Cholestenoic Acid is an important elimination product of cholesterol in the retina: comparison of retinal cholesterol metabolism with that in the brain

Abstract

Purpose: Accumulating evidence indicates a link between cholesterol and age-related macular degeneration. Yet, little is known about cholesterol elimination from the retina and retinal pigment epithelium (RPE), the two layers that are damaged in this blinding disease. Several different pathways of enzymatic cholesterol removal exist in extraocular tissues. The authors tested whether metabolites from these pathways could also be quantified in the bovine and human retina and RPE. For comparison, they measured cholesterol oxidation products in two regions of the bovine and human brain and in the bovine liver and adrenal glands.

Methods: Sterol quantification was carried out by isotope dilution gas chromatography-mass spectrometry. Bovine tissues were used first to optimize analytical procedures and to investigate postmortem changes in oxysterol concentrations. Then human specimens were analyzed for oxysterol concentrations.

Results: Qualitatively, oxysterol profiles were similar in the bovine and human tissues. In the human retina and RPE, the authors could not detect 27-hydroxycholesterol but unexpectedly found that its oxidation product, 5-cholestenoic acid, is the most abundant oxysterol, varying up to threefold in different persons. 24S-Hydroxysterol and pregnenolone were also present in the retina, but at much lower quantities and without significant interindividual variability. In the brain, the predominant oxysterol was 24S-hydroxycholesterol.

Conclusions: The oxysterol profile of the retina suggests that all known pathways of cholesterol elimination in extraocular organs are operative in the retina and that they likely vary depending on specific cell type. However, overall oxidation to 5-cholestenoic acid appears to be the predominant mechanism for cholesterol elimination from this organ.

Figures

Figure 1.

Schematic representation of cholesterol elimination pathways in different extrahepatic organs and cytochrome P450 enzymes involved in these pathways. Bold lines and bold font indicate the major contributors and products of cholesterol elimination.

Figure 2.

Flow chart outlining sample processing for quantification of different oxysterols.

Figure 3.

Effect of death-to-preservation interval on oxysterol concentrations in bovine retina, RPE, and choroid. Open bars: concentrations of the free sterols; dashed bars: concentrations of the total sterols. White: cholesterol; pink: 27-COOH; green: 22-OH; orange: Preg; blue: 24-OH.

Figure 4.

Oxysterol concentrations in different bovine organs. White: cholesterol; pink: 27-COOH; green: 22-OH; orange: Preg; blue: 24-OH.

Figure 5.

Oxysterol concentrations in the retina and RPE of donors 8, 9, and 11. White: cholesterol; pink: 27-COOH; green: 22-OH; orange: Preg; blue: 24-OH.

Figure 6.

Oxysterol concentrations in two regions of the brains of donors 3 and 4. White: cholesterol; pink: 27-COOH; green: 22-OH: orange: Preg; blue: 24-OH; violet: 27-OH.

Figure 7.

Expression of cholesterol-metabolizing P450 in different cell types, as assessed by immunohistochemical studies.– Illustration of the seven main classes of cell types and cell layers found in the vertebrate retina is reprinted by permission from Macmillan Publishers Ltd. Dyer MA, Cepko CL. Regulating proliferation during retinal development. Nat Rev Neurosci. 2001;2:333–342. Copyright 2001. RPE, retinal pigment epithelium; ONL, outer nuclear layer; OPL, outer plexiform layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer; NFL, neural fiber layer.