Figure 4. Slc38a5 is enriched in oxygen-induced retinopathy (OIR) pathological neovessels and its deficiency suppresses pathological angiogenesis in OIR.

(A) Slc38a5 mRNA expression was measured by RT-qPCR at P8, P12, P14, and P17 in C57BL/6J OIR retinas compared with age-matched normoxic control mice. Slc38a5 mRNA levels were decreased during hyperoxia stage (P8 and P12) and increased in hypoxia stage (P14 and P17). (B) Slc38a5 mRNA expression was analyzed using RT-qPCR in laser capture micro-dissected pathological neovessels from P17 unfixed C57BL/6J OIR retinas compared with normal vessels isolated from P17 normoxic retinas. Images on the left are representative retinal cross-sections from normal and OIR retinas stained with isolectin B₄ (red) and DAPI (blue), with dotted lines highlighting micro-dissected retinal vessels. GCL: ganglion cell layer, INL: inner nuclear layer, ONL: outer nuclear layer. (C) Protein levels of SLC38A5 (52 kDa) were increased in C57BL/6J OIR retinas at P17 compared with normoxic controls using Western blot and quantified with densitometry. Proteins were normalized to β-ACTIN (42 kDa). (D) Slc38a5^−/− exposed to OIR had decreased levels of pathological NV (neovascularization) compared with WT OIR controls bred in the same colony at P17. There was no significant difference in VO (vaso-obliteration) between the two groups. Scale bar: 50 µm (B), 1 mm (D). Each dot represents one retina. Data are expressed as mean ± SEM. n=3–6 per group (A–C), n=20 per group (D). *p≤0.05; **p≤0.01; n.s.: not significant.

Figure 4—figure supplement 1. Quantification of vaso-obliteration of P12 wild type (WT) and Slc38a5^−/− retinas in oxygen-induced retinopathy shows comparable levels.

(A) Flat mounts of P12 WT and Slc38a5^−/− retinas showing central avascular area of vaso-obliteration (VO) absence of isolectin B₄ staining (red). (B) Quantification of VO. n=10–12/group. NS: not significant. Scale bars: 500 µm.