Inhibitory Effect of Periodontally Diseased Root Extracts on the Growth of Human Gingival Fibroblasts

Abstract

Cementum shavings obtained from periodontally diseased and nondiseased areas of 100 removed, single-rooted teeth were extracted with either pyrogen-free water (PFW) for 5 minutes, 1 m citric acid for 5 minutes or 45% phenol-PFW for 90 minutes at 65°C. The extracts were membrane-filtered, dialyzed exhaustively versus PFW, lyophilized, weighed and then dissolved in complete growth medium. The phenol-water or citric acid extracts of cementum shavings from periodontally diseased roots were positive for endotoxin by the limulus lysate assay (LLA). Pyrogen-free water extracts of diseased or phenol-water extracts of nondiseased cementum shavings were negative, or only slightly positive, respectively, for endotoxin by LLA. Media containing the various extracts were added to logarithmically growing cultures of human gingival fibroblasts (HGF). Separate cultures of HGF were exposed to Escherichia coli endotoxin at concentrations of 50, 100, 250 and 500 μg/ml to determine the growth-inhibitory effects of a known endotoxin. Cell growth was analyzed by measuring the incorporation of tritiated thymidine into cells. Suppression of HGF growth from 30 to 49% by E. coli endotoxin was concentration-dependent and linear over the concentration range of endotoxin tested. Pyrogen-free water extracts of diseased (endotoxin negative) or phenol-water extracts of nondiseased cementum shavings (slightly endotoxin positive) did not effect HGF growth. However, citric acid or phenol-water extracts of diseased cementum shavings (highly endotoxin positive) significantly suppressed HGF growth 58% and 61%, respectively. These results indicate that citric acid is effective in removing cytotoxic substances, presumably endotoxin, from cementum shavings and suggest that citric acid treatment is effective clinically in detoxifying periodontally diseased root surfaces.