Profiling of genes associated with the murine model of oxygen-induced retinopathy.

PURPOSE

To compare the clinical features and gene expression patterns of the physiologic development of retinal vessels and oxygen-induced retinopathy (OIR) in a mouse model, with the aim of identifying differential regulators of physiologic and pathological angiogenesis in the retina.

METHODS

C57BL/6J mice were used. Seven-day-old pups were subjected to OIR induction following the standard protocols of entering a hyperoxic chamber on day 7 (P7) and returning to a normoxic condition (relative hypoxia) on day 12 (P12). The retinal vasculatures in the OIR model 24 h (P8-O) or 5 days (P12-O) after switching to the hyperoxic environment and 24 h (P13-O) after returning to normoxic conditions were evaluated with retinal flat mounts and compared with those of age-matched controls (i.e., P8-N, P12-N, P13-N). Gene expression profiling was performed using Phalanx Mouse Whole Genome OneArray microarrays. Normal 9-day-old mice were considered representative of physiologic angiogenesis and compared with 30-day-old mice. A bioinformatics analysis was performed on differentially expressed genes using various comparisons, and real-time reverse-transcription PCR was used to confirm the changes in the genes of interest.

RESULTS

The sequential orders and patterns of vasculature development in normal mice and the OIR models were significantly different. In brief, in the early days (P1 to P7) for normal mice, retinal vessels grew from the optic disc into the non-vascularized retina in a radial fashion. In the hyperoxic stage of the OIR model, the main central retina became devoid of a vascular network, and when the mice returned to the normoxic room, the vessels grew from peripheral perfused areas toward the center of the retina, but the development of intermediate and deep layers of vasculature was significantly delayed. Gene profiling at three critical time points (P8, P12, and P13) showed that 162 probes were upregulated to ≥1.5-fold or downregulated to ≤0.67-fold at one or more time points in the OIR model compared to the controls. In the 45 upregulated genes for the P8-O/P8-N group, enriched genes were mainly related to cytoskeleton formation, whereas the 62 upregulated genes for P13-O/P13-N participated in various pathological processes. In the physiologic conditions on P9, however, 135 genes were upregulated compared with P30; the gap junction and Fc gamma R-mediated phagocytosis were the two main enriched pathways for these genes. Fifty-three probes, including vascular endothelium growth factor A, annexin A2, and endothelin 2, changed at P13-O but not at P9-N, and these changed genes might reflect the modulation of pathological neovascularization.

CONCLUSIONS

Angiogenesis in physiologic and pathological conditions is characterized by the differential presentation of vasculature and gene expression patterns. Investigation of those genes unique to the OIR model may help develop new strategies and therapies for intervening in retinal neovascularization.