Lipid transport, oxidation and toxicity in the retina

National Eye Institute

Investigators

Linked publications & trials

Abstract

Recently, research done at the MRDS has demonstrated that the retina uptakes much of its lipids from circulating low density lipoproteins (LDL) via LDL-receptors in the RPE and choriocapillaris (4). Inside the RPE the lipids are processed and delivered as HDL-like particles to HDL-receptors (SR-BI and SR-BII) in the photoreceptor cells (3) and other cells of the inner retina. We have demonstrated that the retina expresses all of the main genes involved in the well-known systemic reverse cholesterol transport pathway. The retina has adapted this pathway to sustain its own particular needs for lipid transport and turnover by controlling the expression and location of the different transporters and receptors in the pathway (3). We hypothesize that the retina requires a high turnover of lipids because of the high susceptibility of this class of molecules to oxidation and particularly to photooxidation. One of the main priorities of the MRDS is to identify the mechanism by which oxidized lipids, which may be highly toxic, are metabolized and excreted from the retina. We are presently pursuing this project by generating transgenic rats with RPE and photoreceptor-specific targeted overexpression and knockdowns of selective genes. [unreadable] [unreadable] Another related area of interest is the formation of oxidized lipids and their cytotoxicity. One particular molecule, 7-ketocholesterol, is of particular interest because it is known to be highly cytotoxic to various cell types and is the major toxic component in atherosclerotic plaques. This oxysterol is formed by copper and/or iron mediated oxidation of cholesterol-esters in LDL deposits but is also known to be readily formed by the photooxidation of cholesterol. The toxicity of 7-ketocholesterol is due to its ability to form destabilizing crystals in mitochondrial membranes leading to cytochrome-c leakage and apoptosis. Our studies have found small amounts of 7-ketocholesterol in normal primate retina mostly associated with oxidized LDL deposits in Bruchs membrane and choriocapillaris (unpublished results). However, in light-damaged albino rats 7-ketocholesterol levels greatly increase throughout the retina especially in the ganglion cells, RPE and photoreceptor inner segments (unpublished results). This suggests that photooxidation is a plausible mechanism for generating 7-ketocholesterol in the retina. Moreover, this also suggests that chronic mitochondrial damage due to 7-ketocholesterol formation could be a factor in aging diseases of the retina. We have also found that 7-ketocholesterol can induce VEFG release from cultured RPE cells and from the RPE and choriocapillaris of rats injected with oxidized LDL. This may be a potential mechanism for some forms of AMD that are characterized by choroidal neovascularization. [unreadable] [unreadable] Our experiments have also shown that lipoproteins like LDL can be readily photoxidized forming a mixture of oxidized lipids of similar composition to those found in atherosclerotic plaques. The polyunsaturated fatty acids are also of interest because they are components of numerous lipid classes and are highly oxidizable at their double bonds. Docosahexaenoic acid (DHA) is of particular interest since it comprises approximately 50% of the lipids in the outer segment membranes. This fatty acid is highly susceptible to photooxidation and its transport and metabolism in the retina is not fully understood.[unreadable] [unreadable] Enzymes that protect the mitochondria from oxidative damage or from 7-ketocholesterol toxicity are also a major area of interest. The cytochrome P450 CYP27A1 is a sterol hydroxylase capable of hydroxylating 7-ketocholesterol and is highly expressed in the mitochondria of rod photoreceptors and RPE (1). 7-Ketocholesterol when hydroxylated at the side chain carbons loses its toxicity (1). Sulfation of 7-ketocholesterol is another potential mechanism of detoxification and very small amounts of sulfated 7-ketocholesterol have been dettected in primate retina. Although hydroxylation seems to be the major detoxification pathway for 7-ketocholesterol in the retina, sterol sulfation may be an important regulatory feed-back mechanism to provide antagonists to the liver X-receptors (LXRs). LXRs are transcription factors that regulate many of the genes in the reverse cholesterol transport pathway and are highly expressed in the retina. It is well known that oxysterols are agonists of LXRs while some sulfated sterols are antagonists. We are presently searching for a retinal-expressed sulfotransferase capable of sulfating 7-ketocholesterol under physiological conditions. [unreadable] [unreadable] Another group of protective enzymes of interest to the MRDS are the methionine sulfoxide reductases (MSRs). These groups of enzymes (MSRAs and MSRBs) convert oxidized methionines in proteins back to methionine often resulting in the restoration of lost function. Overxpression of MSRA in the fruit fly increased their lifespan and fertility. Mice lacking MSRA have greatly reduced lifespans and are highly susceptible to oxidative damage. Previous studies on MSRA suggest that it play a key role in aging and age-related disease. We have found that MSRA is highly expressed in the macular RPE (2). The retina contains high levels of both MSRA and MSRB activities (2). We also discovered that MSRA is controlled by two distinct promoters. One promoter (P1) makes an mRNA whose product is targeted to the photoreceptor synaptic mitochondria and the other promoter (P2) makes two mRNAs whose products are targeted to the cytosol and nucleus. The P2 promoter is highly expressed in RPE cells. We are presently characterizing the P2 promoter and have identified several relevant transcription factors involved in regulating its expression. [unreadable] [unreadable] In summary the MRDS is pursuing several basic research projects investigating lipid transport, oxidation and protective mechanisms with the goal of obtaining a better understanding of the processes involved in aging and the pathogenesis mechanisms of diseases like the age-related macular degenerations.