0000000000717778
AUTHOR
Thomas Montange
Activation of a caspase-3-independent mode of cell death associated with lysosomal destabilization in cultured human retinal pigment epithelial cells (ARPE-19) exposed to 7beta-hydroxycholesterol
International audience; Purpose: To characterize the possible cytotoxic effects of oxysterols (7-hydroxycholesterol (7-OH), 25-hydroxycholesterol (25-OH)) in human retinal pigment epithelial cells (ARPE-19) and to detail the relationships between some of these effects. Methods: ARPE-19 cells were treated with 7-OH and 25-OH. Cell viability was measured with the MTT assay. Membrane permeability, mitochondrial potential, and lysosomal integrity were measured by flow cytometry with propidium iodide, DiOC6(3), and acridine orange, respectively. Cell death was characterized by staining with Hoechst 33342, transmission electron microscopy, and analysis of the DNA fragmentation pattern. Caspase ac…
Cytométrie en flux, microbilles et analyses biomoléculaires multiplexes
Resume En cytometrie en flux, les microbilles constituent des outils importants. Initialement developpees pour optimiser les signaux de diffraction et de fluorescence detectes par les cytometres, les microbilles sont maintenant largement utilisees pour la quantification d'antigenes. Actuellement, leur utilisation comme support reactionnel permet aussi de detecter et de quantifier simultanement par cytometrie en flux plusieurs analytes tels que des proteines, des nucleotides, et des ions, mais aussi des agents infectieux dans quelques microlitres d'echantillon. Pour cela, des microbilles de fluorescences et/ou de tailles differentes couplees a des proteines ou a des oligonucleotides sont le …
Assessment of the toxicity of cholesterol oxides on retinal pigment epithelial cultured cells
International audience; Purpose: Among the retinal lipid deposits found in AMD (age related macular degeneration), esterified and unesterified cholesterol have been identified. Cholesterol that accumulates in Bruch's membrane could be oxidized and lead to cholesterol oxides known as oxysterols. These compounds have cytotoxic activities on many vascular cell types, and we conducted this study to evaluate the cytotoxics effects of oxysterols on retinal pigment epithelial (RPE) cells. Methods: Human RPE cells (ARPE–19) were cultured and treated by 7–ketocholesterol (7–keto), 7–betahydroxycholesterol (7–beta) and 25–hydroxycholesterol (25–OH). ARPE–19 cells viability was determined with MTT ana…
Toxicity of Branched-Chain Fatty Acids in Conjunctival Cell Line
International audience; Purpose:: Tear film lipid composition is modified in patients suffering from chronic blepharitis. We previously found that branched chain fatty acids (BCFA) were significantly increased in tears of these patients, suggesting that BCFA may be involved in the disease. The toxicity of different BCFA as potent inducers of apoptosis has already been established in various cell lines but not in the conjunctiva. The aim of this work was to assess the toxic effects of BCFA in cultures of Chang conjunctival cells. Methods:: Chang’s human conjunctival cells were cultured during 4 or 24h with various concentrations of IsoC16 and IsoC20 as BCFA (from 50 to 200µM), by comparison …
Analogies between atherosclerosis and age-related maculopathy: expected roles of oxysterols
The pathogenesis of age-related macular degeneration (ARMD) is not clearly understood. Like other age-related diseases, it is associated with abnormal deposits called drusen. These drusens are localized in Bruch's membrane. Recent investigations have shown a link between drusen formation and inflammatory and immunologic reactions. The involvement of oxidative stress is supported by available data as an important contributing factor in the developement of ARMD. The data regarding the nature and the source of the deposits suggest that ARMD may share similar pathways with atherosclerosis. The role of oxydized products of cholesterol, the oxysterols, in the pathogenesis of atherosclerosis is we…
Absence of oxysterol-like side effects in human monocytic cells treated with phytosterols and oxyphytosterols
Oxysterols, found in some commonly consumed foods, can induce a wide range of cytotoxic effects, which have been extensively studied. On the other hand, the side effects of phytosterols and oxyphytosterols are less well-known. Over the past few years, different types of foods have been enriched with phytosterols on the basis of the properties of these compounds that reduce circulating cholesterol levels in certain experimental conditions. It is therefore important to gain better knowledge of the risks and benefits of this type of diet. In this study, conducted in human monocytic U937 cells, the ability of phytosterols (sitosterol, campesterol) and oxyphytosterols (7 beta-hydroxysitosterol, …
Phospholipidosis and down-regulation of the PI3-K/PDK-1/Akt signalling pathway are vitamin E inhibitable events associated with 7-ketocholesterol-induced apoptosis
International audience; Among the oxysterols accumulating in atherosclerotic plaque, 7-ketocholesterol (7KC) is a potent apoptotic inducer, which favours myelin figure formation and polar lipid accumulation. This investigation performed on U937 cells consisted in characterizing the myelin figure formation process; determining the effects of 7KC on the PI3-K/PDK-1/Akt signalling pathway; evaluating the activities of vitamin E (Vit-E) (α-tocopherol) on the formation of myelin figures and the PI3-K/PDK-1/Akt signalling pathway and assessing the effects of PI3-K inhibitors (LY-294002, 3-methyladenine) on the activity of Vit-E on cell death and polar lipid accumulation. The ultrastructural and b…