title: Native low-density lipoprotein-dependent interleukin-8 production through pertussis toxin-sensitive G-protein coupled receptors and hydrogen peroxide generation contributes to migration of human aortic smooth muscle cells.
abstract: OBJECTIVE
  Stimulation of human aortic smooth muscle cells (hAoSMCs) with native low-density lipoprotein (nLDL) induced the production of interleukin-8 (IL-8) that is involved in the pathogenesis of cardiovascular diseases. However, the process of signal transduction of nLDL was currently uncharacterized. Therefore, the aim of this study was to investigate the signal transduction pathway of nLDL-dependent IL-8 production and the effect of IL-8 on hAoSMCs migration.
  
  
  METHODS
  nLDL was prepared by ultracentrifugation with density-adjusted human serum of normocholesterolemia. In hAoSMCs, IL-8 secreted to medium was measured using ELISA assay, and Western blot analysis was performed to detect p38 MAPK activation as a key regulator of IL-8 production. nLDL-dependent H₂O₂ generation was determined by microscopic analysis using 2',7'-dichlorofluoroscein diacetate (DCF-DA). IL-8-induced migration of hAoSMCs was evaluated by counting the cell numbers moved to lower chamber using Transwell plates.
  
  
  RESULTS
  nLDL-induced IL-8 production was completely blocked by preincubation of hAoSMCs with pertussis toxin (PTX), which inhibited nLDL-dependent p38 MAPK phosphorylation. PTX-sensitive G-protein coupled receptor was responsible for nLDL-dependent H₂O₂ generation that was abrogated with preincubation of the cells with of polyethylene glycol-conjugated catalase (PEG-Cat). Pretreatment of PEG-Cat prevented nLDL-induced p38 MAPK phosphorylation and IL-8 production, which was partly mimicked by treatment with exogenous H₂O₂2. Finally, IL-8 increased hAoSMCs migration that was completely blocked by incubation with IL-8 neutralizing antibody.
  
  
  CONCLUSIONS
  PTX-sensitive G-protein coupled receptor-dependent H₂O₂ generation by nLDL plays a critical role in IL-8 production in hAoSMC, and IL-8 may contribute to atherogenesis through increased migration of hAoSMCs.
authors.affiliation: Department of Biology, Kangwon National University, Chuncheon 200-701, Korea.
authors.forename: Hyun Kyo
authors.initials: HK
authors.lastname: Lim
authors.forename: Sungwoo
authors.initials: S
authors.lastname: Ryoo
mesh_terms: D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D016209:Interleukin-8 / Q000096:biosynthesis*; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D037342:Pertussis Toxin / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D017382:Reactive Oxygen Species / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000502:physiology*; D015398:Signal Transduction; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D016209:Interleukin-8; D008077:Lipoproteins, LDL; D017382:Reactive Oxygen Species; D043562:Receptors, G-Protein-Coupled; D006861:Hydrogen Peroxide; D037342:Pertussis Toxin; D048051:p38 Mitogen-Activated Protein Kinases