title: Impaired degradation of matrix collagen in human gingival fibroblasts by the antiepileptic drug phenytoin.
abstract: BACKGROUND
  Gingival overgrowth (GO) is a serious adverse effect associated with the administration of phenytoin (PHT), with PHT-induced GO characterized by a massive accumulation of extracellular matrix components, especially collagen, in gingival connective tissues. However, the etiology of such collagen accumulation is still largely unknown. We examined the effects of PHT on the collagen degradation process leading to collagen accumulation in human gingival fibroblasts (HGF).
  
  
  METHODS
  HGFs were cultured with various concentrations of PHT and viable cell numbers and collagen amounts were determined. Gene and protein expressions of matrix metalloproteinases (MMP) and tissue inhibitors of MMPs (TIMP) were quantified with reverse transcription-polymerase chain reaction (RT-PCR) analyses and Western blotting, respectively. Cellular endocytosis of collagen was assayed using flow-cytometric analysis. The effects of PHT on extracellular signal-regulated kinase 1/2 (ERK1/2) and inhibitor kappaB-alpha (IkappaB-alpha) were assayed.
  
  
  RESULTS
  The proliferation of HGFs was not affected by PHT, whereas it significantly increased collagen accumulation. Further, the expressions of MMP-1, -2, and -3 were markedly suppressed by PHT, whereas that of TIMP-1 was induced in a dose- and time-dependent manner. PHT also markedly prevented collagen endocytosis by HGFs, which was associated with the suppression of alpha2beta1-integrin expression. In addition, the phosphorylation of ERK1/2 and IkappaB-alpha degradation were suppressed by PHT.
  
  
  CONCLUSIONS
  These results suggest that PHT causes an impaired degradation of collagen by suppression of enzymatic degradation with MMPs/TIMP-1 and alpha2beta1-integrin-mediated endocytosis. Those alterations are likely mediated through the cellular signaling pathways of ERK1/2 and nuclear factor kappaB. These synergistic effects may cause collagen accumulation, leading to GO.
authors.affiliation: Department of Oral Frontier Biology, Osaka University Graduate School of Dentistry, Suita-Osaka, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Kato
authors.forename: Nobuo
authors.initials: N
authors.lastname: Okahashi
authors.forename: Shinji
authors.initials: S
authors.lastname: Kawai
authors.forename: Takafumi
authors.initials: T
authors.lastname: Kato
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Inaba
authors.forename: Ichijiro
authors.initials: I
authors.lastname: Morisaki
authors.forename: Atsuo
authors.initials: A
authors.lastname: Amano
mesh_terms: D000927:Anticonvulsants / Q000494:pharmacology*; D049109:Cell Proliferation / Q000187:drug effects; D003094:Collagen / Q000187:drug effects / Q000378:metabolism; D004705:Endocytosis / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects*; D005881:Gingiva / Q000166:cytology / Q000187:drug effects*; D019214:Gingival Overgrowth / Q000139:chemically induced; D006801:Humans; D038982:Integrin alpha2beta1 / Q000187:drug effects; D020782:Matrix Metalloproteinases / Q000187:drug effects / Q000378:metabolism; D010672:Phenytoin / Q000009:adverse effects / Q000494:pharmacology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019714:Tissue Inhibitor of Metalloproteinases / Q000187:drug effects / Q000378:metabolism
chemical_list: D000927:Anticonvulsants; D038982:Integrin alpha2beta1; D019714:Tissue Inhibitor of Metalloproteinases; D010672:Phenytoin; D003094:Collagen; D020782:Matrix Metalloproteinases