Identification of Ghrelin in Human Saliva: Production by the Salivary Glands and Potential Role in Proliferation of Oral Keratinocytes

Abstract

Abstract Background: We investigated whether ghrelin is present in human saliva, is produced by salivary glands, and physiologic consequences of these findings. Methods: Expression of ghrelin and specific receptor mRNA was determined by PCR. Proteins were identified by immunoblotting and size-exclusion fast protein liquid chromatography (FPLC) with consecutive RIA. Specific RIAs were used for quantification of salivary total and bioactive ghrelin. Distribution of ghrelin was investigated by immunohistochemistry in cryosections of the salivary glands. The effect of ghrelin on incorporation of 5-bromo-2′-deoxyuridine as a measure of cell proliferation was investigated in primary oral keratinocytes. Results:Ghrelin is produced by the salivary glands. The hormone was identified in saliva and glands by immunoblotting and by FPLC fractionation of saliva. Immunohistochemistry demonstrated ghrelin distribution in the salivary glands. The receptor was also produced by the glands and by oral keratinocytes and was shown to be functional. Comparison of total ghrelin values for healthy individuals (body mass index, 18–27 kg/m2) showed significantly lower concentrations in saliva than in serum (P <0.01). The correlation between both matrices was r2 = 0.56 (P <0.001) with a negative correlation to body mass index (r2 = 0.314; P <0.01). Bioactive acylated ghrelin was also present in saliva. Incubation of keratinocytes with ghrelin led to significantly increased cell proliferation (P <0.001). This effect could be completely suppressed by co-incubation with NOX-B11 (50 nmol/L), a novel specific inhibitor of acylated ghrelin. Conclusions: Ghrelin in saliva is produced and released by salivary glands. The effect of ghrelin on oral cell proliferation adds to the pro-proliferative action of other salivary growth factors.