title: Antisense inhibition of 85-kDa cPLA2 blocks arachidonic acid release from airway epithelial cells.
abstract: Inflammatory cytokines play a critical role in the initiation and perpetuation of inflammation. Several cytokines are known to increase the production of arachidonic acid (AA) metabolites, which may mediate cytokine-induced acute and chronic inflammation. Although cytokines upregulate phospholipase A2 (PLA2) in several target cells, the contribution of individual PLA2 to cytokine-induced AA release and eicosanoid production remains unclear because of the existence of various forms of cellular PLA2. To examine the role of 85-kDa cytosolic PLA2 (cPLA2) in cytokine-induced AA release, a system was developed to inhibit the expression of cPLA2 in a human bronchial epithelial cell line (BEAS-2B cells) by antisense RNA. Cells stably expressing antisense cPLA2 exhibited decreased cPLA2 protein levels as well as decreased cPLA2 activity assayed in vitro. The effects of cytokines interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), and interleukin-1 alpha (IL-1 alpha) on the release of prelabeled [3H]AA were then tested in cells stably transfected with vector alone as well as cells transfected with cPLA2 antisense plasmid. IFN-gamma (300 U/ml), TNF-alpha (20 ng/ml), and IL-1 alpha (20 ng/ml) all induced a significantly increased release of prelabeled [3H]AA after 15 min to 2 h of treatment in control cells, and their effects were significantly reduced in cells transfected with cPLA2 antisense vector. These results demonstrate a critical role of cPLA2 in inflammatory cytokine-induced AA metabolism.
authors.affiliation: Critical Care Medicine Department, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: T
authors.initials: T
authors.lastname: Wu
authors.forename: S J
authors.initials: SJ
authors.lastname: Levine
authors.forename: M
authors.initials: M
authors.lastname: Cowan
authors.forename: C
authors.initials: C
authors.lastname: Logun
authors.forename: C W
authors.initials: CW
authors.lastname: Angus
authors.forename: J H
authors.initials: JH
authors.lastname: Shelhamer
mesh_terms: D016375:Antisense Elements (Genetics) / Q000378:metabolism / Q000494:pharmacology*; D016718:Arachidonic Acid / Q000378:metabolism*; D001980:Bronchi / Q000166:cytology / Q000378:metabolism*; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D009693:Nucleic Acid Hybridization; D010741:Phospholipases A / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D054467:Phospholipases A2; D012333:RNA, Messenger / Q000378:metabolism; D012260:Ribonucleases; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D016375:Antisense Elements (Genetics); D004791:Enzyme Inhibitors; D007375:Interleukin-1; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D016718:Arachidonic Acid; D007371:Interferon-gamma; D012260:Ribonucleases; D010741:Phospholipases A; D054467:Phospholipases A2