Retinopathy in Mice Induced by Disrupted All-trans-retinal Clearance

Abstract

The visual (retinoid) cycle is a fundamental metabolic process in vertebrate retina responsible for production of 11-cis-retinal, the chromophore of rhodopsin and cone pigments. 11-cis-Retinal is bound to opsins, forming visual pigments, and when the resulting visual chromophore 11-cis-retinylidene is photoisomerized to all-trans-retinylidene, all-trans-retinal is released from these receptors. Toxic byproducts of the visual cycle formed from all-trans-retinal often are associated with lipofuscin deposits in the retinal pigmented epithelium (RPE), but it is not clear whether aberrant reactions of the visual cycle participate in RPE atrophy, leading to a rapid onset of retinopathy. Here we report that mice lacking both the ATP-binding cassette transporter 4 (Abca4) and enzyme retinol dehydrogenase 8 (Rdh8), proteins critical for all-trans-retinal clearance from photoreceptors, developed severe RPE/photoreceptor dystrophy at an early age. This phenotype includes lipofuscin, drusen, and basal laminar deposits, Bruch's membrane thickening, and choroidal neovascularization. Importantly, the severity of visual dysfunction and retinopathy was exacerbated by light but attenuated by treatment with retinylamine, a visual cycle inhibitor that slows the flow of all-trans-retinal through the visual cycle. These findings provide direct evidence that aberrant production of toxic condensation byproducts of the visual cycle in mice can lead to rapid, progressive retinal degeneration.What discriminates the eye from other organs is its light sensitivity and associated metabolic transformations that restore the light-sensitive chromophore (1). It is unclear if aberrations in the visual cycle and byproduct accumulation could be an underlying cause of retinopathy or merely a nonspecific nonpathogenic reflection of impaired metabolism. To distinguish between these two possibilities, we disrupted two genes critical for clearance of light-generated all-transretinal from rhodopsin and cone visual pigments (2, 3). Both the photoreceptor-specific ATP-binding cassette transporter (ABCA4) (4) and all-trans-retinol dehydrogenases (RDHs) 2 are involved in removal of all-trans-retinal from photoreceptors (5) (Scheme 1).ABCA4, also known as ABCR or the rim protein, localizes to the rim of photoreceptor discs and transfers all-trans-retinal from the inside to the outside of disc membranes once alltrans-retinal is released from visual pigments (4). Diretinoidpyridinium-ethanolamine (A2E) (6, 7) and retinal dimer (RALdi) conjugates (8) are the major fluorophores of lipofuscins produced from all-trans-retinal. Even in the presence of a functional transporter, both A2E and RALdi can accumulate as a consequence of aging (9) and produce toxic effects on RPE cells (10, 11). Patients affected by age-related macular degeneration (AMD), Stargardt disease with a disabled ABCA4 gene, or other retinal diseases associated with lipofuscin accumulation develop retinal degeneration. ABCA4 mutations also are linked with a high risk of AMD (12). However, no such ...

Enzymatic Degradation of A2E, a Retinal Pigment Epithelial Lipofuscin Bisretinoid

“…Measured as a decline in amounts of A2E, the therapeutic effect observed with small molecules that target the visual cycle in mouse models have ranged from 30% to 60%. 12a,b,21 …”

“…These approaches have included gene therapies based on viral vector mediated delivery of the wild-type gene 11 and systemic administration of compounds that limit the retinoid cycle so as to decrease all- trans -retinal formation. 12 Nevertheless, neither gene nor drug therapy can reverse the accumulation of lipofuscin bisretinoids such as A2E, once it has already occurred. Thus, in in vitro experiments described here, we have tested an additional treatment method.…”

Serum levels of lipid metabolites in age‐related macular degeneration

“…Such mice used in this study were homozygous for the Leu450 allele of Rpe65 as determined by a published genotyping protocol (36) and free of the Crb1/rd8 (37) and rd/rd (38) mutations. All mice were genotyped by PCR with the following primers: ABCR1 (59-GCCCAGTGGTCGATCTGTCTAGC-39) and ABCR2 (59-CGGACACAAAGGCCGCTAGGACCACG-39) for wildtype (619 bp) and A0 (59-CCACAGCACACATCAGCATTTCT-CC-39) and N1 for the targeted deletion (455 bp) as published previously (29). Animals were provided with standard chow (LabDiet 5053; Purina Mills St. Louis, MO, USA) and maintained under a 12/ 12-h light-dark cycle.…”

“…Though genetically manipulated rodents and other small rapidly reproducing animals do not have maculae, they can serve as limited surrogate models of photoreceptor/retinal pigmented epithelium degeneration with drusen and lipofuscin accumulation similar to that observed with humans with AMD. One of these models is a mouse with a double knockout of both the ATP-binding cassette transporter 4 and retinol dehydrogenase 8, a transporter and enzyme involved in alltrans-retinal clearance (29). In this model, the pathology and death of photoreceptors can be induced in a cohort of animals by simple exposure to light in a synchronized manner.…”

Two-photon microscopy reveals early rod photoreceptor cell damage in light-exposed mutant mice

“…Brief exposure of Abca4 −/− Rdh8 −/− mice to intense light results in acute retinal degeneration, which allows investigators to follow the precise sequence of degenerative events at both a cellular and molecular level (18,29). Notably, such retinal degeneration can be prevented by inhibition of atRAL production with retinylamine, a retinoid cycle inhibitor (30), or by sequestration of atRAL by producing Schiff-base adducts of atRAL with drugs containing a primary amine group (12,29).…”