μM SSA). Cell viability was assessed using the CCK-8 assay, and apoptosis was evaluated by flow cytometry and by analyzing expression of BAX, BCL-2, and cleaved CASPASE-3. Oxidative stress levels were assessed via malondialdehyde (MDA), reactive oxygen species (ROS), and glutathione peroxidase (GSH-Px). mRNA levels of Ptgs2 (COX-2), Nos2 (iNOS), Hmox1 (HO-1), and Sod1 (SOD1) were detected by qRT-PCR. Protein expression of KEAP1, NFE2, and phosphorylated NFE2 was examined by western blot, including analysis of nuclear translocation. All experiments were independently performed in triplicate. SSA increased cell viability by approximately 40% (P < 0.01) and reduced apoptotic rate by 70% (P < 0.001) in GO-treated HEI-OC1 cells. SSA also decreased MDA and ROS levels and restored GSH-Px activity (P < 0.01). Moreover, SSA downregulated Ptgs2 and Nos2 expression, while upregulating Hmox1 and Sod1. At the protein level, SSA suppressed KEAP1 expression, enhanced NFE2 and p-NFE2 levels, and promoted NFE2 nuclear translocation. SSA alleviates GO-induced oxidative stress and apoptosis in HEI-OC1 cells by activating the KEAP1/NFE2 signaling pathway. These findings support the potential application of SSA in protecting cochlear hair cells from oxidative damage, warranting further in vivo investigation.