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Oxidized low-density lipoprotein (ox-LDL) binding to lectin-like ox-LDL receptor-1 (LOX-1) in cultured bovine articular chondrocytes increases production of intracellular reactive oxygen species (ROS) resulting in the activation of NF-kappaB.
- Source :
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Osteoarthritis and cartilage [Osteoarthritis Cartilage] 2004 Jul; Vol. 12 (7), pp. 568-76. - Publication Year :
- 2004
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Abstract
- Objective: To examine the effect of oxidized low-density lipoprotein (ox-LDL) on the intracellular production of reactive oxygen species (ROS) in bovine articular chondrocytes (BACs) and to investigate whether this increase occurs through binding to the receptor lectin-like ox-LDL receptor-1 (LOX-1). Furthermore, to ascertain whether the binding of ox-LDL to LOX-1 results in NF-kappaB activation.<br />Design: BACs were preincubated with 2',7'-dichlorofluorescin diacetate (DCFH-DA), a dye that allows the monitoring of intracellular ROS production for DCF by spectrofluorometry. BACs were incubated with native LDL and ox-LDL (10, 50, and 100 microg/ml) for 5 min at 37 degrees C and DCF formation was observed. BACs were also preincubated with anti-LOX-1 mAb (40 microg/ml) or ascorbic acid (10 microM). Nuclear extracts from BACs treated for the indicated periods with 50 microg/ml ox-LDL, and preincubated with anti-LOX-1 mAb or ascorbic acid, were prepared and analyzed by electrophoretic mobility shift assay (EMSA).<br />Results: ox-LDL induced a significant dose-dependent increase in ROS production after 5-min incubation with BACs (P < 0.001). ROS formation was markedly reduced in BACs preincubated with anti-LOX-1 mAb and ascorbic acid (P < 0.001). Activation in BACs of the transcription factor NF-kappaB was evident after 5-min incubation with ox-LDL and was attenuated by anti-LOX-1 mAb and ascorbic acid.<br />Conclusion: ox-LDL binding to LOX-1 in BACs increased the production of intracellular ROS and activated NF-kappaB. Reduction of NF-kappaB activation by ascorbic acid indicates that the activation, at least in part, is ROS-dependent. These observations support the hypothesis that hypercholesterolemia is one of several risk factors for arthritis, and that lipid peroxidation products such as ox-LDL are involved in cartilage matrix degradation.<br /> (Copyright 2004 OsteoArthritis Research Society International)
- Subjects :
- Animals
Ascorbic Acid metabolism
Cattle
Cells, Cultured
Dose-Response Relationship, Drug
Fluoresceins analysis
Microscopy, Confocal methods
Oxidation-Reduction
Reactive Oxygen Species antagonists & inhibitors
Receptors, LDL analysis
Receptors, Oxidized LDL
Reverse Transcriptase Polymerase Chain Reaction methods
Cartilage, Articular metabolism
Chondrocytes metabolism
Lipoproteins, LDL metabolism
NF-kappa B metabolism
Reactive Oxygen Species metabolism
Receptors, LDL metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1063-4584
- Volume :
- 12
- Issue :
- 7
- Database :
- MEDLINE
- Journal :
- Osteoarthritis and cartilage
- Publication Type :
- Academic Journal
- Accession number :
- 15219572
- Full Text :
- https://doi.org/10.1016/j.joca.2004.04.005