Macrophages play a crucial role in inflammatory-mediated bone loss. Orthodontic tooth movement (OTM) is associated with inflammatory bone remodeling. However, whether and how macrophages contribute to mechanical force-induced OTM remains unknown. In this study, we hypothesized that polarization of M1-like macrophages may contribute to the OTM. Orthodontic nickel-titanium springs were applied to the upper first molars of rats or mice to induce OTM. The distance of OTM gradually increased after mechanical force was applied to the rats for 5 and 10 d. M1-like macrophage polarization and expression of M1 cytokine tumor necrosis factor (TNF)-α also increased after force application. More importantly, monocyte/macrophage depletion in mice by injection of clodronate liposomes decreased the distance of OTM and the number of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts and CD68(+) macrophages, accompanied by reduced expressions of M1 markers TNF-α and inducible nitric oxide synthase (iNOS), whereas systemic transfusion of M1 macrophages in mice increased them. Further experiments showed that injection of recombinant TNF-α increased the distance of OTM and the number of TRAP-positive osteoclasts and CD68(+) macrophages, as well as upregulated the expression of TNF-α and iNOS. Blockage of TNF-α by etanercept injection reduced the distance of OTM and the number of TRAP-positive osteoclasts and CD68(+) macrophages, as well as decreased the levels of TNF-α and iNOS. These data suggest that M1-like macrophage polarization promotes alveolar bone resorption and consequent OTM after mechanical force application.

Mechanical force-induced orthodontic root resorption is a major clinical challenge in orthodontic treatment. Macrophages play an important role in orthodontic root resorption, but the underlying mechanism remains unclear. In this study, we examined the mechanism by which the ratio of M1 to M2 macrophage polarization affects root resorption during orthodontic tooth movement. Root resorption occurred when nickel-titanium coil springs were applied on the upper first molars of rats for 3 to 14 d. Positively stained odontoclasts or osteoclasts with tartrate-resistant acid phosphatase were found in resorption areas. Meanwhile, M1-like macrophages positive for CD68 and inducible nitric oxide synthase (iNOS) persistently accumulated on the compression side of periodontal tissues. In addition, the expressions of the M1 activator interferon-γ and the M1-associated pro-inflammatory cytokine tumor necrosis factor (TNF)-α were upregulated on the compression side of periodontal tissues. When the coil springs were removed at the 14th day after orthodontic force application, root resorption was partially rescued. The number of CD68(+)CD163(+) M2-like macrophages gradually increased on the compression side of periodontal tissues. The levels of M2 activator interleukin (IL)-4 and the M2-associated anti-inflammatory cytokine IL-10 also increased. Systemic injection of the TNF-α inhibitor etanercept or IL-4 attenuated the severity of root resorption and decreased the ratio of M1 to M2 macrophages. These data imply that the balance between M1 and M2 macrophages affects orthodontic root resorption. Root resorption was aggravated by an enhanced M1/M2 ratio but was partially rescued by a reduced M1/M2 ratio.

The immune system plays a pivotal role during bone remodeling process. Orthodontic tooth movement (OTM) induces local inflammation in periodontium, but whether systemic immune response is involved in OTM remains unknown. In this study, we show that tooth movement distance was significantly reduced in T-cell-deficient immunocompromised mice compared with wild-type (WT) mice. Intravenous infusion of allogeneic T cells to the immunocompromised mice rescued the OTM distance. Correspondingly, increased numbers of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts were detected around the alveolar bone after OTM in WT mice but were barely detected in immunocompromised mice. Moreover, intravenous infusion of T cells rescued the number of TRAP-positive osteoclasts in the OTM area of the immunocompromised mice, thus suggesting T cells are required for OTM. We then reveal that OTM induced a significant elevation of type 1 T helper cell (Th1) cytokines tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) around periodontal tissue in WT but not in immunocompromised mice. Infusion of T cells could increase the levels of TNF-α and IFN-γ in periodontal tissues of immunocompromised mice. More interestingly, intraperitoneal injection of TNF-α inhibitor etanercept significantly reduced the distance of OTM in T-cell-infused immunocompromised mice. In summary, this study demonstrates a previously unrecognized mechanism that T cells are required for OTM depending on Th1-associated cytokines.

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