Bacterial proteases in gingival crevicular fluid before and after periodontal treatment

Our recent work has developed specific assays for bacterial dipeptidyl peptidase (DPP) and trypsin-like proteases and we have found them in gingival crevicular fluid (GCF). The purpose of this study was to determine whether their levels reduce following periodontal treatment of chronic periodontitis patients. The probing depth, probing attachment level, gingival index, gingival bleeding index and plaque index were measured at mesio-buccal sites of molars and premolars in 25 untreated patients. At a second visit GCF was collected on filter paper strips for 30 seconds. GCF volumes were determined with a Periotron and the samples eluted into buffer. The patients than received oral hygiene instruction, supra- and subgingival scaling and other appropriate non-surgical treatment. Four weeks later GCF was collected from the same 16 sites and the clinical parameters were measured again. DPP-like activity was determined fluorometrically with Ala-Pro-AFC at pH 8.0 with and without heating to 60 degrees C for 30 minutes. The heat sensitive portion was taken as bacterial DPP activity. Bacterial trypsin-like protease activity was assayed with Z-Val-Lys-Lys-Arg-AFC at pH 7.0 with 2 mM dithiothreitol and 10 microM Z-Phe-Ala-CH2. Following treatment there were marked reductions in clinical parameters, enzyme total activities and concentrations. All reductions were statistically significant at patient and site level using either individual patient or pooled patient data. Bacterial proteases appear to reflect the clinical status and may be of value in monitoring chronic inflammatory periodontal disease.