title: TNF-α stimulates MMP-3 production via PGE2 signalling through the NF-kB and p38 MAPK pathway in a murine cementoblast cell line.
abstract: BACKGROUND
  Cementoblasts are considered to play an important role in the homeostasis of periodontal tissues under both physiologic and pathologic conditions. Matrix metalloproteinases (MMPs) is the key family of enzymes participating in extracellular matrix remodelling. In the present study, the effects and regulatory mechanisms of tumour necrosis factor (TNF)-α on the expression of MMPs and their inhibitors (tissue inhibitor of metalloproteinases; TIMPs) were investigated.
  
  
  METHODS
  OCCM-30, an immortalised murine cementoblast cell line, was stimulated with TNF-α at 1 and 10ng/ml for 24h. The expression of Mmp-2, Mmp-3, Mmp-13, Mmp-14, Timp-1, and Timp-2 as well as PGE2 was determined. Inhibitors of MAPKs, PI3K/Akt, NF-kB and Cox-2 were employed to reveal possible TNF-α induced regulatory signalling pathway(s). The mRNA and protein expression were analysed by (semi)quantitative real-time PCR and enzyme-linked immunosorbent assay (ELISA), respectively.
  
  
  RESULTS
  TNF-α dose-dependently stimulated MMP-3 expression by cementoblasts. This was found for mRNA as well as protein expression. No significant differences were found in the mRNA expression of Mmp-2, Mmp-13, Mmp-14, Timp-1, and Timp-2 upon TNF-α stimulation. The level of PGE2, however, was significantly increased along with MMP-3. Treatment with a selective Cox-2 inhibitor resulted in partial suppression of TNF-α-induced Mmp-3 mRNA expression. Addition of PGE2 enhanced Mmp-3 mRNA in a dose dependent manner, suggesting an inductive effect of TNF-α partly via PGE2. The up-regulation of Mmp-3 by TNF-α was completely suppressed by a combination of NF-kB and p38 MAPK inhibitors, while partial suppression was found with each inhibitor. The effect of PGE2 on Mmp-3 expression was abolished by treating cells with an NF-kB inhibitor; a p38 MAPK inhibitor had only a small effect.
  
  
  CONCLUSIONS
  The present study indicates that cementoblasts respond to TNF-α by increasing MMP-3 production partially via PGE2 and signalling through the NF-kB and p38 MAPK pathway. MMP-3 may participate in periodontal tissue degradation/remodelling.
authors.affiliation: Research Unit of Mineralized Tissue, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Department of Anatomy, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand. Electronic address: sneeracha@yahoo.com.
authors.forename: Neeracha
authors.initials: N
authors.lastname: Sanchavanakit
authors.affiliation: Research Unit of Mineralized Tissue, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand. Electronic address: saengtong_weerayut@hotmail.com.
authors.forename: Weerayut
authors.initials: W
authors.lastname: Saengtong
authors.affiliation: Research Unit of Mineralized Tissue, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Department of Anatomy, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand. Electronic address: aorjm711@gmail.com.
authors.forename: Jeeranan
authors.initials: J
authors.lastname: Manokawinchoke
authors.affiliation: Research Unit of Mineralized Tissue, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Department of Anatomy, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand. Electronic address: prasit215@gmail.com.
authors.forename: Prasit
authors.initials: P
authors.lastname: Pavasant
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D003739:Dental Cementum / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D015232:Dinoprostone / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000378:metabolism*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
chemical_list: D016328:NF-kappa B; C560822:TIMP1 protein, human; C560823:TIMP2 protein, human; D019715:Tissue Inhibitor of Metalloproteinase-1; D014409:Tumor Necrosis Factor-alpha; D019716:Tissue Inhibitor of Metalloproteinase-2; D048051:p38 Mitogen-Activated Protein Kinases; C511768:MMP3 protein, human; D019278:Matrix Metalloproteinase 3; D020778:Matrix Metalloproteinase 2; D015232:Dinoprostone
keywords: Cementoblasts; Cox-2; MMP-3; PGE(2); TNF-α