Formononetin attenuates early diabetic retinal and renal microvascular injury associated with EGR1/VEGF-related endothelial responses.

ETHNOPHARMACOLOGICAL RELEVANCE: Formononetin (FMN) is a natural isoflavone found in several medicinal plants. Its effects on diabetic retinal and renal microvascular injury and EGR1/VEGF-related endothelial responses remain incompletely defined. AIM OF THE STUDY: To assess whether FMN attenuates early diabetes-associated retinal and renal injury and whether EGR1 is involved. MATERIALS AND METHODS: Alloxan-induced diabetic rabbits received FMN orally at 25 mg/kg/day from Week 4 to Week 16 after pretreatment renal and OCT/OCTA assessment. Renal function, histopathology, retinal flat mounts, longitudinal OCTA, and tissue EGR1/VEGF expression were examined. High-glucose-treated retinal, renal microvascular, and umbilical vein endothelial cells were used for functional assays. EGR1 overexpression, VEGFA promoter reporter assays, SPR, docking, and molecular dynamics simulations were also performed. RESULTS: FMN lowered serum creatinine, blood urea nitrogen, and urinary albumin-to-creatinine ratio, and reduced renal histopathological injury. It also attenuated retinal capillary loss, retinal thinning, and OCTA abnormalities despite persistent hyperglycemia. Retinal and renal tissues from FMN-treated rabbits showed lower EGR1 and VEGF expression. In endothelial cells, FMN reduced EGR1 and VEGF expression, abnormal tube formation, permeability, invasion, ROS, and inflammatory mediator release, while increasing NO-related signals. EGR1 overexpression weakened several effects and increased VEGFA promoter activity through the predicted EGR1-binding sequence. SPR detected FMN-EGR1 binding with a K_D of 3.245 μM. Computational analyses provided structural context. CONCLUSIONS: FMN attenuated early diabetes-associated retinal and renal injury and was linked to EGR1/VEGF-related endothelial responses. The data support EGR1 involvement but do not establish it as the sole target or mediator of FMN.