The role of osteocytes during experimental orthodontic tooth movement: A review.

OBJECTIVE: To explore the types of orthodontic force-induced mechanical stimuli that regulate osteocyte function. DESIGN: In orthodontics, a tooth can be moved through the alveolar bone when an appropriate orthodontic force is applied. These mechanical loads stimulate cells within the bone tissue around the tooth. These cellular responses lead to bone resorption on the side of the tooth where the pressure has been applied and bone deposition on the side of the tooth experiencing tension. Recently, osteocytes were identified to function as mechano-sensory cells in bone tissue that direct bone resorption and bone formation. Based on recent literature, the proposed function of osteocytes during orthodontic tooth movement is explored with better understanding. RESULTS: Several stimuli regulating osteocyte function have been highlighted, and their potential roles in events initiating osteocyte sensing of orthodontic force have been explored in detail. The most popular hypotheses for osteocyte response include stress-induced bone matrix deformation/microcrack formation and fluid-flow shear stress. CONCLUSIONS: Understanding osteocyte function under mechanical stress may have profound implications in future orthodontic treatments.

The Relationship between Number of Natural Teeth and Chewing Qat Habit among Adult Yemeni People in Sana'a: A Pilot Study.

AIMS AND OBJECTIVES: Several studies have reported associations between the habit of chewing qat and the deterioration of periodontal health among adults in Yemen; however, qat's effect on the number of teeth has not yet been evaluated. The purpose of this observational study was to examine the relationship between the number of natural teeth and qat-chewing status among Yemeni participants. MATERIALS AND METHODS: The participants were interviewed with a structured questionnaire by a researcher on the day of the dental check-up. The surveyed items were age, sex, chewing-qat status (nonchewer; current chewer, nonsmoker; current chewer and smoker; current chewer, ex-smoker; ex-chewer, never smoked; ex-chewer and smoker). The dental examination was conducted with dental mirrors by a single examiner under sufficient artificial light. A linear regression was performed, stratified by sex, with the number of natural teeth as the dependent variable and with age and qat status as independent variables using SAS/STAT software. RESULTS: Results show that age was a significant factor for both males and females having few natural teeth. Females had a higher tooth loss than males. The habit of chewing qat did not affect the number of natural teeth for both males and females. In males, a small, but not significant, relationship between chewing-qat status and the number of natural teeth was found. All smokers were qat chewers. CONCLUSIONS: This study highlights for the first time the relationship between having fewer teeth and the habit of chewing qat and provides useful findings for dental health workers to advise people about the benefits of cessation of both qat chewing and smoking.

Prevalence of prematurely lost primary teeth in 5-10-year-old children in Thamar city, Yemen: A cross-sectional study.

OBJECTIVES: The premature loss of primary teeth is a potential risk factor for poor arch length development. Adequate arch length is important to the progression of the permanent teeth. Poor arch length can lead to crowding, ectopic eruption, or impaction of these teeth. This study is designed to assess the prevalence of premature loss of primary teeth in the 5-10-year-old age group. MATERIALS AND METHODS: The study group included 185 children, that is, 91 boys and 94 girls. The dental examination was conducted by an experienced examiner under sufficient artificial light. Data including patient age and missing teeth were collected. Descriptive statistics were applied for data analysis, and from the results, Chi-square tests were used at a level of significance of 5% (P < 0.05). RESULTS: We observed a 40.54% prevalence of premature loss of primary teeth with no statistically significant difference between genders. The lower left primary second molar was the most commonly absent tooth in the dental arch (13.5%). CONCLUSION: The status of premature loss of primary teeth was high in the study group. Implementation of efficient educational and preventive programs to promote oral health would help children maintain a healthy primary dentition and eventually prevent the disturbances in the future development of normal occlusion. Early detection and management of the space problems associated with the early loss of primary teeth would help in reducing malocclusion problems.

Compressive force-produced CCN2 induces osteocyte apoptosis through ERK1/2 pathway.

Osteocytes produce various factors that mediate the onset of bone formation and resorption and play roles in maintaining bone homeostasis and remodeling in response to mechanical stimuli. One such factor, CCN2, is thought to play a significant role in osteocyte responses to mechanical stimuli, but its function in osteocytes is not well understood. Here, we showed that CCN2 induces apoptosis in osteocytes under compressive force loading. Compressive force increased CCN2 gene expression and production, and induced apoptosis in osteocytes. Application of exogenous CCN2 protein induced apoptosis, and a neutralizing CCN2 antibody blocked loading-induced apoptosis. We further examined how CCN2 induces loaded osteocyte apoptosis. In loaded osteocytes, extracellular signal-regulated kinase 1/2 (ERK1/2) was activated, and an ERK1/2 inhibitor blocked loading-induced apoptosis. Furthermore, application of exogenous CCN2 protein caused ERK1/2 activation, and the neutralizing CCN2 antibody inhibited loading-induced ERK1/2 activation. Therefore, this study demonstrated for the first time to our knowledge that enhanced production of CCN2 in osteocytes under compressive force loading induces apoptosis through activation of ERK1/2 pathway.

