Immunological protein expression profile in Ccl2/Cx3cr1 deficient mice with lesions similar to age-related macular degeneration

Age-related macular degeneration (AMD) is the leading cause of blindness in the United States. Ccl2 knock-out (KO) mice sporadically develop the cardinal features of AMD in their senescent stage. Humans bearing a loss of function variant or single nucleotide polymorphism (SNP) in CX3CR1 are at increased risk of developing AMD. We recently developed Ccl2−/−/Cx3cr1−/− mice, which consistently develop retinal degeneration with many AMD features. Since there is strong evidence for an immunological role in AMD pathogenesis, we examined ocular immune protein expression levels in Ccl2−/−/Cx3cr1−/−, Ccl2−/−, Cx3cr1−/−, and age-matched wild-type (WT) mice. Immunohistochemistry revealed increased complement C3d in Bruch's membrane, retinal pigment epithelium (RPE), choroidal capillaries, and particularly drusen of the Ccl2−/−/Cx3cr1−/− mice relative to the WT controls. No change was detected in single KO mice. Real-time RT-PCR revealed a 2.5-fold increase in C3 expression in the Ccl2−/−/Cx3cr1−/−. While the retinas of four month old WT and Ccl2−/− showed minimal immunoreactivity for markers of macrophages and microglia, infiltrates of these mononuclear phagocytic cells were detected in the Ccl2−/−/Cx3cr1−/−retinal lesions and a few foci in the Cx3cr1−/− retina. The Ccl2−/−/Cx3cr1−/− had reduced toll-like receptor 4 (TLR4) expression in the RPE. Following LPS injection, the Ccl2−/−/Cx3cr1−/− had significantly reduced endotoxin-induced uveitis scores and showed a diminished increase in Tlr4 mRNA expression. No changes in TLR4 expression were detected in either single KO. Autoantibodies against the retina and photoreceptors were also detected in the Ccl2−/−/Cx3cr1−/− serum. Real-time RT-PCR revealed significant increases in Ccl5 transcript in the Ccl2−/−/Cx3cr1−/− relative to the WT. These results suggest that innate immunity and possibly adaptive immunity play an important role in Ccl2−/−/Cx3cr1−/− retinal degeneration. Moreover, since human AMD patients show similar immunopathological profiles, these results support the Ccl2−/−/Cx3cr1−/− as a suitable model for human AMD.