Nrf2 regulates the temporal dynamics of retinal vascular development in mice

The murine retinal vasculature is a model for studying angiogenesis, but the precise temporal sequence of vascular plexus formation and the role of hyaloid vessels remain incompletely defined. Retinas from C57BL/6 J wild-type and Nrf2 knockout (Nrf2^-/-) mice from postnatal day (P)1 to P21 were analyzed by whole-mount immunofluorescence and frozen section staining. In wild-type mice, superficial vascular plexus (SVP) formation began at P1 from the optic nerve head, reaching the retinal margin by P8, with vascular coverage increasing from 2.78% to 97.61% of total retinal area. Before SVP completion, the hyaloid artery (HA) and vasa hyaloidea propria (VHP) were already present on the inner retinal surface. At P4, VHP remnants were sparse and composed of larger-diameter vessels, distinct from the developing SVP. Between P9-P12, SVP sprouts formed the deep vascular plexus (DVP), followed by the intermediate plexus (IVP) by P17; all three layers matured by P21. In Nrf2^-/- mice, vascular development was delayed: SVP density reduced at P9 (recovered by P12), DVP reduced at P12 (recovered by P17), and IVP density reduced at P17 with increased tip cells. Importantly, P30 assessment showed no genotype differences in vascular density or fluorescein leakage, confirming the vascular delay was reversible without permanent deficits. This study delineates the developmental timeline of murine retinal vessels, describes hyaloid vessel morphology during early SVP formation, and identifies Nrf2 as a regulator of angiogenic timing.