Early AMD-like defects in the RPE and retinal degeneration in aged mice with RPE-specific deletion of or .

PURPOSE

To examine the effects of autophagy deficiency induced by RPE-specific deletion of or in mice as a function of age.

METHODS

Conditional knockout mice with a floxed allele of or were crossed with inducible transgenic mice. -directed RPE-specific Cre recombinase expression was induced with doxycycline feeding in the resulting mice. Cre-mediated deletion of floxed or resulted in RPE-specific inactivation of the or gene. Plastic and thin retinal sections were analyzed with light and electron microscopy for histological changes. Photoreceptor outer segment (POS) thickness in plastic sections was measured using the Adobe Photoshop CS4 extended ruler tool. Autophagic adaptor p62/SQSTM1 and markers for oxidatively damaged lipids, proteins, and DNA were examined with immunofluorescence staining of cryosections. Fluorescence signals were quantified using Image J software.

RESULTS

Accumulation of p62/SQSTM1 reflecting autophagy deficiency was observed in the RPE of the and mice. 3-nitrotyrosine, advanced glycation end products (AGEs), and 8-hydroxy-2'-deoxyguanosine (8-OHdG), markers for oxidatively damaged proteins and DNA, were also found to accumulate in the RPE of these mice. We observed retinal degeneration in 35% of the mice and 45% of the mice at 8 to 24 months old. Degeneration severity and the number of mice with degeneration increased with age. The mean POS thickness of these mice was 25 µm at 8-12 months, 15 µm at 13-18 months, and 3 µm at 19-24 months, compared to 35 µm, 30 µm, and 24 µm in the wild-type mice, respectively. Early age-related macular degeneration (AMD)-like RPE defects were found in all the and mice 13 months old or older, including vacuoles, uneven RPE thickness, diminished basal infoldings, RPE hypertrophy/hypotrophy, pigmentary irregularities, and necrosis. The severity of the RPE defects increased with age and in the mice with retinal degeneration. RPE atrophy and choroidal neovascularization (CNV) were occasionally observed in the and mice with advanced age.

CONCLUSIONS

Autophagy deficiency induced by RPE-specific deletion of or predisposes but does not necessarily drive the development of AMD-like phenotypes or retinal degeneration.