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

Objective: 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).

Methods: 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.

Results: 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.

Conclusion: 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.

HEI-OC1 cells; KEAP1/NRF2 pathway; Saikosaponin A; cell apoptosis; cell viability; cochlear hair cells; oxidative damage; protective effect.