Experimental model of tooth mobility in the human "in vivo"]

The aim of the present study was to investigate experimentally the mechanical properties of tooth deflection under external loading. These properties have a significant impact on tooth movement during orthodontic treatment. The stresses and strains caused by tooth movement influence bone remodelling, which is the basis of orthodontic treatment. The movement of a tooth as a direct reaction to the forces acting on it is termed "initial" movement. It is nonlinear and has a clearly time-dependent component. While the initial tooth movement represents the totality of the reaction mechanisms of all the tissues of the tooth unit, it is determined primarily by the mechanical properties of the periodontal ligament (PDL). The PDL is the softest tissue of the tooth unit and is therefore subject to the largest deformations when forces act on the crown of the tooth. The objective of orthodontic treatment is to achieve as precise and rapid tooth movement as possible, without provoking such undesired effects as bone and root resorption. To enable the implementation of an optimal orthodontic force system that meets these requirements, a thorough knowledge of the biomechanics of tooth movement is a must.     

Expression of genes for gelatinases and tissue inhibitors of metalloproteinases in periodontal tissues during orthodontic tooth movement

Orthodontic tooth movement progresses by a combination of periodontal ligament (PDL) tissue and alveolar bone remodeling processes. Besides the remodeling of alveolar bone around the moving teeth, the major extracellular matrix (ECM) components of PDLs, collagens, are degenerated, degraded, and restructured. Matrix metalloproteinases (MMPs) and their specific inhibitors, tissue inhibitors of metalloproteinases (TIMPs), act in a co-ordinated fashion to regulate the remodeling of periodontal tissues. We hypothesized that the expression levels of the genes for MMP-2, MMP-9, and TIMPs 1–3 are increased transiently in the periodontal tissue during orthodontic tooth movement. To test this hypothesis, we employed an animal model of tooth movement using rats, as well as in situ hybridization to analyze the expression levels of Mmp-2, Mmp-9, and Timps 1-3. The expression levels of these genes increased transiently in cells of periodontal tissues, which include cementoblasts, fibroblasts, osteoblasts, and osteoclasts, at the compression side of the moving teeth. The transient increases in gene expression at the tension side were mainly limited to osteoblasts and cementoblasts. In conclusion, the expression levels of Mmp-2, Mmp-9, and Timps 1-3 increase transiently during orthodontic tooth movement at both the tension and compression sides. The expression of these genes is regulated differentially in the periodontal tissue of the tension side and compression side. This altered pattern of gene expression may determine the rate and extent of remodeling of the collagenous ECM in periodontal tissues during orthodontic tooth movement.     

Detection of immature dendritic cells in the enamel organ of rat incisors by using anti-cystatin C and anti-MHC class II immunocytochemistry.

Dendritic cells in the enamel organ of rat incisors were examined with immunocytochemistry using an anti-cystatin C antibody for immature dendritic cells and macrophages, OX6 for MHC Class II, ED1 for macrophages and dendritic cells, and ED2 for macrophages. Single cells positive for anti-cystatin C appeared in the enamel organ in zones at which ameloblasts secrete enamel matrix proteins. They were also present in transition and enamel maturation zones. In addition, ameloblasts, osteocytes, and osteoclasts were labeled by anti-cystatin C. ED1 and ED2 immunocytochemistry revealed that there was no macrophage population in the enamel organ of secretion, transition, or enamel maturation zone. A double labeling study showed that most anti-cystatin C-positive cells in the enamel maturation zone were also positive for OX6, whereas anti-cystatin C-positive and OX6-negative cells were prevalent in the secretion zone. The results suggest that immature dendritic cells penetrate the enamel organ of the secretion zone and begin to mature in the zones of transition and enamel maturation. (J Histochem Cytochem 48:1243-1255, 2000)     

Cellular response to orthodontically-induced short-term hypoxia in dental pulp cells

