Epidermal growth factor in gingival crevicular fluid and its binding capacity in inflamed and non-inflamed human gingiva

Abstract

Epidermal growth factor (EGF) is a pro-inflammatory small peptide (6000 Da) with a variety of biological activities including stimulation of cell differentiation and mediation of proteolysis by binding to its specific receptor on the cell surface. The purpose of this study was to determine the levels of EGF in gingival crevicular fluid (GCF) and the EGF-binding capacity to its receptor in gingival tissue. The GCF samples were collected from six patients by inserting paper strips into shallow (< 5 mm) and deep pockets (≥5 mm) for 30 s. The strips were soaked in 0.2 M acetate for extraction and the EGF in the supernatants was analysed by radioimmunoassay. To determine the binding capacity of EGF to its receptor, inflamed gingival tissues (pocket depth ≥ 5 mm, Gingival index = 1, 2 or 3) were collected during periodontal flap surgery and non-inflamed gingival tissues (pocket depth < 5 mm, Gingival Index = 0) were collected during surgical ‘crown lengthening’ for aesthetic purposes. The tissues were pooled by group, homogenized for membrane preparation and the supernatants obtained after centrifugation were used in a 125ÍEGF binding assay. To determine the effect of inflammation on gingival EGF receptor, inflamed and non-inflamed gingival tissues were collected from six patients and prepared similarly to the binding assay. Gingival preparations were then electrophoresed for Western blot anslysis with EGF receptor antiserum. The EGF level in GCF was significantly lower ( P < 0.05) in the samples collected from pockets ≥ 5 mm (0.9 ± 0.6 ng/ml) than in those from pockets < 5 mm (2.4 ± 2.1 ng/ml). The average Gingival Index was higher (2.6 ± 0.6) in pockets ≥ 5 mm than in pockets < 5 mm (1.4 ± 1.0). Specific binding of 125I-EGF to its receptor in inflamed gingiva was 2.7-fold higher than in non-inflamed gingiva (14.4 ± 4.9 vs 5.4 ± 1.8 fmol/g wet tissue). Western blot analysis showed two major immunoreactive bands (180 and 120 kDa), which represent EGF receptor and its degradation products, in inflamed gingiva. The findings show that inflammation activates EGF binding capacity in gingiva and that the up-regulation of EGF receptor in inflamed gingiva might be associated with a lowered concentration of EGF in GCF produced adjacent to inflamed gingiva. This up-regulation of EGF receptor during inflammation might be an important mechanism in the pathogenesis of periodontal disease.