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Michael R. Powers, Michael H. Davies, Stephen T. Magill; Altered Vascular Response in MicroRNA 132-212 Knockout Mice in the Model of Oxygen-Induced Retinopathy. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6295.
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MicroRNAs (miR) are small noncoding RNAs that regulate gene expression by silencing target mRNA transcripts. They function in numerous processes, including angiogenesis, cell proliferation and apoptosis. miR-132 acts as a positive angiogenic switch that induces neovascularization (NV). This study investigates the role of miR-132 in pathologic retinal angiogenesis.
miR-132/212-/- (miR-/-) and C57BL/6 (B6) control mice were exposed to 75% oxygen from postnatal day 7 (P7) to P12 and recovered in room air. Eyes from hyperoxia exposed and room air control mice were collected at various time points. Quantitative analysis of retinal vascular development in miR-/- and B6 mice were performed at P7 and P12 from lectin-labeled retinal whole mounts. Retinal vaso-obliteration was quantified from retinal whole mounts at P12, P17 and P21 in both miR-/- and B6 mice. NV was quantified from H&E stained cross-sections by counting pre-retinal vascular nuclei from. Relative levels of VEGF mRNA were quantified by qRT-PCR.
Normal vascular development was altered in miR-/- mice at P7 with 99% of the superficial retina vascularized (n=3) compared to 75% (n=8) in B6 mice. Additionally, there was a small but significant increase in the vascular density of the deep vessels at P12, along with a 2-fold increase in VEGF mRNA levels in miR-/- mice. There was no significant difference in central vaso-obliteration in miR-/- mice compared to B6 mice after hyperoxia-exposure at P12 (n=8-10). Hyperoxia-exposure caused a significant decrease in VEGF levels in both strains, although VEGF expression was relatively higher in miR-/- mice compared to B6 mice. At P17 after hyperoxia exposure, there was no difference in central retinal avascularity, however, there was a significant decrease in NV in miR-/- (13.3±2.8 neovascular nuclei per section) compared to B6 mice (24.3±2.5; p=0.01, n=6-8). At P21, after hyperoxia exposure, miR-/- mice had more neovascular nuclei per section (28.8±3.3) compared to in B6 mice (7.1±1.8; p<0.0001; n=8). Interestingly, miR-/- mice showed delayed re-vascularization of the central retina with 10% avascular area compared to 2% in B6 mice (p=0.0007). Finally, at P24, the miR-/- continued to exhibit a significantly delayed neovascular regression, with 12.5±2.6 preretinal neovascular nuclei as compared to 4.8±1.0 in controls (p=0.02, n=8-10).
The absence of miR-132/212 results in altered retinal vascular development in room air mice. Hyperoxia exposure induced a consistent disease phenotype at P12 in the miR-/- mice. The delayed recovery and resolution of the retinal avascularity and NV in the miR-/- mice suggests a role for miR-132/212 in the retinal vascular response to ischemia.
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