title: Tumor necrosis factor-α induces matrix metalloproteinases-3, -10, and -13 in human periodontal ligament cells.
abstract: BACKGROUND
  Various biologic mediators, including matrix metalloproteinases (MMPs), that are implicated in periodontal tissue breakdown can be induced by cytokines. MMPs are known to degrade periodontal ligament attachment, and bone matrix proteins and tissue inhibitors of metalloproteinase (TIMPs) inhibit the activity of MMPs. The aim of this study is to investigate the effect of tumor necrosis factor (TNF)-α on the expression of MMPs in human periodontal ligament (PDL) cells in vitro and establish which MMPs are expressed specifically in response to that stimulus.
  
  
  METHODS
  Cultured PDL cells were stimulated with TNF-α and analyzed with an MMP antibody array. Real-time polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and western blot with cell lysate and zymography were used to measure messenger RNA (mRNA) and protein levels of MMP-3, -10, and -13. To examine TNF receptor (TNFR) expression, PDL cells were examined by flow cytometry, and expression of MMP-3, -10, and -13 was observed after blocking the TNFR with an antagonist. Results from real-time PCR, ELISA, and western blot were analyzed by paired t test.
  
  
  RESULTS
  The antibody array showed that the protein most strongly upregulated by TNF-α stimulation was MMP-3, followed by MMP-13 and MMP-10. The TNF-α receptor blocker specifically inhibited expression of MMP-3 and -13. In addition, TNF-α increased levels of MMP mRNAs in MMP-3, -13, and -10 (in decreasing order). However, ELISAs showed that MMP-13 was the most upregulated protein, followed by MMP-10 and MMP-3. Western blotting indicated that TNF-α increased MMP-3 and -13 levels but had no significant effect on the level of MMP-10, and zymography showed that TNF-α increased the activities of all forms of MMP-3 and -13, but MMP-10 was not detected. Flow cytometry demonstrated that the majority of PDL cells expressed TNFR1.
  
  
  CONCLUSIONS
  TNF-α (10 ng/mL) upregulates levels of MMP-3, -10, and -13 in human PDL cells. These results suggest that these proteins play an important role in the inflammation of PDLs.
authors.affiliation: Kyung Hee University Dental Hospital at Gangdong, Seoul, Korea.
authors.forename: Su-Jin
authors.initials: SJ
authors.lastname: Ahn
authors.forename: Eun-Mi
authors.initials: EM
authors.lastname: Rhim
authors.forename: Ji-Yoen
authors.initials: JY
authors.lastname: Kim
authors.forename: Kyung-Hee
authors.initials: KH
authors.lastname: Kim
authors.forename: Hyeon-Woo
authors.initials: HW
authors.lastname: Lee
authors.forename: Eun-Cheol
authors.initials: EC
authors.lastname: Kim
authors.forename: Sang Hyuk
authors.initials: SH
authors.lastname: Park
mesh_terms: D000328:Adult; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D008297:Male; D053506:Matrix Metalloproteinase 10 / Q000187:drug effects*; D053509:Matrix Metalloproteinase 13 / Q000187:drug effects*; D019278:Matrix Metalloproteinase 3 / Q000187:drug effects*; D061965:Matrix Metalloproteinase Inhibitors / Q000494:pharmacology; D010513:Periodontal Ligament / Q000166:cytology / Q000187:drug effects*; D060888:Real-Time Polymerase Chain Reaction; D018124:Receptors, Tumor Necrosis Factor / Q000032:analysis / Q000037:antagonists & inhibitors; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000032:analysis; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation
chemical_list: D061965:Matrix Metalloproteinase Inhibitors; D018124:Receptors, Tumor Necrosis Factor; D047888:Receptors, Tumor Necrosis Factor, Type I; D014409:Tumor Necrosis Factor-alpha; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13; C511768:MMP3 protein, human; D019278:Matrix Metalloproteinase 3; C511770:MMP10 protein, human; D053506:Matrix Metalloproteinase 10