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S. E. Barker, A. J. Smith, U. F. O. Luhmann, J. W. B. Bainbridge, R. E. MacLaren, R. R. Ali; Development of a Murine Model of AMD by Overexpression of Complement Component 3 in the RPE of MCP-1-/- and Wild-Type Mice. Invest. Ophthalmol. Vis. Sci. 2008;49(13):207.
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Age-related macular degeneration (AMD) is the most common cause of late-onset blindness in the developed world. The disease is characterised by photoreceptor degeneration in the macula, secondary to pathological changes in the RPE. Recently, the importance of the immune system in disease pathogenesis has became apparent. Aged monocyte chemoattractant protein-1 (MCP-1) knock-out mice have been shown to develop AMD-like symptoms such as drusen and, in some mice, choroidal neovascularisation, but disease progression is slow and penetration of some symptoms is incomplete. This model would provide a more useful research tool if it were possible to induce pathological changes in younger mice.
The cDNA encoding murine C3 was cloned into a HIV-1 based lentiviral vector, which transduces the RPE after subretinal injection. Vector-driven C3 production was demonstrated by western blot, and protein function was tested in a complement-mediated erythrocyte lysis assay. Two microlitres of LNT-C3 was injected subretinally in the superior hemisphere of wild-type or MCP-1-/- mice, while control mice received LNT-GFP or PBS. Fundoscopy to detect gross changes and fluorescein angiography to determine changes in vessel permeability was carried out after 9 -12 months. Eyes were harvested and retinal flatmounts were prepared to identify areas of autofluorescence. The remaining eyes were cryosectioned and stained for C3, vitronectin and C5.
In vitro data showed that LNT-C3 produced functional protein. Fundoscopy showed equivalent responses to injection of vector in wild-type and MCP-1-/- mice and that LNT-C3 did not appear to cause gross RPE damage. Fluorescein angiography showed some changes in the vascular permeability following administration of LNT-C3 in wild-type and MCP-1-/- mice compared with controls. Histological analysis showed there was some degeneration of photoreceptor layers and thickening of the RPE around the site of injection in a proportion of the MCP-1-/- mice.
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