A method for observing silver-stained osteocytes in situ in 3-microm sections using ultra-high voltage electron microscopy tomography.

Osteocytes are surrounded by hard bone matrix, and it has not been possible previously to directly observe the in situ architecture of osteocyte morphology in bone. Electron microscope tomography, however, is a technique that has the unique potential to provide three-dimensional (3D) visualization of cellular ultrastructure. This approach is based on reconstruction of 3D volumes from a tilt series of electron micrographs of cells, and resolution at the nanometer level has been achieved. We applied electron microscope tomography to thick sections of silver-stained osteocytes in bone using a Hitachi H-3000 ultra-high voltage electron microscope equipped with a 360 degrees tilt specimen holder, at an accelerating voltage of 2 MeV. Osteocytes with numerous processes and branches were clearly seen in the serial tilt series acquired from 3-microm-thick sections. Reconstruction of young osteocytes showed the 3D topographic morphology of the cell body and processes at high resolution. This morphological data on osteocytes should provide useful information to those who study osteocyte physiology and the several models used to explain their mechanosensory properties.

The alteration of a mechanical property of bone cells during the process of changing from osteoblasts to osteocytes.

Osteocytes acquire their stellate shape during the process of changing from osteoblasts in bone. Throughout this process, dynamic cytoskeletal changes occur. In general, changes of the cytoskeleton affect cellular mechanical properties. Mechanical properties of living cells are connected with their biological functions and physiological processes. In this study, we for the first time analyzed elastic modulus, a mechanical property of bone cells. Bone cells in embryonic chick calvariae and in isolated culture were identified using fluorescently labeled phalloidin and OB7.3, a chick osteocyte-specific monoclonal antibody, and then observed by confocal laser scanning microscopy. The elastic modulus of living cells was analyzed with atomic force microscopy. To examine the consequences of focal adhesion formation on the elastic modulus, cells were pretreated with GRGDS and GRGES, and then the elastic modulus of the cells was analyzed. Focal adhesions in the cells were visualized by immunofluorescence of vinculin. From fluorescence images, we could distinguish osteoblasts, osteoid osteocytes and mature osteocytes both in vivo and in vitro. The elastic modulus of peripheral regions of cells in all three populations was significantly higher than in their nuclear regions. The elastic modulus of the peripheral region of osteoblasts was 12053+/-934 Pa, that of osteoid osteocytes was 7971+/-422 Pa and that of mature osteocytes was 4471+/-198 Pa. These results suggest that the level of elastic modulus of bone cells was proportional to the stage of changing from osteoblasts to osteocytes. The focal adhesion area of osteoblasts was significantly higher than that of osteocytes. The focal adhesion area of osteoblasts was decreased after treatment with GRGDS, however, that of osteocytes was not. The elastic modulus of osteoblasts and osteoid osteocytes were decreased after treatment with GRGDS. However, that of mature osteocytes was not changed. There were dynamic changes in the mechanical property of elastic modulus and in focal adhesions of bone cells.

Skeletal Class lll severe openbite treatment using implant anchorage.

A female patient with a skeletal Class III severe anterior openbite was treated using miniplates as the anchorage. The patient was 15 years and 10 months of age when she reported to our university hospital with a chief complaint of anterior openbite and reversed occlusion. The patient had an anterior openbite with an overjet of -3.0 mm and overbite of -5.0 mm and a Class III molar relationship. The cephalometric analysis showed a skeletal Class III relationship (ANB 0 degrees ). After the extraction of the bilateral mandibular third molars, miniplates were placed in the mandibular external oblique line. The mandibular dentition was retracted using elastic chain and miniplates. After treatment, an Angle Class I molar relationship was achieved and overjet and overbite had become 2.0 mm and 1.5 mm. A good facial appearance and occlusal relationship were obtained. The total active orthodontic treatment period was 23 months. Wrap-around type retainers were placed on both jaws and a lingual bonded retainer was also attached in the mandibular incisors. After 1 year of retention, the occlusion was stable, and a good facial profile was also retained. The mandibular deviation to the left was improved and the strain in the circumoral musculature during lip closure disappeared. An appropriate interincisal relationship was achieved by the uprighting of mandibular dentition without changing the vertical intermaxillary relationship. A panoramic radiograph showed no marked root resorption. Our results suggest that implant anchorage is useful for correction of skeletal Class III severe anterior openbite cases.

Actin and microtubule cytoskeletons of the processes of 3D-cultured MC3T3-E1 cells and osteocytes.

