Evaluation of Antimicrobial Effect of Air-Polishing Treatments and Their Influence on Human Dental Pulp Stem Cells Seeded on Titanium Disks

Abstract

Dental implants are one of the most frequently used treatment options for tooth replacement, and titanium is the metal of choice due to its demonstrated superiority in resisting corrosion, lack of allergic reactions and mechanical strength. Surface roughness of titanium implants favors the osseointegration process; nevertheless, its topography may provide a suitable substrate for bacterial biofilm deposition, causing peri-implantitis and leading to implant failure. Subgingival prophylaxis treatments with cleansing powders aimed to remove the bacterial accumulation are under investigation. Two different air-polishing powders-glycine and tagatose-were assayed for their cleaning and antimicrobial potential against a Pseudomonas biofilm and for their effects on human dental pulp stem cells (hDPSCs), seeded on sandblasted titanium disks. Immunofluorescence analyses were carried out to evaluate cell adhesion, proliferation, stemness and osteogenic differentiation. The results demonstrate that both the powders have a great in vitro cleaning potential in the early period and do not show any negative effects during hDPSCs osteogenic differentiation process, suggesting their suitability for enhancing the biocompatibility of titanium implants. Our data suggest that the evaluated cleansing systems reduce microbial contamination and allow us to propose tagatose as an adequate alternative to the gold standard glycine for the air-polishing prophylaxis treatment.

Keywords: air-polishing powders; anti-microbial effect; glycine; hDPSCs; stem cell properties; tagatose; titanium implants.

Figure 1

Immunophenotypic characterization of hDPSCs. Immunophenotypic characterization of c-Kit⁺/STRO-1⁺ hDPSCs by FACS analysis against mesenchymal surface antigens.

Figure 2

Titanium surface topography. Scanning electron microscopy (SEM) analysis at different magnifications was carried out on titanium surfaces from the three experimental groups (CTRL, GLY, TAGAT) in order to evaluate the surface topography. Scale bars: 100 µm (left) and 20 µm (magnifications on the right).

Figure 3

Stem cell morphology, adhesion and proliferation. (A) hDPSCs morphology was evaluated by immunofluorescence analysis of phalloidin (PHA)-stained cells at different time points (24 h, 48 h, 72 h, 7 d) on titanium surfaces previously cleansed with GLY and TAGAT. Control group (CTRL) consisting of hDPSCs seeded on untreated titanium surfaces. Nuclei were counterstained with DAPI. Scale bar = 50 µm. (B) Histograms represent cell proliferation (24 h, 48 h, 72 h, 7 d) of hDPSCs seeded on disks from different experimental groups and one-way ANOVA followed by Newman-Keuls post hoc test. Experiments were performed in triplicate.

Figure 4

Assessment of stemness markers on immunoselected hDPSCs. Evaluation by immunofluorescence analysis of stemness markers STRO-1 (green) and c-Kit (red) after 48 h of culture on hDPSCs seeded on titanium disks previously cleansed with GLY and TAGAT. Control group (CTRL) consisting of hDPSCs seeded on untreated titanium surfaces. Nuclei were counterstained with DAPI. Scale bar = 10 µm.

Figure 5

Angiogenic phenotype of hDPSCs after different air-polishing treatments. Analysis of VEGF expression by immunofluorescence analysis on hDPSCs seeded on titanium disks previously cleansed with GLY and TAGAT. Control group (CTRL) consisting of hDPSCs seeded on untreated titanium surfaces. Red squares show the morphology of hDPSCs after staining with phalloidin (PHA). Nuclei were counterstained with DAPI. Pseudocolor analysis of VEGF is shown on the right side. Scale bar = 10 µm.

Figure 6

Osteogenic commitment of hDPSCs on titanium disks. Confocal immunofluorescence analysis of osteocalcin (OCN) and RUNX-2 were carried out on hDPSCs seeded on titanium disks previously cleansed with GLY and TAGAT. Control group (CTRL) consisting of hDPSCs seeded on untreated titanium surfaces. Nuclei were counterstained with DAPI. Scale bar = 10 µm.