Evidence for Calcifying Nanoparticles in Gingival Crevicular Fluid and Dental Calculus in Periodontitis

Abstract

Calcifying nanoparticles (CNPs), also known as nanobacteria, can produce carbonate apatite on their cell walls and initiate pathologic calcification. The objective of this study was to determine whether CNPs are present in the gingival crevicular fluid (GCF) from subjects with periodontal disease and whether they can induce the pathologic calcification of primary cultured human gingival epithelial cells. GCF and dental calculus samples were collected from 10 subjects with gingivitis and 10 subjects with chronic periodontitis. CNPs in GCF and calculus filtrates were detected with nanocapture enzyme-linked immunosorbent assay kits. The CNPs in cultures of dental calculus filtrates were also identified using immunofluorescence staining, transmission electron microscopy (TEM), and chemical analysis. Pathologic changes in the CNP-treated gingival epithelial cells were observed with TEM, alizarin red staining, and disk-scanning confocal microscopy. CNPs were found in GCF samples from two subjects with chronic periodontitis. Based on chemical analysis, the surface-associated material from CNPs isolated and cultured from calculus has a composition similar to dental calculus. The pathologic calcification of CNP-treated gingival epithelial cells was also observed. Self-replicating calcifying nanoparticles can be cultured and identified from dental calculus. This raises the issue of whether CNPs contribute to the pathogenesis of periodontitis.