Purchase this article with an account.
T.E. de Gooyer, K.A. Stevenson, P. Humphries, D.A. C. Simpson, T.A. Gardiner, A.W. Stitt; Contribution of Rod Photoreceptor Loss to the Reduction of Retinal Hypoxia and Hypoxia–Regulated Gene Expression in Rho–/– Mice . Invest. Ophthalmol. Vis. Sci. 2005;46(13):1667.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: Since photoreceptors are among the main consumers of oxygen in the retina, it has been hypothesised that rod loss in retinitis pigmentosa (RP) leads to a reduced metabolic demand and lower overall net retinal oxygen usage. In the current study, we sought to evaluate whether regions of the retina become hypoxic during periods of high oxygen consumption, and if outer retina removal (in RP) reduces hypoxia and hypoxia–mediated changes in gene expression. As vascular attenuation has been observed in RP, we also sought to assess whether the retinal vasculature changes concomitantly with alterations in retinal hypoxia. Methods: Retina from rhodopsin knockout (Rho–/–) mice and C57Bl/6 (wild type; WT) controls were collected at 1, 3, and 6 months of age. Retinal hypoxia was assessed using the bio–reductive drug pimonidazole (Hypoxyprobe–HP), which forms irreversible adducts when pO2<10mmHg. HP–immunoreactivity was analysed using confocal microscopy in retinal flatmounts and sections. HP deposition was quantified using competitive ELISA. mRNA expression of the hypoxia–regulated genes hypoxia–inducible factor–1α (HIF1α) and vascular endothelial growth factor–A (VEGF–A) was quantified using real–time RT–PCR. HIF1α protein levels were assessed using a DNA protein binding assay kit. Vascular density was qualitatively assessed in flatmount retina incubated for ADPase activity (Lutty and McLeod, Arch Oph 110: 267–276, 1992). Results: HP adducts were observed in the inner nuclear and ganglion cell layers in retina of WT mice. HP adduct formation was significantly reduced at 6 months in Rho–/– compared to 1 month (p=0.034), and significantly lower than WT at 3 (p=0.001) and 6 months (p=0.0001), and was unchanged in WT retinae. Hypoxia related gene expression mirrored the pattern of HP–immunoreactivity. Both HIF1α and VEGF–A expression in Rho–/– retina were significantly reduced at 3 (p=0.0001) and 6 months (p=0.0001) compared to 1 month, and to WT (3mth:p=0.0001, 6mth:p=0.0001). HIF1α protein levels corresponded with mRNA expression. At 6 months, retina from the Rho–/– displayed vascular attenuation in capillary beds. Conclusions: This study has demonstrated that hypoxia occurs in the normal retina, which may account for constitutively high levels of VEGF and HIF1α. Photoreceptor loss with RP may reduce retinal oxygen usage and levels of relative hypoxia, which in turn leads to attenuation of the retinal vasculature.
This PDF is available to Subscribers Only