Mechanisms of Retinal Diseases Section
The Mechanisms of Retina Diseases Section (MRDS) performs basic research on the biology of the retina especially regarding lipid transport, oxidation and cytotoxicity. The section is focused on determining the role of oxidized lipids on the pathogenesis of age-related macular degeneration (AMD).
The MRDS has shown that the retina uptakes circulating LDL (2, 6) and distributes the lipids to the different cellular layers (6). To perform this lipid uptake and transport, the retina expresses the same proteins used by the systemic "reverse cholesterol pathway" (7, 13). Monkey retina has been found to express apoA1, apoE, apoB, ABCA1, LDLR, SR-BI and II, CD36, CETP, and LCAT (7). The particular locations where these molecules are expressed within the retina suggest an internal lipid transport based on HDL-like lipid particles (7, 13). The MRDS is presently investigating this pathways and mechanisms.
Recently the MRDS has found that the retina contains significant levels of the highly toxic oxysterol 7-ketocholesterol (10, 11). In the monkey retina 7-ketocholesterol is found in association with oxidized lipoprotein deposits in the choriocapillaris and Bruch's membrane (10). However, in photodamaged rats 7-ketocholesterol is found in areas of high mitochondrial content especially the RPE, photoreceptor inner segments and ganglion cells (11). Intermediates identified in these rats by LCMS indicate that the 7-ketocholesterol was formed via a free radical mediated mechanism. This mechanism requires a transition metal catalyst which is this case is likely Fe+2 (11). The source of the iron was not conclusively identified but light is known to cause the release of iron from ferritin and possibly cytochrome c. The source of the iron and mechanisms of its release are being actively investigated.
7-Ketocholesterol is found in very high levels in atherosclerotic plaques and is suspected of causing foam cell formation and toxicity to vascular endothelial cells. In cultured human RPE and vascular endothelial cells the MRDS has found that 7-ketocholesterol is a potent inducer of VEGF (10) and other cytokines including IL-6 and IL-8 (13). The pharmacological properties of 7-ketocholesterol are complex and seem to be dose dependent. At low doses 7-ketocholesterol is pro-inflammatory while at higher doses it can induce cell death by necrosis or apoptosis depending on the cell type. In most cell types 7-ketocholesterol-induced inflammation is dependent on reactive oxygen species (ROS) formation. However, in cultured RPE-derived cells the pro-inflammatory pathway seems to be independent of ROS (13). The MRDS is actively investigating the 7-ketocholesterol-mediated inflammatory and death pathways in the retina in vivo. In addition, the MRDS is testing the anti-angiogenic properties of certain antagonist to 7-ketocholesterol inflammation using a CNV rat model (laser treated).
The metabolism of 7-ketocholesterol by the neural retina and RPE are another topic of interest to the MRDS. The retina expresses significant levels of the mitochondrial CYP27A1 (5), a cytochrome P450 enzyme capable of hydrolylating 7-ketocholesterol at the side chain 27-carbon. This hydroxylated form of 7-ketocholesterol is non-toxic (5) but repeated attempt at detecting it retinal extracts have been unsuccessful. Two other enzymes capable of metabolizing 7-ketocholesterol have been investigated, the 24-cholesterol hydroxylases (CYP46A1) and the sulfotransferases (SULT2B1). However, both of these enzymes are present in the retina only in trace amounts and their potential metabolites are also undetectable (13). The retina seems to metabolize 7-ketocholesterol by excreting it to circulation. Preliminary evidence suggests transporters such as ABCG1 which is present in the basal aspect of the RPE may be involved in the elimination of 7-ketocholesterol. Once in circulation 7-ketocholesterol is known to be quickly metabolized by the liver.