Cell shape is the most critical determinant of cell function and is potentially influenced by the organization of a cell's cytoskeletal components. It has been reported that three-dimensionally cultured osteoblasts have a morphology that closely resembles that of osteocytes, most notably including formation of processes. We have previously shown the critical differences between cytoskeletal components in osteoblasts and osteocytes in two-dimensional culture. We have now extended that investigation to the cytoskeletal components of 3D-cultured osteoblasts and osteocytes using 3D cultures of the osteoblast cell line, MC3T3-E1, and primary osteocytes grown in collagen gel. Three-dimensional fluorescent image reconstructions for actin, fimbrin, alpha-actinin, myosin, tropomyosin, and microtubules were made using IMARIS software. Actin, fimbrin, alpha-actinin, myosin, and tropomyosin all appeared in the processes of both cell types, but fimbrin and myosin showed differences in their distribution patterns between cell types. Microtubules were limited in distribution to the proximal region of osteocyte processes but extended the entire length of MC3T3-E1 cell processes. Microtubules were essential for the integrity and formation of MC3T3-E1 cell processes, but osteocyte processes were dependent on actin. These results showed that there are significant differences between the actin and microtubule cytoskeletons in the processes of 3D-cultured MC3T3-E1 cells and in the processes of 3D-cultured primary osteocytes. These differences in the cytoskeleton of the processes of 3D-cultured osteoblasts and of osteocyte dendrites suggest that osteoblast processes may have a different functional role than the osteocyte dendritic network.

Primary cultures of chick osteocytes retain functional gap junctions between osteocytes and between osteocytes and osteoblasts.

The inaccessibility of osteocytes due to their embedment in the calcified bone matrix in vivo has precluded direct demonstration that osteocytes use gap junctions as a means of intercellular communication. In this article, we report successfully isolating primary cultures of osteocytes from chick calvaria, and, using anti-connexin 43 immunocytochemistry, demonstrate gap junction distribution to be comparable to that found in vivo. Next, we demonstrate the functionality of the gap junctions by (1) dye coupling studies that showed the spread of microinjected Lucifer Yellow from osteoblast to osteocyte and between adjacent osteocytes and (2) analysis of fluorescence replacement after photobleaching (FRAP), in which photobleaching of cells loaded with a membrane-permeable dye resulted in rapid recovery of fluorescence into the photobleached osteocyte, within 5 min postbleaching. This FRAP effect did not occur when cells were treated with a gap junction blocker (18alpha-glycyrrhetinic acid), but replacement of fluorescence into the photobleached cell resumed when it was removed. These studies demonstrate that gap junctions are responsible for intercellular communication between adjacent osteocytes and between osteoblasts and osteocytes. This role is consistent with the ability of osteocytes to respond to and transmit signals over long distances while embedded in a calcified matrix.

Fluid shear stress induces less calcium response in a single primary osteocyte than in a single osteoblast: implication of different focal adhesion formation.

UNLABELLED: The immediate calcium response to fluid shear stress was compared between osteocytes and osteoblasts on glass using real-time calcium imaging. The osteoblasts were responsive to fluid shear stress of up to 2.4 Pa, whereas the osteocytes were not. The difference in flow-induced calcium may be related to differences in focal adhesion formation. INTRODUCTION: To explore the immediate response to mechanical stress in a bone cell population, we examined flow-induced calcium transients. In addition, the involvement of focal adhesion-related calcium transients in response to fluid flow in the cells was studied. MATERIALS AND METHODS: Bone cells were isolated from 16-day-old embryonic chicken calvaria by serial treatment with EDTA and collagenase. Single cells on glass without intercellular connections were subjected to fluid flow, and intracellular calcium concentration was measured using imaging with fluo-3. The identification of cell populations in the same field was performed with a chick osteocyte-specific antibody, OB7.3, and an alkaline phosphatase substrate, ELF-97, for osteoblast identification afterward. Immunofluorescence staining of vinculin was performed to visualize focal adhesions. RESULTS: The percentage of cells responding to fluid shear stress at 1.2 Pa was 5.5% in osteocytes, 32.4% in osteoblasts, and 45.6% in OB7.3/ELF-97-negative cells. Furthermore, osteoblasts and OB7.3/ELF-97-negative cells were more responsive to 2.4 Pa than 1.2 Pa, whereas osteocytes were less responsive. The elevation of calcium transients over baseline did not show any significant differences in the populations. To elucidate the mechanism accounting for the fact that single osteocytes are less sensitive to fluid shear stress of up to 2.4 Pa than osteoblasts, we studied focal adhesion-related calcium transients. First, we compared focal adhesion formation between osteocytes and osteoblasts and found a larger number of focal adhesions in osteoblasts than in osteocytes. Next, when the cells were pretreated with GRGDS (0.5 mM) before flow treatment, a significant reduction of calcium transients in osteoblasts (18%) was observed, whereas calcium transients in osteocytes were not changed by GRGDS. Control peptide GRGES did not reduce the calcium transients in either cell type. Furthermore, we confirmed that osteoblasts in calvaria showed a marked formation of vinculin plaques in the periphery of the cells. However, osteocytes in calvaria showed faint vinculin plaques only at the base of the processes. CONCLUSIONS: On glass, single osteocytes are less sensitive to fluid shear stress up to 2.4 Pa than osteoblasts. The difference in calcium transients might be related to differences in focal adhesion formation. Shear stress of a higher magnitude or direct deformation may be responsible for the mechanical response of osteocytes in bone.