Mesenchymal stem cell-derived extracellular vesicles protect against radiation-induced retinal injury in human retinal organoids via OSER1-mediated ferroptosis suppression

Radiation retinopathy (RR) is a sight-threatening complication of radiotherapy with no effective treatment. The scarcity of human retinal specimens has impeded mechanistic studies. Human embryonic stem cell-derived retinal organoids (hESC-ROs) offer a promising platform to model the early-stage cellular responses to radiation-induced retinal injury. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) show therapeutic potential for retinal diseases, but their role in radiation injury remains unexplored. We established an in vitro human retinal radiation injury model using 60 Co-γ-irradiation of hESC-derived organoids, followed by bone marrow MSC-EVs treatment. Cell death, ROS, mitochondrial membrane potential and ferroptosis markers (GSH, MDA, Fe 2+ ) were assessed by flow cytometry and biochemical assays. RNA-seq characterized cell death mechanisms, validated by Western blotting, gene knockout and overexpression in hESCs, followed by retinal organoid differentiation, assessed gene roles in radiation-induced ferroptosis. hESC-ROs exposed to 32 Gy radiation induced significant cell death and mitochondrial dysfunction, accompanied by canonical ferroptosis features including GSH depletion, lipid peroxidation accumulation, Fe 2+ overload and GPX4 down-regulation. MSC-EVs significantly attenuated these effects. RNA-seq and GSEA identified ferroptosis as a top enriched pathway. Furthermore, the ferroptosis inducer RSL3 reversed the protection conferred by MSC-EVs, confirming that ferroptosis inhibition is a major mechanism of action. Notably, OSER1 was identified as a critical gene upregulated by MSC-EVs. Importantly, the protective effect of MSC-EVs was largely abrogated in OSER1-knockout organoids, while OSER1 overexpression rescued organoids, establishing OSER1 as a necessary downstream mediator of MSC-EVs function. hESC-ROs provide a valuable platform for modeling radiation-induced retinal injury. MSC-EVs protect against this injury by activating OSER1 to suppress ferroptosis, identifying OSER1 as a potential therapeutic target for radiation-induced retinal damage. These findings lay the groundwork for developing cell-free, EV-based strategies to preserve vision in patients suffering from RR.