Figure 5.

Caffeine’s protection against OIR is independent of A₁R activation. A) Mouse retinas from room air/OIR and water/caffeine (1 g/L) groups were harvested on P17 and analyzed for A₁R mRNA levels in retinas by quantitative PCR. Caffeine reversed the down-regulation of retinal A₁R mRNA expression in OIR. **P < 0.01, ***P < 0.001; 2-way ANOVA, Bonferroni post hoc test (n = 6/group). B) Retinal vasculature was visualized by whole-mount isolectin B4 staining at P17 of OIR. Representative images show avascular area (red dotted line) from WT and A₁R-KO mice (with or without caffeine treatment). Scale bar, 500 μm. C) Retinal pathologic angiogenesis (red arrow) from WT and A₁R-KO mice (with or without caffeine treatment) were detected by using hematoxylin and eosin staining at P17. Scale bar, 20 μm. D) Avascular areas (%) from WT and A₁R-KO mice (with or without caffeine treatment) in 3 groups were quantified as percentages of the whole retinal surface. Whereas A₁R KO increased avascular areas, caffeine treatment still reduced avascular areas in A₁R-KO mice. ***P < 0.001; 1-way ANOVA, Bonferroni post hoc test (n = 8/group). E) The number of vascular nuclei on the vitreal side of the inner limiting membrane in 3 groups was quantified and analyzed. Caffeine treatment reduced intravitreal vascular nuclei in A₁R-KO mice. **P < 0.01, ***P < 0.001; 1-way ANOVA, Bonferroni post hoc test (n = 6–8/group).