In situ localization and characterization of active proteases in chronically inflamed and healthy human gingival tissues

Background: Studies have indicated an important role for host-derived prote ases in the pathogenesis of periodontal disease. The objectives of this stu dy were: 1) to develop an assay measuring protease activity in situ and 2) to localize and characterize the enzymatic activity in intact inflamed and healthy gingiva. Methods: Gingival specimens were prepared and overlaid with a quenched fluo rescent substrate. Protease activity was visualized by fluorescence microsc opy and correlated with histologic features. Results: In inflamed tissues, enzymatic activity was detected mainly in the connective tissue (predominantly matrix metalloproteases) and, to some ext ent, in the epithelium (predominantly serine proteases). In contrast, clini cally healthy tissues failed to exhibit significant amounts of protease act ivity. Quantitative and qualitative characteristics of protease activity in intact tissues were found to be pH dependent. Conclusions: The method described here enabled assessment of active proteas es in intact tissues where cell-cell and cell-matrix interactions had been maintained. Our results indicate that there are substantial differences in the distribution of specific proteases between clinically healthy and infla med periodontal tissues.