Extraction and quantification of heparin in "clinically normal" and inflamed gingiva of the dog

Heparin present in tissue has been reported to be a potential locally active agent responsible for bone resorption by the stimulation of collagenase via osteoclast activation and increased collagenase synthesis by the osteoblast. The purpose of this study was to extract, identify and quantify heparin in the "clinically normal", mildly and severely inflamed gingiva of Beagle dogs, using a simple and reliable technique. The extraction was carried out by homogenization, fat elimination, proteolytic digestion and precipitation, with organic solvents. Identification was performed by microelectrophoresis conducted on an agarose coated microscope slide. Quantification was performed by measuring the optical density of the metachromatic toluidine blue stained spot an comparing with the standard reference curve of heparin run simultaneously. The results showed that the difference in concentration of heparin in units per gram of wet tissue, was not significant when the "clinically normal" (26 +/- 1.9) was compared with mildly inflamed (24.4 +/- 4.7) gingiva. However, the concentration of heparin in severe gingivitis (79 +/- 7) was significantly higher. Gingival heparin could play important role in the established periodontal lesions, acting as a local factor or co-factor in periodontal bone destruction.