Objective: Retinal neovascularization is an iconic change in retinopathies. Vascular Endothelial Growth Factor (VEGF) and α-crystallins have been identified to mediate the pathogenesis of retinopathy. However, the special and temporal changes in their expression associated with retinal neovascularization have not yet been determined. Therefore, we examined the expression and distribution of VEGF, αA- and αB-crystallins in the retina using a mouse model of oxygen-induced retinopathy (OIR).
Methods: 90 C57/BL mice were randomly divided into the OIR and control groups. The OIR group at postnatal day 7 (P7) were kept at high oxidation state (75 ± 5%) for 5 days before returned to normal environment. Retinal tissue was cut into sections. Oxygen induced retinal neovascularization and vascular structural changes were evaluated using retinal fluorescein angiography. The number of endothelial cell nuclei breaking through the retinal internal limiting membrane was counted after H&E staining. The mRNA expression levels of VEGF, αA- and αB-crystallins in the mouse retina were determined using real-time RT-PCR. The distribution of αA- and αB-crystallins in the retina was detected by fluorescent immunohistochemistry staining.
Results: Oxygen induction triggered new blood vessel formation in the retina and impaired the structure of the retinal vascular network. The number of endothelial cell nuclei breaking through the retinal internal limiting membrane was significantly increased in the OIR group compared to the control group at P13, P17 and P21 (P < 0.01), reaching the peak on P17. The expression levels of VEGF, αA- and αB-crystalllins were also significantly different between the OIR and control groups. VEGF expression was highest on P15, αA-crystallin expression was highest on P17, whereas αB-crystallin expression kept increasing during the time frame of our study. Both αA- and αB-crystallins were expressed in the ganglion cell layer and the inner nuclear cell layer. While αA- and αB-crystallins were only located on the cell membrane in the outer ganglion cell layer, they were observed both on the cell membrane and in the cytoplasm in the inner layer of cells.
Conclusion: Using our mouse model of oxygen-induced retinopathy, we showed that the expression patterns of VEGF, αA- and αB-crystallins during retinal neovascularization in both spatially and temporally manners, providing significant insights into the molecular mechanisms of retinopathy and the associated neovascularization.
Keywords: VEGF; gene expression pattern; mouse model; neovascularization; oxygen induced retinopathy; αA-crystallin; αB-crystallin.