Orthodontic force application is well known to induce sterile inflammation, which is initially caused by the compression of blood vessels in tooth-supporting apparatus. The reaction of periodontal ligament cells to mechanical loading has been thoroughly investigated, whereas knowledge on tissue reactions of the dental pulp is rather limited. The aim of the present trial is to analyze the effect of orthodontic treatment on the induction and cellular regulation of intra-pulpal hypoxia. To investigate the effect of orthodontic force on dental pulp cells, which results in circulatory disturbances within the dental pulp, we used a rat model for the immunohistochemical analysis of the accumulation of hypoxia-inducible factor-1α in the initial phase of orthodontic tooth movement. To further examine the regulatory role of circulatory disturbances and hypoxic conditions, we analyze isolated dental pulp cells from human teeth with regard to their specific reaction under hypoxic conditions by means of flow cytometry, immunoblot, ELISA and real-time PCR on markers (Hif-1α, VEGF, Cox-2, IL-6, IL-8, ROS, p65). In vivo experiments showed the induction of hypoxia in dental pulp after orthodontic tooth movement. The induction of oxidative stress in human dental pulp cells showed up-regulation of the pro-inflammatory and angiogenic genes Cox-2, VEGF, IL-6 and IL-8. The present data suggest that orthodontic tooth movement affects dental pulp circulation by hypoxia, which leads to an inflammatory response inside treated teeth. Therefore, pulp tissue may be expected to undergo a remodeling process after tooth movement.     

Simultaneous immunohistochemical demonstration of antigen expression and 5-bromodeoxyuridine incorporation in plastic embedded sections

Summary In this study a double immunohistochemical staining procedure is described for the simultaneous demonstration of antigen expressing cells and replicating cells in rat thymus. As markers for cell surface antigen expression a monoclonal antibody against Ia-expressing cells (His 19) and a monoclonal antibody against cells of the monocyte-macrophage lineage (ED2) were used. Replicating cells were demonstrated by the incorporation of 5-bromodeoxyuridine (BrdUrd). Tissue pieces were fixed in a periodate-lysine-paraformaldehyde fixative and embedded in glycol methacrylate. To demonstrate Ia-expressing cells or ED2-positive macrophages in plastic embedded sections a digestion with trypsin is necessary. The staining procedure was applied sequentially and was performed with a peroxidase and an alkaline phosphatase labeled reagent yielding respectively a brown and a blue reaction product. Results with this staining procedure on plastic embedded sections of rat thymus, an organ with a high DNA synthesizing capacity, showed incorporation of BrdUrd predominantly in the cortex. ED2-positive macrophages were only found in the cortex. The la-positive epithelial reticular cells demonstrated extremely well their stellate form.     

Simultaneous immunohistochemical demonstration of antigen expression and 5-bromodeoxyuridine incorporation in plastic embedded sections

In this study a double immunohistochemical staining procedure is described for the simultaneous demonstration of antigen expressing cells and replicating cells in rat thymus. As markers for cell surface antigen expression a monoclonal antibody against Ia-expressing cells (His 19) and a monoclonal antibody against cells of the monocyte-macrophage lineage (ED2) were used. Replicating cells were demonstrated by the incorporation of 5-bromodeoxyuridine (BrdUrd). Tissue pieces were fixed in a periodate-lysine-paraformaldehyde fixative and embedded in glycol methacrylate. To demonstrate Ia-expressing cells or ED2-positive macrophages in plastic embedded sections a digestion with trypsin is necessary. The staining procedure was applied sequentially and was performed with a peroxidase and an alkaline phosphatase labeled reagent yielding respectively a brown and a blue reaction product. Results with this staining procedure on plastic embedded sections of rat thymus, an organ with a high DNA synthesizing capacity, showed incorporation of BrdUrd predominantly in the cortex. ED2-positive macrophages were only found in the cortex. The Ia-positive epithelial reticular cells demonstrated extremely well their stellate form.     

