Flotillin- 1 ameliorates experimental diabetic retinopathy by inhibiting ferroptosis in blood-retinal barrier

Diabetic retinopathy (DR) is a chronic disease that severely impairs the vision of working individuals and is closely linked to blood-retinal barrier (BRB) dysfunction. Flotillin- 1 (FLOT1), a protein located in membrane lipid rafts, is essential for various intracellular biological processes. However, its role in the pathogenesis of DR remains unclear. Ferroptosis in high-glucose was assessed using Cell counting kit- 8 (CCK- 8), Malondialdehyde (MDA), Glutathione (GSH), Fe²⁺ assays, and transferrin expression. BRB disruption was evaluated with Evans blue staining. The interaction between FLOT1 and NF-E2-related factor 2 (Nrf2) was confirmed by immunoprecipitation and ferroptosis mechanisms were explored by inhibiting Nrf2 with ML385. In db/db mice (a type 2 diabetes model) was intravitreal injection of an adeno-associated virus (AAV) overexpressing FLOT1. Expression levels of Nrf2, solute carrier family 7 member 11 (SLC7 A11), and glutathione peroxidase 4 (GPX4) were evaluated in retina. Our study indicated that FLOT1 significantly alleviated BRB damage in DR, reversing high-glucose induced reductions in GPX4 and GSH, and inhibited the elevation of MDA and Fe²⁺. FLOT1 also suppressed ROS accumulation. Mechanistically, FLOT1 activates the Nrf2 pathway by enhancing its expression and promoting its nuclear translocation, thereby stimulating the SLC7 A11/GPX4 pathway to inhibiting lipid peroxidation and ferroptosis. We have identified ferroptosis is a key mechanism driving BRB damage in DR.