Temporomandibular joint synovial fibroblasts mediate serine proteinase dependent Type I collagen degradation.
Degenerative temporomandibular joint (TMJ) disorders are characterized by the excessive turnover of collagen. In addition to the matrix metalloproteinase (MMP) and cathepsin mediated collagen degradation pathways, a serine proteinase dependent pathway has recently been identified in TMJ fibroblasts. This study focused on further characterizing this serine proteinase pathway utilizing a media-mediated collagen degradation assay and zymography. The conditioned media from cell-mediated collagen degradation assays were incubated with Type I collagen at pH 7.5 with or without a MMP inhibitor (1,10-phenanthroline), serine proteinase inhibitors (alpha1-antitrypsin and soybean trypsin inhibitor, STI), or cysteine proteinase inhibitors. The data showed that 1,10-phenanthroline and STI reduced the collagen cleavage by 12.33% and 47.78%, respectively. The cysteine proteinase inhibitors had no effect. The combination of alpha1-antitrypsin and 1,10-phenanthroline inhibited the cleavage by 79.22%, while STI and 1,10-phenanthroline together blocked the cleavage by 85.44%. Zymography identified a proteinase at approximate 22.5 kDa, which was more effectively blocked by serine proteinase inhibitors than by MMP or cysteine proteinase inhibitors. Reverse transcript-PCR and real-time PCR results demonstrated that TMJ cells did not express trypsinogen-2 mRNA, a collagen cleaving serine proteinase. This study demonstrated that TMJ fibroblasts can predominantly utilize a serine proteinase to mediate collagen degradation, which is not trypsinogen-2.
Song
F
F
Indiana University School of Dentistry, Department of Oral Biology, 1121 West Michigan Street, Room 271, Indianapolis, IN 46202, USA.
Bergdoll
A S
AS
Windsor
L J
LJ
D045744:Cell Line, Tumor; D024042:Collagen Type I / Q000378:metabolism*; D017077:Culture Media, Conditioned; D005347:Fibroblasts / Q000378:metabolism*; D005780:Gelatin / Q000378:metabolism; D006801:Humans; D010618:Phenanthrolines / Q000494:pharmacology; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D012697:Serine Endopeptidases / Q000378:metabolism*; D013583:Synovial Membrane / Q000166:cytology / Q000378:metabolism; D013704:Temporomandibular Joint / Q000378:metabolism*; D014357:Trypsin / Q000032:analysis; D014362:Trypsinogen / Q000032:analysis; D000515:alpha 1-Antitrypsin / Q000494:pharmacology
D024042:Collagen Type I; D017077:Culture Media, Conditioned; D010618:Phenanthrolines; D000515:alpha 1-Antitrypsin; C064831:PRSS2 protein, human; D010664:Phenylmethylsulfonyl Fluoride; D005780:Gelatin; D014362:Trypsinogen; D012697:Serine Endopeptidases; D014357:Trypsin; C025205:1,10-phenanthroline