Histological evidence that metformin reverses the adverse effects of diabetes on orthodontic tooth movement in rats

This study evaluated the effects of metformin on orthodontic tooth movement in a rat model of type 2 diabetes mellitus. Rats were fed a high-fat diet for 4 weeks to induce fat accumulation and insulin resistance, and then injected with a low dose of streptozotocin (35 mg/kg) intraperitoneally to induce type 2 diabetes. An orthodontic appliance was placed in normoglycemic, type 2 diabetes, and type 2 diabetes with metformin-administrated rats. After 14 days, type 2 diabetes rats exhibited greater orthodontic tooth movement and had a higher number of tartrate-resistant acid phosphatase-positive osteoclasts, stronger cathepsin K expression, and weaker alkaline phosphatase immunostaining than normoglycemic rats. Metformin administration resulted in normalization of osteoclast numbers, cathepsin K immunostaining, and of tooth movement as well as partly recovery of alkaline phosphatase expression in diabetic rats. Metformin also reduced sclerostin expression and improved the immunolocalization of dentin matrix protein 1 in osteocytes of type 2 diabetes rats. These results suggest that metformin administration reversed the adverse effects of diabetes on orthodontic tooth movement.     

Simulation of bone remodelling in orthodontic treatment

Orthodontic treatments not only displace irregular teeth but also induce responses in surrounding bone tissues. Bone remodelling is regarded as the regulatory mechanism triggered by mechanical loading. This study was aimed at investigating the effect of orthodontic loading on both tooth movement and neighbouring bone density distribution. A set of computational algorithms incorporating both external and internal remodelling mechanisms was implemented into a patient-specific 3D finite element (FE) model to investigate and analyse orthodontic treatment under four typical modes of orthodontic loading. The consequence of orthodontic treatment was reproduced numerically by using this FE-based technique. The results indicated that the diverse modes of orthodontic loading would result in different magnitudes of tooth movement and particular morphology of bone density distribution. It is illuminated that the newly developed algorithms may replicate the clinical situation more closely compared with the previous proposed method.     

Effect of orthodontic force on the expression of PI3K,Akt, and P70S6 K in the human periodontal ligament during orthodontic loading

The mammalian target of rapamycin (mTOR) is an atypical serine/threonine protein kinases involved in the regulation of cell growth, proliferation, and differentiation through the PI3K/Akt/mTOR/P70S6 K signalling pathway. P70S6 K as a downstream molecule of mTOR is activated by phosphorylation and subsequently promotes the synthesis of ribosomal and translational proteins. In this study, we investigated the role of PI3K, Akt, and P70S6 K in human periodontal tissue remodelling during orthodontic loading. The prepared tissue specimens taken from 4 extracted premolars were processed for immunolabelling. The changes in the expression of PI3K, Akt, and P70S6 K in the periodontal tissues were detected by real‐time quantitative‐polymerase chain reaction and Western blot analysis. The results from real‐time quantitative‐polymerase chain reaction and Western blot both showed that the expression of PI3K, Akt, and P70S6 K in the experimental group began to increase at 3 days and increased significantly at 10 days, then decreased approaching the control group level at 28 days. Our findings showed that the expression of PI3K, Akt, and P70S6 K in human periodontal ligament demonstrated a variability during the orthodontic loading, which suggested that the PI3K/Akt/mTOR/P70S6 K signal pathway was involved in orthodontic tooth movement and played a role in the process of periodontium remodelling.     

Effect of interrupted eruption on the enamel organ of the rat incisor

The aim of this study was to investigate the behavior of rat incisor tissues during the inhibition of tooth eruption. Twenty Sprague-Dawley rats were used in this study, and incisor eruption was inhibited by a screw pin. Animals were sacrificed 1, 3, 7 and 14 days after the start of the experiment. Cross-sections at the mesial point of the mandibular first molar and sagittal sections of the mandibular tooth germ area were examined using immunohistochemical and immunofluorescence methods. For morphometric analysis, numbers of TRAP-positive cells were calculated against the total number of cells. In cross-sections from the experimental group, dentin was thickened and pulp tissue was constricted day by day. On days 1, 3 and 7, nestin-positive cells were observed in all odontoblast cell bodies and processes, while on day 14 fewer nestin-positive cells were seen than in the control group. On day 14, the mesial area of the periodontal ligament was constricted and the number of TRAP-positive cells in the mesial area was significantly higher than in the control group. In sagittal sections, enamel formation was found to be increased on days 7 and 14. Furthermore, in the enamel matrix amelogenin was expressed more strongly than in the control group. PCNA-positive cells were significantly increased in cells of the tooth germ compared with the control group. These results suggest that inhibition of tooth eruption accelerates the apical elongation with resorption of the mesial area of the alveolar bone and stimulates cell proliferation with thickened enamel towards the apical end.     

