Fig. 1: Endothelial knockdown of RAB5IF inhibits retinal neovascularization in oxygen-induced retinopathy mice.

A UMAP (uniform manifold approximation and projection) dimensionality reduction plot showing the cell annotations of GSE150703 data. B Group information displayed in the UMAP plot for oxygen-induced retinopathy (OIR) and age-matched normoxia mice (NORM). C UMAP plot illustrating the distribution of RAB5IF in OIR retinas. D Bidirectional bar graph comparing the fold changes of RAB5IF expression, with adjusted P-values shown. Blue: significant downregulation, red: significant upregulation, gray: not significant (N.S.). E Volcano plot highlighting differentially expressed genes (DEGs) between OIR and age-matched normal mice groups, with RAB5IF annotated. F qRT-PCR and Western blot analyses showed elevated RAB5IF levels in primary mRMECs from P17 OIR mice versus normoxia controls (pooled from 4 mice/sample, n = 5 samples/group, 15 μg protein/lane for WB). G Immunofluorescence of P17 OIR retinal flat mounts revealed RAB5IF (green) localized to pathological neovascular regions, co-stained with IB4 (red) and NG2 (blue). Scale bar = 40 μm. (Representative of three independent experiments). H Retinal endothelial populations from GSE150703 were subdivided by classical markers; UMAP and dot plots demonstrated RAB5IF expression patterns within endothelial subtypes. I–K The retro-orbital injection of endothelial-targeted AAV (1 × 10¹¹ viral particles/7.5 µL/mouse) for RAB5IF knockdown was performed in neonatal P3 mice, followed by qRT-PCR and Western blotting analysis on P17 OIR to assess RAB5IF mRNA and protein levels in primary mRMECs (pooled from 4 mice/sample, n = 5 samples/group, 15 μg protein/lane for WB) (I). IB4 staining at P17 OIR, showing representative images along with the quantification of neovascular area (white areas) and non-perfused avascular area (purple-shaded areas) in OIR mice (n = 5 mice per group). Scale bar = 1 mm ( J). ELISA analysis of VEGFA levels in the vitreous in P17 OIR mice (n = 10 mice per group) (K). Data are presented as means ± S.D. “N.S.” indicates no statistical difference (P > 0.05); Wilcoxon rank sum test with Bonferroni’s post hoc test (D, E), Two-tailed Student’s t-test (F) and one-way ANOVA with Bonferroni’s post hoc test (I–K). Source data are provided as a Source Data file.