Selective Clearance of Senescent Chondrocytes in Osteoarthritis by Targeting Excitatory Amino Acid Transporter Protein 1 to Induce Ferroptosis

Aims: This study aimed at exploring the mechanism of ferroptosis (an iron-dependent form of nonapoptotic cell death) resistance in senescent chondrocytes (SenChos). Results: In this study, by utilizing metabolomics and single-cell RNA sequencing, we found that hyperactivation of ferroptosis metabolism was one of the most prominent metabolic features in SenChos. Interestingly, however, SenChos were able to survive in this state and were resistant to ferroptosis-induced cell death. Next, we elucidated that this survival mechanism of SenChos could be primarily attributed to overexpression of the membrane protein excitatory amino acid transporter protein 1 (EAAT1), which can increase intracellular glutamate (Glu) levels and activate the glutathione system to counteract ferroptosis. In addition, 2-amino-5,6,7,8-tetrahydro-4-(4-methoxyphenyl)-7-(naphthalen-1-yl)-5-oxo-4H-chromene-3-carbonitrile (UCPH-101) (a specific inhibitor of EAAT1) and siRNA-EAAT1 were able to substantially increase the sensitivity of SenChos to ferroptosis and to induce cell death, with no apparent effects on the normal cells. Administration of an intraarticular injection of UCPH-101 caused inhibition of EAAT1 selectively, cleared SenChos from cartilage, improved the cartilage homeostasis, and significantly delayed the progression of osteoarthritis (OA). Innovation: This work supports a relevant role for EAAT1 in ferroptosis resistance mechanism for SenChos, revealing a potential therapeutic target of OA. Conclusions: EAAT1-Glu-glutathione peroxidase 4 anti-ferroptosis axis is a key survival mechanism for SenChos, and EAAT1 is an effective and specific target for anti-senescence therapy in OA. Antioxid. Redox Signal. 39, 262–277.