Background <p>Diabetic nephropathy (DN) represents a leading cause of chronic kidney diseases worldwide. While solute carrier family 40 member 1 (SLC40A1) is involved in the regulation of iron homeostasis and iron transport in diabetic kidney disease, its specific role and underlying mechanism in regulating ferroptosis during DN progression remain elusive.</p> Methods <p>Protein and mRNA expression levels were detected using western blot and quantitative real-time polymerase chain reaction (qRT-PCR), respectively. Cell viability and apoptosis were measured using the cell counting kit-8 (CCK8) and flow cytometry assays. The tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), ferrous iron (Fe²⁺), reactive oxygen species (ROS), glutathione (GSH), and malondialdehyde (MDA) levels were determined using corresponding assay kits. Bioinformatics analysis and methylated RNA Immunoprecipitation (MeRIP), or RNA immunoprecipitation (RIP) was performed to examine the relationship between SLC40A1 and methyltransferase-like protein 14 (METTL14), YTH domain family protein 3 (YTHDF3), or E2F transcription factor 4 (E2F4). The interaction between SLC40A1 and E2F4 was examined by chromatin immunoprecipitation (ChIP) and dual-luciferase reporter gene assays.</p> Results <p>SLC40A1 levels were decreased in high glucose (HG)-induced HK-2 cells. Notably, SLC40A1 overexpression abolished HG-induced action of cell viability, cell apoptosis, inflammation, and ferroptosis. Mechanistically, silencing SLC40A1 greatly reversed the protective roles of METTL14 knockdown in HG-induced damage and ferroptosis of HK-2 cells. In addition, in vivo, SLC40A1 mitigated kidney damage, inflammation, and ferroptosis.</p> Conclusions <p>METTL14 knockdown remits HG-induced HK-2 cell apoptosis, inflammation, and ferroptosis via promoting SLC40A1. In addition, E2F4 enhances SLC40A1 transcription. This research provides new potential targets and insights for the treatment of DN.</p>