Methodological approaches for dissecting ferroptosis in lens epithelial cells and its role in diabetic cataract progression.

Diabetic cataract has become the second-largest blind causing disease among the complications of diabetes mellitus in eyes. Ferroptosis is responsible for many complications associated with diabetes mellitus, but the role of ferroptosis in diabetic cataract has not yet been studied. Here, we first ascertained that ferroptosis was triggered during diabetic cataract, as identified by elevated levels of transferrin receptor protein 1 (CD71) and solute carrier family 7 member 11 (SLC7A11), lipid peroxidation , total iron and Fe2+ under conditions of excessive glucose. Moreover, voltage-dependent anion channel 3 (VDAC3) was found to decrease under high levels of glucose both in vivo and in vitro. VDAC3 knockdown resulted in increased reactive oxygen species (ROS) levels and reduced nicotinamide adenine dinucleotide (NADH) levels, indicating that decreased VDAC3 modulates ferroptosis in diabetic cataract via ROS-induced lipid peroxidation. Furthermore, we verified that the level of ubiquitin-specific peptidase 47 (USP47) was decreased in high glucose-treated lens epithelial cells. We found that USP47 bound to and modulated the deubiquitination of VDAC3. This binding between USP47 and VDAC3 was weakened and VDAC3 was subsequently ubiquitinated and degraded under high glucose conditions, which ultimately resulted in ferroptosis and diabetic cataract. Overall, we verified that the USP47-VDAC3-ROS/NADH axis regulates ferroptosis in high glucose-treated lens epithelial cells (LECs) and in an animal model of diabetic cataract. Our findings increase the understanding of human diabetic cataract and provide important mechanistic insight into potential ferroptosis treatments in medicine and technology for treating diabetic cataract.