Macrophage polarization in aseptic bone resorption around dental implants induced by Ti particles in a murine model

Titanium particles/ions detected in peri-implant tissues have been considered as a potential etiologic factor for crestal bone loss around oral implants. However, the definite impact of titanium wear particles on the health of surrounding structures remains undetermined. The purpose of this study was to investigate the effects of titanium particles-induced foreign body reaction on peri-implant bone level and the related mechanism by using clodronate liposomes to deplete macrophages.

Sprague Dawley rats with custom-made titanium screw implanted in bilateral maxillary first molar area for 4 weeks to obtain osseointegration were randomly divided into four groups. Twenty microgram titanium particles were introduced into the peri-implant tissue to induce aseptic foreign body reaction, and macrophages were depleted by the local injection of 100 μL clodronate liposome immediately and re-injection every 3 days until the sacrifice of the rats (Ti + LipClod group). Titanium-injected rats also treated with phosphate buffer solution (Ti + PBS) or empty liposome (Ti + Lip) as well as rats injected with PBS alone (Control) were included as controls. Eight weeks later, animals were sacrificed and samples containing implants were collected. Half of the samples were analyzed radiologically to measure bone level change, and macrophage markers (CD68, CCR7, CD163) was also characterized by immunofluorescence to evaluate macrophage number, density, and phenotype distribution (CCR7+M1/CD163+M2). The rest of the samples were used to determine the relative mRNA expression levels of TNF-α, IL-1β, IL-6, and RANKL with real-time PCR analysis.

No obvious bacterial contamination was found in all titanium-injected areas, and the implant survival rate was 100% with no implant loss. Compared with Ti + PBS and Ti + Lip group, macrophage density (1.64 ± 0.86%) infiltrated into peri-implant tissue and bone loss (0.17 ± 0.03 mm) around implant decreased significantly in the Ti + LipClod group. Immunofluorescence analysis showed that more macrophage infiltrated into peri-implant tissue in the Ti + PBS and Ti + Lip groups, predominantly with M1 phenotype. In contrast, the macrophage density was lower and M2 phenotype was dominant in the Control group, while macrophages density was significantly reduced and the M1 type macrophages were slightly more than M2 type in the Ti + LipClod group. Accordingly, TNF-α, IL-1β, IL6, and RANKL mRNA expression increased significantly in the Ti + PBS and Ti + Lip groups compared with Control and Ti + LipClod groups.

Titanium particles had a negative effect on peri-implant tissue by activating macrophages which induced an M1 macrophage phenotype promoting local secretion of inflammatory cytokines. It was found that clodronate liposome treatment attenuated the severity of inflammation and bone loss by depletion of macrophages. Therefore, the present study revealed the marked impact of macrophage polarization with respect to peri-implant bone loss caused by titanium particles.