Analysis of in situ protease activity in chronic adult periodontitis patients: expression of activated MMP-2 and a 40 kDa serine protease.
BACKGROUND
Periodontitis is characterized by extensive destruction of the gingival tissues and associated supporting structures of the teeth. Although the pathogenesis of the various forms of this disease is not completely understood, host-derived proteases are believed to have an important role. In this study, we analyzed human tissue samples from chronic adult periodontitis patients to assess the levels of specific proteases and determine the effect of pH and tetracyclines on their activity.


METHODS
Gingival tissue samples were obtained from patients with chronic adult periodontitis (probing depths ranged from 5 to 9 mm) and periodontally healthy controls. Tissue extracts were prepared and analyzed for protease activity by zymography and Western blotting.


RESULTS
Maximal protease activity from clinically normal and diseased tissues was observed at pH 8. Latent matrix metalloproteinase (MMP)-9 and MMP-2 were expressed in all samples examined, while active MMP-2 was detected only in tissues obtained from patients with clinical disease. The MMP activities were differentially inhibited by derivatives of tetracycline. At pH 6, a protease with a mass of approximately 40 kDa was observed in diseased samples. The enzymatic activity was inhibited by phenylmethylsulfonyl fluoride, suggesting it is a serine protease.


CONCLUSIONS
The results of the current study substantiate the proposed role of host-derived proteases in the pathogenesis of chronic adult periodontitis. Specifically, they indicate that activated MMP-2 and a 40 kDa serine protease are involved in tissue destruction associated with this form of periodontal disease and also suggest that tissue pH influences protease activity in situ.
Korostoff
J M
JM
Department of Microbiology, School of Dental Medicine, University of Pennsylvania, Philadelphia 19104, USA. jkorosto@pobox.upenn.edu
Wang
J F
JF
Sarment
D P
DP
Stewart
J C
JC
Feldman
R S
RS
Billings
P C
PC
D000328:Adult; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015153:Blotting, Western; D002908:Chronic Disease; D015331:Cohort Studies; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic; D005881:Gingiva / Q000201:enzymology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D020778:Matrix Metalloproteinase 2 / Q000032:analysis* / Q000187:drug effects; D020784:Matrix Metalloproteinase 8 / Q000032:analysis; D061965:Matrix Metalloproteinase Inhibitors; D010514:Periodontal Pocket / Q000188:drug therapy / Q000201:enzymology; D010518:Periodontitis / Q000188:drug therapy / Q000201:enzymology*; D010664:Phenylmethylsulfonyl Fluoride / Q000494:pharmacology; D012697:Serine Endopeptidases / Q000032:analysis*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D013754:Tetracyclines
D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D061965:Matrix Metalloproteinase Inhibitors; D015842:Serine Proteinase Inhibitors; D013754:Tetracyclines; D010664:Phenylmethylsulfonyl Fluoride; D012697:Serine Endopeptidases; D020778:Matrix Metalloproteinase 2; D020784:Matrix Metalloproteinase 8