title: MicroRNA-199a* regulates the expression of cyclooxygenase-2 in human chondrocytes.
abstract: OBJECTIVE
  Cyclooxygenase-2 (COX-2) expression is associated with the pathogenesis of chronic inflammation and pain in osteoarthritis (OA). A study was undertaken to determine whether interleukin-1β (IL-1β)-mediated induction of COX-2 can be regulated by microRNAs (miRNAs) in OA.
  
  
  METHODS
  Human chondrocytes were stimulated with IL-1β in vitro. Total RNA was prepared using Trizol reagent. Gene expression was quantified using TaqMan Assays and miRNA targets were identified using bioinformatics. Transfection with reporter construct and premiRNA and antimiRNA was employed to verify suppression of target mRNA. Expression of COX-2 proteins was determined by immunoblotting. The role of activated p38-MAPKs was evaluated using specific inhibitor.
  
  
  RESULTS
  The 3'UTR of COX-2 mRNA contained the 'seed-matched' sequences for miR-199a* and miR-101_3. Increased expression of COX-2 correlated with the downregulation of miR-199a* and miR-101_3 in IL-1β-stimulated normal and OA chondrocytes. miR-199a* directly suppressed the luciferase activity of a COX-2 3'UTR reporter construct and inhibited the IL-1β-induced expression of COX-2 protein in OA chondrocytes. Modulation of miR-199a* expression also caused significant inhibition of IL-1β-induced upregulation of mPGES1 and prostaglandin E(2) production in OA chondrocytes. Activation of p38-MAPK downregulated the expression of miR-199a* and induced COX-2 expression. Treatment with antimiR-101_3 increased COX-2 expression in IL-1β-stimulated chondrocytes, but overexpression of miR-101_3 had no significant effect on COX-2 protein expression.
  
  
  CONCLUSIONS
  miR-199a* is a direct regulator of COX-2 expression in OA chondrocytes. IL-1β-induced activation of p38-MAPK correlates inversely with miR199a* expression levels. miR-199a* may be an important regulator of human cartilage homeostasis and a new target for OA therapy.
authors.affiliation: Department of Medicine/Rheumatology, MetroHealth Medical Center/Case Western Reserve University, Cleveland, Ohio 44109, USA.
authors.forename: Nahid
authors.initials: N
authors.lastname: Akhtar
authors.forename: Tariq M
authors.initials: TM
authors.lastname: Haqqi
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D019902:Chondrocytes / Q000187:drug effects / Q000502:physiology*; D051546:Cyclooxygenase 2 / Q000235:genetics* / Q000378:metabolism; D015232:Dinoprostone / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology; D017930:Genes, Reporter; D006801:Humans; D053583:Interleukin-1beta / Q000494:pharmacology; D035683:MicroRNAs / Q000235:genetics*; D010003:Osteoarthritis / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D020413:3' Untranslated Regions; D051546:Cyclooxygenase 2; D015232:Dinoprostone; D053583:Interleukin-1beta; D035683:MicroRNAs; D012333:RNA, Messenger; D048051:p38 Mitogen-Activated Protein Kinases; C534072:MIRN101 microRNA, human; C528163:MIRN26 microRNA, human; C531408:mirn199 microRNA, human