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M. E. Kleinman, B. J. Raisler, A. Takeda, R. J. C. Albuquerque, M. Nozaki, J. Baffi, W. W. Hauswirth, J. Ambati; Prevention and Reversal of Drusen-Like Phenotype in a Mouse Model of Amd. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2354.
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Mice deficient in the monocyte chemoattractant protein Ccl2 or its receptor Ccr2 develop drusen-like basal laminar deposits as they age. We employed targeted gene therapy coupled with bone-marrow transplantation (BMT) to prevent or reverse the early AMD-like phenotype in young and aged mice, respectively.
Adeno-associated virus (AAV) vectors encoding Ccl2 or green fluorescent protein (GFP) were injected subretinally into fellow eyes of 8-9 month-old Ccl2-/- mice with drusen-like spots (n=9) or 4-8 week-old Ccl2-/- mice without drusen-like spots (n=9). BMT from wild-type mice constitutively expressing LacZ was performed in a group of AAV treated Ccl2-/- x Ccr2-/- mice (n=6). Fundus photography, electron microscopy, and immunohistochemical analyses were conducted on treated eyes at 14, 28, and 72 days following viral injections. F4/80+ CD11c- macrophage infiltration into the retina or choroid at 72 days was quantified by flow cytometry.
AAV-mediated expression of Ccl2 prevented and reversed the drusen-like phenotype in Ccl2 deficientmice. Fundus and ultrastructural examinations showed decreased sub-RPE basal laminar deposits and hard drusen-like excrescences in eyes injected with AAV-Ccl2 compared with AAV-GFP. BMT treated animals exhibited similar findings. Immunohistochemistry demonstrated persistent expression of Ccl2 in AAV-Ccl2 injected eyes and the presence of LacZ+ bone-marrow derived cells in the eyes of BMT animals. Significantly more macrophages were mobilized in the AAV-Ccl2 treated eyes compared to AAV-GFP with a predilection for migration to the choroid.
These rescue data confirm a critical role of the Ccl2-Ccr2 signaling axis in the prevention and regression of drusen-like basal laminar deposits. Gene therapy and BMT were able to upregulate Ccl2 expression and demonstrate that the drusen-like phenotype is both preventable and reversible by restoration of macrophage mobilization.
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