Cathepsin B in gingival crevicular fluid of adult periodontitis patients: identification by immunological and enzymological methods.

Cathepsin B (EC 3.4.22.1), a typical lysosomal cysteine proteinase was identified immunologically with anti-human cathepsin B antibody in inflammatory exudate, gingival crevicular fluid (GCF) of adult periodontitis patients. The sensitive enzyme immunoassay (EIA) system initially developed, was rarely influenced by the presence of endogenous cysteine proteinase inhibitors, cystatin(s), indicating that it is possible to quantify the gross amount of cathepsin B including free enzyme forms and enzyme-inhibitor complex forms using this EIA system. The cathepsin B levels in GCF as determined by EIA and the activity measured with Z-Arg-Arg-MCA showed positive and significant correlation with various clinical parameters. Immunoblotting analysis revealed that the molecular form was a 29 kDa mature enzyme. More than 95% of Z-Arg-Arg-MCA hydrolytic activity in each GCF sample was inhibited by CA-074, specific inhibitor of cathepsin B. These results strongly suggested that the gross amount of cathepsin B in GCF as well as its activity level is closely associated with the severity of the disease and that cathepsins B play an important role in the pathogenesis of periodontitis.