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M. R. Powers, M. H. Davies, A. J. Stempel; Bim-Deficient Mice Exhibit Reduced Retinal Vaso-Obliteration and Subsequent Neovascularization in the Mouse Model of Oxygen-Induced Retinopathy. Invest. Ophthalmol. Vis. Sci. 2010;51(13):59.
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Previously our laboratory has demonstrated that the extrinsic apoptotic pathway plays an important role in retinal neovascularization (NV) during oxygen-induced retinopathy (OIR) via death ligands. We used Bim deficient (-/-) mice in the model of OIR to determine if this proapoptotic Bcl-2 family member (BH3-only) contributes to the control of retinal NV via the intrinsic apoptotic pathway.
Bim expression was evaluated in room air and hyperoxia exposed C57BL/B6 (B6) retinas by RT-PCR and Western blot. Vascular development was assessed at postnatal day 5 (P5) and P7 in B6 and Bim-/- mice in lectin stained retinal whole mounts. B6 and Bim-/- mice were exposed to 75% oxygen from P7 to P12. Retinal vaso-obliteration was quantified from lectin-labeled retinas collected at P8 and P12. NV was quantified from H&E stained cross-sections by counting pre-retinal vascular tuft nuclei from P17 and P21 retinas. Qualitative analysis of retinal vascularization was performed in lectin-labeled retinal whole mounts in B6 and Bim-/- mice on P17.
Bim expression was confirmed at both the RNA and protein level in room air and hyperoxia exposed B6 retinas on P8, P12, P17 and P21. There was no significant difference in retinal vascular development in Bim-/- mice compared to controls. At P8, after hyperoxia exposure, 0.7±0.05% of the retina was avascular in Bim-/- mice compared to 39.3±2.6% avascular in B6 mice (p<0.0001; n=6). At P12 following hyperoxia exposure, 9.7±0.4% of the retina was avascular in Bim-/- mice compared to 30.3±1.7% in B6 mice (p<0.0001; n=5-6). Subsequently, at P17, Bim-/- mice exhibited a significant reduction in NV (9.2±1.2 neovascular nuclei per section) compared to B6 mice (25.1±1.2; p<0.0001; n=8-10 eyes). At P21, Bim-/- mice had 5.1±0.4 neovascular nuclei per section compared to 9.8±2.5 in B6 mice (p=0.179; n=6-10 eyes). These findings were qualitatively confirmed in lectin-labeled retinal whole mounts.
The immature retinal vasculature was dramatically protected from vaso-obliteration in Bim-/- mice during hyperoxia exposure. Bim is known to be strongly induced by growth factor withdrawal. We speculate that Bim contributes to retinal endothelial cell death and vaso-obliteration as a result of VEGF down-regulation from hyperoxia. The reduced NV observed during room air recovery in the Bim-/- mice is likely related to a reduction in retinal ischemia as compared to controls. Modulation of the intrinsic apoptotic pathway offers unique therapeutic opportunities for retinal NV.
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