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George Hoppe, Alan D. Marmorstein, Eric A. Pennock, Henry F. Hoff; Oxidized Low Density Lipoprotein–Induced Inhibition of Processing of Photoreceptor Outer Segments by RPE. Invest. Ophthalmol. Vis. Sci. 2001;42(11):2714-2720.
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purpose. To examine the effects of oxidized low-density lipoproteins (oxLDL) on
phagocytosis and processing of photoreceptor outer segments (OS) by
retinal pigment epithelial (RPE) cells.
methods. Confluent cultures of RPE-J cells were pretreated with oxLDL or LDL,
and the effects of such treatment on the processing of added OS was
determined. Processing was determined either by the degradation of 125I-labeled OS to trichloroacetic acid-soluble label or by
the cleavage of rhodopsin observed on Western blot analysis of cell
lysates separated by sucrose density gradient fractionation. Binding to
and uptake of OS by RPE-J cells was assessed by determining the
fluorescence of FITC-labeled OS before and after quenching with trypan
results. OxLDL induced a significant decrease in the degradation of 125I-OS in RPE-J cells but no reductions in either binding
or uptake, when a 24-hour recovery period was inserted between
treatment with oxLDL and challenge with OS. Rhodopsin cleavage
increased in a time-dependent manner after phagocytosis of OS by RPE-J
cells. The small guanosine triphosphatase (GTPase), Rab5, was first
found in phagosome fractions containing rhodopsin and its cleavage
products, and at later times of challenge, in more dense fractions
representing phagolysosomes. These were assessed by the colocalization
of rhodopsin cleavage products in density fractions with cathepsin D, a
marker of lysosomes. OxLDL induced a reduction in rhodopsin cleavage
product formation and in phagosome-lysosome fusion (maturation). It
also reduced the time-dependent shift of rhodopsin to higher density
fractions containing more cathepsin D without any detectable reduction
in the expression of cathepsin D or in acid protease activity.
conclusions. OxLDL induces a reduction in the processing of OS by RPE by perturbing
the fusion of lysosomes with phagosomes.
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