Exfoliated oral mucosa cells as bioindicators of short- and long-term systemic titanium contamination

Background: Humans are exposed to exogenous sources of titanium-containing particles that can enter the body mainly by inhalation, ingestion, or dermal absorption. Given the widespread use of biomaterials in medicine, the surface of a titanium (Ti) biomedical device is a potential endogenous source of Ti ions and/or Ti-containing particles, such as TiO2 micro-(MPs) and nano-particles (NPs), resulting from biotribocorrosion processes. Ti ions or Ti-containing particles may deposit in epithelial cells of the oral mucosa, and the latter may therefore serve as bioindicators of short and long-term systemic Ti contamination. The aim of the present study was to histologically and quantitatively evaluate the presence of Ti traces in cells exfoliated from the oral mucosa as possible bioindicators of systemic contamination with this metal at short and long-term experimental time pointsMethods: Thirty Wistar rats were intraperitoneally injected with a suspension of titanium dioxide (TiO2) (0.16 g/ 100 g body weight of TiO2 in 5 ml of NaCl 0.9%) using 5 nm NPs (Group: TiO2-NP5; n = 10), 45 mu m MPs (Group: TiO2-MP45; n = 10), or vehicle alone (Control group; n = 10). At one and six months post-injection, right-cheek mucosa cells were obtained by exfoliative cytology using a cytobrush; they were spray fixed and stained using Safranin or the Papanicolaou technique. The smears were cytologically evaluated (light microscopy) to deter-mine the presence of particulate material, which was also analyzed microchemically (SEM-EDS). Left-cheek mucosa cells were similarly obtained and re-suspended in 5 ml of PBS (pH: 7.2-7.4); the samples correspond-ing to each group were pooled together and analyzed spectrometrically (ICP-MS) to determine Ti concentration in each of the studied groups. Blood samples were obtained for histological determination of the presence of particulate material on Safranin-stained blood smears and determination of plasma concentration of Ti by ICP-MSResults: Different size and shape metal-like particles were observed inside and outside epithelial cells in TiO2-NP5 and TiO2-MP45 cytological smears at both one and six months post-injection. EDS analysis showed the presence of Ti in the particles. ICP-MS revealed higher Ti concentrations in both TiO2 injected groups compared to the control group. In addition, Ti concentration did not vary with time or particle size. Monocytes containing par-ticles were observed in blood smears of TiO2-exposed animals one-and six-months post-injection. Plasma levels of Ti were significantly higher in TiO2-NP5-and TiO2-MP45-exposed animals than in controls (p < 0.05), and Ti concentration was significantly higher at one month than at six months in both TiO2-exposed groups (p < 0.05).Conclusions: Cells exfoliated from the oral mucosa could be used as bioindicators of short-and long-term systemic contamination with Ti. Exfoliative cytology could be used as a simple, non-invasive, and inexpensive diagnostic method for monitoring biotribocorrosion of Ti implants and patient clinical follow-up.