Acid phosphatase activity is detected preferentially in the osteoclastic lineage by pre-treatment with cyanuric chloride

We previously reported a simple method to detect osteoid matrices in decalcified bone sections by pre-treatment with cyanuric chloride. We have applied this technique to identify osteoclasts and their precursors in rats. In JB-4 sections prepared from untreated bone tissues with cyanuric chloride, both acid phosphatase (ACP) and tartrate-resistant acid phosphatase (TRAP) were found not only in osteoclasts and bone marrow mononuclear cells but also in osteoblasts. In contrast, treatment of bones with cyanuric chloride resulted in staining ACP preferentially in osteoclasts and mononuclear cells adjacent to the bone surface. In the osteoclasts and most of the ACP-positive mononuclear cells, autoradiography showed calcitonin binding. Decalcification with EDTA did not affect the staining for ACP activity in bones treated with cyanuric chloride. It was possible to simultaneously identify ACP and osteoid matrix in a decalcified section. In soft tissues without treatment with cyanuric chloride, both ACP and TRAP were detected in splenic macrophages, alveolar macrophages, and proximal convoluted ducts in kidney. Neither ACP nor TRAP was found in these cell types in the tissues treated with cyanuric chloride. This procedure provides a new, simple method to identify a more restricted population in the osteoclastic lineage than that detected by TRAP staining.     

正畸牙移动相关信号通路研究进展

<正>畸牙移动是在机械力的作用下,通过对牙周膜产生牵张或压缩的力来引起牙周组织在生理限度内的组织改建,从而达到牙齿移动、矫治畸形的目的。由于没有明显的年龄限制,正畸矫治在全球范围已变得越来越普遍。因此,相关的研究也日益增多。牙齿移动的生物学基础是正畸力作用于牙周组织激活一系列信号转导通路,进而引起牙周膜的修复改建。为指导临床、加速正畸矫治疗程提供新的思路,本文综述了近年来有关正畸牙移动相关信号通路的研究进展。发现最新的研究集中在MAPK信号通路,Wnt/β-catenin信号通路,PI3K/AKt/m TOR信号通路,BMP-2信号通路,Caspase-3介导的凋亡通路较多。但是正畸牙移动引起的牙周组织改建是一个多种生物力学信号转导通路相互调节相互作用的过程,对于上述信号通路之间的相互关系还有待于我们更进一步的探索。     

Mechano-transduction in periodontal ligament cells identifies activated states of MAP-kinases p42/44 and p38-stress kinase as a mechanism for MMP-13 expression

Background  

Mechano-transduction in periodontal ligament (PDL) cells is crucial for physiological and orthodontic tooth movement-associated periodontal remodelling. On the mechanistic level, molecules involved in this mechano-transduction process in PDL cells are not yet completely elucidated.     

A visco-elastic model for the prediction of orthodontic tooth movement

This study presents a biomechanical model of orthodontic tooth movement. Although such models have already been presented in the literature, most of them incorporate computationally expensive finite elements (FE) methods to determine the strain distribution in the periodontal ligament (PDL). In contrast, the biomechanical model presented in this work avoids the use of FE methods. The elastic deformation of the PDL is modelled using an analytical approach, which does not require setting up a 3D model of the tooth. The duration of the lag phase is estimated using the calculated hydrostatic stresses, and bone remodelling is predicted by modelling the alveolar bone as a viscous material. To evaluate the model, some typically used motion patterns were simulated and a sensitivity analysis was carried out on the parameters. Results show that despite some shortcomings, the model is able to describe commonly used motion patterns in orthodontic tooth movement, in both single- and multi-rooted teeth.     

