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Changdong Liu, Yu Wang, Shutong Cao, Rosalyn Le, David P. Bingaman, Richard Ornberg, Naj Sharif, Carl Romano; Natural History of Collagen and Endothelial Markers in Mouse Laser-induced CNV/Subretinal Fibrosis. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1786.
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Subretinal fibrosis (SRF) secondary to choroidal neovascularization (CNV) in patients with age-related macular degeneration (AMD) is linked to vision loss and possibly non-responsiveness to anti-VEGF therapies. Laser-induced CNV in mice also is associated with SRF. Therefore, the natural history of collagen types and vascular endothelial markers was investigated in the mouse CNV/SRF model over a three month time-course.
Laser-induced rupture of Bruch’s membrane was performed in 4 and 10 month old C57BL/6J mice. Eyes were collected at 7, 14, 30, and 90 days post-laser following systemic perfusion with fluorescein-labeled dextran, and fixed in 4% paraformaldehyde. Choroidal flat mounts were labeled with anti-Col IV/or anti-Col I antibodies and biotinylated Iso-B4 lectin I followed by secondary labeling with Alexa 594 and streptavidin-conjugated DyLight 649. Fluorescence image analysis was used to segment the hyperfluorescence of each label and determine the mean area of label/lesion.
Laser-induced CNV/SRF lesions did not regress over 90 days in 4 & 10 month old mice. Col IV and the endothelial marker, Iso-B4-lectin, were consistently expressed at higher levels than Col I at all time points in both age groups. Col IV and endothelial markers did not show a significant change over time in 4 month old mice. However, a time-dependent reduction of these markers was observed in 10 month old mice, where FITC labeling was significantly decreased at post-laser day 30 (↓58.9%, P<0.05) and day 90 (↓46.6%, P<0.05) compared to day 7.
Laser-induced CNV/subretinal fibrosis persisted for 3 months in young and old mice. Vascular endothelial and fibrotic markers were increased by post-laser day 7 and sustained through day 90. A time-dependent reduction in these markers was observed in older, but not younger, mice. The robust staining of Col IV at all time points in both age groups suggests that it may be a viable biomarker for SRF in this model. These findings correlate with the presence of Col IV found in SRF samples from AMD patients (data not shown).
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