Another area of interest to the MRDS are anti-oxidative enzymes specially those that protect the mitochondria. In this regard we have investigated the retinal expression of peroxiredoxin 3 (8) and the methionine sulfoxide reductases (MSRs) (4, 9, 12). These are enzymes capable of reversing the oxidation of methionine in proteins and thus preserving function. There are two major forms of MSRs categorized by their stereospecificity on methionine sulfoxide (MetO). The MSRAs recognize the Met(S)O and the MSRBs recognize the Met(R)O stereoisomers. These enzymes have been shown to be important in preserving the health of cells and in the aging process. The MSRs are highly expressed in the retina especially in the macular region (4, 9). These enzymes protect cultured cells from multiple forms of oxidative damage including damage caused by 7-ketocholesterol (12). The MRDS is actively investigating the role of these enzymes in preventing the light-mediated iron release from protein like ferritin.
|Ignacio R. Rodriguez
|Jung Wha Lee||Staff Scientistfirstname.lastname@example.org|
|Iranzu Pascual||Visiting Fellowemail@example.com|
|Jian-dar Huang||Visiting Fellowfirstname.lastname@example.org|
|Juam Amaral||Staff Scientistemail@example.com|
1. Rodriguez, I.R. Rapid analysis of oxysterols by HPLC and UV spectroscopy. Biotechniques, 36(6): 952-958, 2004
2. Gordiyenko, N., Campos, M., Lee, J.W., Fariss, R.N., Sztein J., and Rodriguez, I.R. RPE cells internalize low density lipoprotein (LDL) and oxidized LDL in large quantitites in vitro and in vivo. Invest. Ophthalmol. & Vis. Sci. 45(8): 2822-2829, 2004
3. Rodriguez, I.R., Alam, S., Lee, J.W. Cytotoxicity of oxidized low density in cultured RPE cells is dependent on the formation of 7-ketocholesterol. Invest. Ophthalmol. & Vis. Sci. 45(8): 2830-2836, 2004
4. Lee, J.W., Gordiyenko, N.V., Marchetti, M., Tserentsoodol, N., Sagher, D., Deamicis, C., Alam, S., Weissbach, H., Kantorow, M., and Rodriguez, I.R. Gene structure, localization and role in oxidative stress of methionine sulfoxide reductase A (msrA) in the monkey retina. Exp. Eye Res. 82: 816-827, 2006
5. Lee J.W., Fuda, H., Javitt, N., Strott, C., and Rodriguez, I.R. Expression and localization of the sterol 27-hydroxylase (CYP27A1) in monkey retina. Exp. Eye Res. 83: 465-469, 2006
6. Tserentsoodol, N., Sztein, J., Campos, M., Fliesler, S.J., Gordiyenko, N.V., Fariss, R.N., Lee, J.W., and Rodriguez, I.R. Uptake of cholesterol by the retina occurs primarily via an LDL-receptor mediated process. Mol. Vis. 12:1306-1318, 2006
7. Tserentsoodol, N., Gordiyenko, N.V., Pasual, I., Lee, J.W., Fliesler, S.J., and Rodriguez, I.R. Internal lipid transport in the retina is dependent on HDL-like particles and class B scavenger receptors. Mol. Vis. 12:1319-1333, 2006
8. Moreira, E.F., Kantorow, M., and Rodriguez, I.R. Peroxiredoxin 3 (PDRX3) is highly expressed in the primate retina especially in blue cones. Exp. Eye Res. 86:452-455, 2007
9. Pascual I., Larrayoz, I.M., and Rodriguez, IR. All-trans retinoic acid regulates the human methionine sulfoxide reductase A (MSRA) gene via two distinct promoters Genomics 93: 62-71, 2008
10. Moreira, E.F., Larrayoz, I.M., Lee, J.W., and Rodriguez, I.R. 7-Ketocholesterol is present in lipid deposits in the primate retina: potential implications in the induction of VEGF and CNV formation. Invest Ophthalmol Vis Sci. 50:523-532, 2009
11. Rodriguez, I.R. and Fliesler, S.J. Photo-damage generates 7-keto- and 7-hydroxycholesterol in the rat retina via a free radical mediated mechanism. Photochem Photobiol. (in press)
12. Pacual, I., Larrayoz, I.M. and Rodriguez, I.R. Methionine sulfoxide reducatse B2 (MSRB2) is highly expressed in the retina and protects RPE cells from oxidative damage. (submitted)
13. Rodriguez, I.R. and Larrayoz, I.M. Cholesterol Oxidation in the Retina: Implications of 7-Ketocholesterol Formation in Chronic Inflammation and Age-Related Macular Degeneration. (review, submitted)