Numerical study of tension and strain distribution around rat molars]

A knowledge of the mechanical processes triggered in the bone and periodontal ligament (PDL) by orthodontic forces applied to a tooth is of decisive importance for an understanding of the subsequent remodelling around the tooth. To investigate these mechanical relationships, three-dimensional finite element (FE) models of the first lower molar in the rat were established. On the basis of digitized serial histological sections, these FE models were generated semi-automatically. Using various simplified geometrical variations, an appropriate FE model for the analysis of the stress and strain distributions was established. The numerical analyses were carried out under a mesially directed force of 0.1 N. Stress distributions in the bone and PDL showed a similar pattern, while strains in the bone were lower than in the PDL by a factor of 10-5. The data confirm the assumption that strain patterns in the PDL may be the key stimulus of bone remodelling.     

Tartrate-resistant acid phosphatase in mononuclear and multinuclear cells during the bone resorption of tooth eruption

Tartrate-resistant acid phosphatase (TRAP) has been used as a cytochemical marker for the cell mediators of bone resorption, osteoclasts and their mononuclear precursors. We have applied a cytochemical method for TRAP to study the dependence of the osteoclast-mediated bone resorption of tooth eruption on the dental follicle, a connective tissue investment of the developing tooth, by analyzing the TRAP activity of mononuclear cells in the dental follicle before and during pre-molar eruption in dogs. The percentage of TRAP-positive monocyte cells increases until mid-eruption, slightly preceding a previously demonstrated rise in numbers of osteoclasts on adjacent bone surfaces. These data suggest an ontogenetic relationship between follicular mononuclear cells and osteoclasts on adjacent alveolar bone surfaces during tooth eruption. However, because TRAP occurs in other tissues and is not an exclusive indicator of pre-osteoclasts, proof of their relationship will have to await application of more definitive techniques.     

HSPA1A is upregulated in periodontal ligament at early stage of tooth movement in rats

Heat shock proteins (HSPs) are molecular chaperones that maintain intracellular protein homeostasis and ensure survival of cells. Continuous orthodontic force on the tooth is considered to be a type of physical stress loaded to the periodontal ligament (PDL). However, little is known about the role of HSPs during tooth movement. This study was performed to examine the expression of HSPs in the PDL during tooth movement using laser microdissection, microarray analysis, real-time RT-PCR and immunohistochemistry. Gene expression of HSPA1A in the pressure zone of the PDL was higher during 6 h of tooth movement than in the control group. Expression of HSPA1A decreased with time. HSPA1A was also detected in the pressure zone of the PDL at the protein level 24 h after the initial tissue change. These results strongly suggest that expression of HSPA1A in the PDL during early stages of tooth movement is a critical factor for tissue reaction.     

Differential immunohistochemical resolution of the human mononuclear phagocyte system

Four new monoclonal antibodies, termed Ki-M1, Ki-M2, Ki-M3, and Ki-M4, were developed for distinguishing macrophage subgroups. Purified lysosomes of cells of stimulated U-937 cell line were used as immunogen. Specificity control was performed by staining unfixed cryostat sections of fresh human tissue with an immunohistochemical method, which allowed reliable recognition of reactive structures. Ki-M1 reacted with macrophages of lymphoid tissue, lung, and serous cavities. Ki-M2 recognized Kupffer cells and splenic macrophages. Both monoclonal antibodies reacted with interdigitating reticulum cells and Langerhans cells, which are thought to be accessory cells of the T-cell immune response. Accessory cells of the B-cell immune response, on the other hand, showed reactivity with Ki-M3 and Ki-M4. Thus, analogous to T lymphocytes, the human mononuclear phagocyte system (MPS) can be subdivided into different subgroups with the aid of appropriate monoclonal antibodies.     

An immunohistochemical study examining the role of collagen type VI in the rodent periodontal ligament

Summary The distribution of collagen types I and VI has been examined in the periodontal ligaments of rat incisor and molar teeth using cryosections and immunohistochemical staining procedures. The stain for collagen type I was uniform in all the ligaments examined. Uniform staining for collagen type VI was evident only in the ligament of the fully erupted molars, and the stain was absent from the mid-zone of the ligaments of the erupting molars and incisors. The staining pattern of the collagen type VI antibodies is consistent with previous reports which have suggested that the removal of collagen type VI precedes the degradation of major banded collagen fibrils in remodelling connective tissues. The removal of collagen type VI from the mid-zone of the incisor ligament may precede the remodelling events which facilitate tooth eruption.