ZEB1 silencing protects against ferroptosis and mitochondrial dysfunction in osteoarthritis by inhibiting HSPA5 expression

Zinc finger E-box binding homeobox 1 (ZEB1), a ferroptosis-associated gene, is upregulated in osteoarthritis (OA) articular cartilage. However, whether ZEB1 regulates ferroptosis in OA progression remain unclear. ZEB1 protein levels in cartilage specimens from OA patients and normal controls were measured. Interleukin 1β (IL-1β)-induced chondrocyte injury model was established, followed by short hairpin RNA (shRNA)-mediated ZEB1 silencing in chondrocytes. Chondrocyte viability, apoptosis, inflammatory cytokine expression, extracellular matrix (ECM) degradation were assessed. Moreover, ferrous ion (Fe2+) level, reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH), glutathione peroxidase 4 (GPX4), mitochondrial membrane potential (MMP) and ATP level were determined. ZEB1-mediated transcriptional regulation of heat shock protein family A member 5 (HSPA5) was validated. Rescue experiments were conducted to validate the ZEB1/HSPA5 regulatory axis in chondrocyte injury. OA mouse model was constructed, and ZEB1 shRNA was injected into OA mice. The pathological changes in cartilage tissues were detected. ZEB1 was upregulated in OA cartilage tissues. ZEB1 silencing attenuated IL-1β-induced apoptosis, inflammation, and ECM degradation. IL-1β treatment increased Fe2+, ROS, and MDA levels and decreased GPX4 and GSH levels in chondrocytes, while ZEB1 silencing reversed these changes. ZEB1 silencing abrogated IL-1β-induced MMP and ATP reduction. Mechanistic studies revealed that ZEB1 transcriptionally inhibited HSPA5 expression in chondrocytes. HSPA5 silencing abrogated the protective effects of ZEB1 silencing. Additionally, ZEB1 silencing alleviated articular cartilage degradation, inflammatory response, and iron deposition. ZEB1 silencing ameliorated IL-1β-induced chondrocyte injury and OA progression by suppressing ferroptosis and mitochondrial dysfunction via HSPA5 inhibition.