Osteoclast differentiation during orthodontic tooth movement
A scientific essay in Medical Sciences
DOCTORAL THESIS defended in public on 24th of April 2012
Chapter 1 elucidates the background of the biological process leading to orthodontic tooth movement. The principle is that force application to a tooth eventually leads to bone resorption at the compression area and bone formation at the tension area. An unfavorable side-effect is that root resorption also might occur during orthodontic treatment. This is less predictable than bone resorption although the same cells appear to be responsible for both processes. However, if bone resorption and root resorption are conducted by the same cells, then it is unclear why root resorption is not always found during orthodontic tooth movement, and why there is a difference in the timing of the onset of bone resorption and root resorption after the start of orthodontic force application.
Chapter 2 is a literature review describing the putative principal or gradual differences between the mechanisms behind alveolar bone resorption and root resorption during orthodontic tooth movement. The following questions were addressed:
- What are the structural and chemical similarities and differences between the extracellular matrices of alveolar bone and root cementum?
- What are the morphological and physiological similarities and differences between osteoclasts, odontoclasts, and other cells that may be involved in alveolar bone and root resorption during orthodontic tooth movement?
- What are the similarities and differences between the regulatory mechanisms for the differentiation, recruitment, and functioning of cells involved in the processes?
In chapter 3, the inflammatory response of two often used rat models for experimental tooth movement were compared. In the elastic band model an orthodontic elastic band is squeezed between two molars (group 1); and in the coil spring model a force is exerted by a NiTi coil spring (group 3). The inflammatory reactions evoked by these models, were compared with a model in which a silk ligature is applied around the cervix of the upper second molar to induce periodontitis (group 2), and an untreated control group (group 4). After an experimental period of 1, 3, or 5 days, the continuity of the epithelium and the transseptal fibers of the interdental papilla was graded. Furthermore, the intensity of inflammation in the interdental papilla, vertical bone loss, and the number of osteoclasts and their precursors was evaluated. In group 1 and 2, the epithelium and the transseptal fibers of the interdental papilla were completely absent after 5 days, whereas the epithelium and transseptal fibers in group 3 were still comparable to baseline after 5 days. The inflammatory infiltration was abundant within the interdental papilla in group 1 and 2, and in group 3 the inflammatory infiltration was less severe and restricted to the area occlusal to the CEJ. The vertical bone loss and the interdental bony septum resorption was faster and was more severe in group 1 and 2 than in group 3. From this study, we conclude that elastic bands yield similar periodontitis-like effects as silk ligatures. Therefore, the spring model is to be preferred for experimental tooth movement studies.
Chapter 4 describes the spatial and sequential distribution of osteoclast precursors between one and five days of orthodontic tooth movement using an elastic band model. The reason for this study is that the origin of osteoclasts responsible for bone resorption during orthodontic tooth movement is not yet clear. Their precursors may reside within the periodontal ligament (PDL) or could be recruited from the circulation or the adjacent bone marrow. Three differentiation markers were used: receptor for macrophage colony-stimulating factor (c-Fms), a marker for early precursors, receptor activator of nuclear factor-κB (RANK), a marker for late precursors, and calcitonin receptor (CTR), a marker for mature osteoclasts. Before force application, many early precursors and a few late precursors were present in the adjacent bone marrow while no early precursors were observed in the PDL. After force application, the number of both early and late precursors, as well as multinuclear osteoclasts increased rapidly in the adjacent bone marrow whereas only late precursors but not early precursors increased in the PDL. The number of multinuclear osteoclasts in the PDL increased later than in the adjacent bone marrow. The results of this study suggested strongly that osteoclast precursors differentiate within the adjacent bone marrow and then migrate into the PDL during early tooth movement.
Chapter 5 describes a further study on the differentiation and recruitment of osteoclasts between 6 and 120 hours of experimental tooth movement in rats. Here, ED1 is used as a marker for osteoclasts and their potential precursors (macrophage-lineage cells). Matrix metalloproteinase 9 (MMP9) was used as a marker for migrating osteoclasts and their precursors. The upper three molars of Wistar rats at one side were moved mesially, using NiTi coil springs of 10 cN. The contralateral sides served as controls. After force application, the number of osteoclasts first increased in the bone marrow. At compression areas, the number of potential osteoclast precursors first decreased, followed by an increase of the aforementioned cells and migrating osteoclasts. At tension areas, the pre-existing osteoclasts disappeared rapidly while the number of potential osteoclast precursors remained stable. It confirmed the conclusion of chapter 4 with a different animal model that force application induces osteoclast differentiation first within the bone marrow. These osteoclasts migrate subsequently into the compressed PDL.
Chapter 6 describes a study on eNOS and iNOS expression in osteocytes during orthodontic force application. Osteocytes are mechanosensitive and essential for bone remodeling. During mechanotransduction, they translate canalicular flow into cellular signals which can recruit osteoclasts and osteoblasts. Nitric oxide, an important regulator of bone remodeling, is produced by osteocytes through the activity of constitutively expressed endothelial nitric oxide synthase (eNOS) or inducible nitric oxide synthase (iNOS). We hypothesized that these enzymes regulate the tissue response to orthodontic force. The upper rat molars were moved mesially by NiTi coil springs (10 cN, 6-120 hrs) in a split-mouth design. Immunohistochemical staining revealed that, in the tension area, eNOS-positive osteocytes increased from 24 hrs on, while iNOS-positive osteocytes remained largely constant. In the compression area, iNOS-positive osteocytes increased after 6 hrs, while eNOS-positive osteocytes increased after 24 hrs. This suggests that eNOS mediates bone formation in the tension area, while iNOS mediates inflammation-induced bone resorption in the compression area. Both eNOS and iNOS seem to be important regulators of bone remodeling during orthodontic force application.
Chapter 7 is the general discussion. The background of the topic selection of this thesis was elucidated. The results from the different studies were related. Suggestions for future orthodontic research were proposed.