Oxysterols induced inflammation and oxidation in primary porcine retinal pigment epithelial cells

Corinne Joffre, Laurent Leclère, Bénédicte Buteau, Lucy Martine, Stéphanie Cabaret, Laure Malvitte, Niyazi Acar, Gérard Lizard, Alain Bron, Catherine Creuzot-Garcher, Lionel Bretillon
Current Eye Research. 2007-01-01; 32(3): 271-280
DOI: 10.1080/02713680601187951

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1. Curr Eye Res. 2007 Mar;32(3):271-80.

Oxysterols induced inflammation and oxidation in primary porcine retinal pigment
epithelial cells.

Joffre C(1), Leclère L, Buteau B, Martine L, Cabaret S, Malvitte L, Acar N,
Lizard G, Bron A, Creuzot-Garcher C, Bretillon L.

Author information:
(1)Eye and Nutrition Research Group, INRA, UMR1129 FLAVIC, Dijon, France.

PURPOSE: Aging is associated with an accumulation of cholesterol esters in the
Bruch membrane. Cholesterol esters are prone to undergo oxidation and generate
oxysterols that have cytotoxic and proinflammatory properties. We investigated
the effects of three oxysterols on mitochondrial dysfunctions, inflammation, and
oxidative stress in primary cultures of porcine retinal pigment epithelial (RPE)
METHODS: RPE cells were incubated with oxysterols (50 micro M of
24-hydroxycholesterol, 25-hydroxycholesterol, or 7-ketocholesterol) for 24 hr and
48 hr. Oxysterol content was determined in cells and in corresponding media by
gas chromatography. Mitochondrial activity was measured by mitochondrial
dehydrogenase activity. The intracellular formation of reactive oxygen species in
RPE cells was detected by using the fluorescent probe DCFH-DA. IL-8 was assayed
in the supernatants by ELISA, and the corresponding cellular transcripts were
semiquantified by RT-PCR.
RESULTS: Analyses of the oxysterols content in the RPE cells and corresponding
media suggested a high rate of cellular uptake, although some differences were
observed between 7-ketocholesterol on the one hand and 24-hydroxycholesterol and
25-hydroxycholesterol on the other hand. All oxysterols induced slight
mitochondrial dysfunctions but a significant 2- to 4-fold increase in reactive
oxygen species (ROS) production compared with the control. They also enhanced
IL-8 gene expression and IL-8 protein secretion in the following decreasing
order: 25-hydroxycholesterol > 24-hydroxycholesterol > 7-ketocholesterol.
CONCLUSIONS: We conclude that in confluent primary porcine RPE cells,
24-hydroxycholesterol, 25-hydroxycholesterol, and 7-ketocholesterol are potent
inducers of oxidation and inflammation.

DOI: 10.1080/02713680601187951
PMID: 17453947 [Indexed for MEDLINE]

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