USP7-stabilized FOXD1 activates the Gal-3-VEGFA/PI3K/AKT pathway to promote epithelial‒mesenchymal transition and angiogenesis in diabetic retinopathy

Diabetic retinopathy (DR) is a common microvascular complication of diabetes mellitus (DM), and its recurrence often leads to irreversible vision loss. During the process of retinal damage induced by high glucose (HG), epithelial‒mesenchymal transition (EMT) and angiogenesis are the core processes, but the underlying molecular mechanism remains unclear. C57BL/6J mice were induced with streptozotocin (STZ), and ARPE-19 cells and HRMECs were stimulated with HG to establish in vivo and in vitro DR models. The mice were randomly allocated to four groups: non-diabetic control, STZ-induced DR, DR + sh-NC and DR + sh-FOXD1; in the last group, FOXD1 was knocked down by bilateral intravitreal lentiviral injection. H&E staining was used to evaluate pathological damage to retinal tissues. Western blot, immunofluorescence, Transwell, and tube formation assays were performed to detect EMT and angiogenesis. FOXD1 was highly expressed in DR mice. Knocking down FOXD1 could partially reverse weight loss and decrease blood glucose levels in DR mice and attenuate retinal histopathological damage while reducing the levels of CD31 and VEGFA. At the cellular level, knocking down FOXD1 expression led to a decrease in EMT in HG-induced ARPE-19 cells and inhibited HG-induced HRMEC viability and angiogenesis. In terms of molecular mechanisms, USP7 stabilizes FOXD1 expression via deubiquitination, FOXD1 transcriptionally activates Gal-3 in ARPE-19 cells and VEGFA in HRMECs to increase the expression of PI3K/AKT signaling pathway proteins, and the overexpression of Gal-3 or VEGFA and the PI3K agonist 740 Y-P can weaken the inhibitory effect of FOXD1 knockdown. USP7 stabilizes FOXD1 expression through deubiquitination. FOXD1 activates the PI3K/AKT signaling pathway by promoting the transcription of Gal-3 in ARPE-19 cells and VEGFA in HRMECs, thereby promoting EMT in ARPE-19 cells and angiogenesis in HRMECs.