Eliane H. Dutra

The buccinator during mastication: A functional and anatomical evaluation in minipigs

OBJECTIVE The buccinator muscle forms the lateral wall of the oral cavity. It is presumed to aid mastication by maintaining bolus position. Such a function would involve thickening the cheek, possibly compressing the alveolar bone and contributing to malocclusions. However, neither buccinator deformation nor its effect on pressure has been demonstrated. Our objective was to evaluate buccinator EMG during feeding, its changes in length and thickness, and the pressure exerted on its alveolar attachment, using miniature pigs as an animal model. METHODS EMG of the buccinator and other oral muscles was recorded with fine-wire electrodes. Anteroposterior length and mediolateral thickness of the buccinator were evaluated with implanted sonomicrometry crystals, and pressure was measured by flat transducers placed beneath the mandibular origin of the buccinator. Recordings were made during feeding and muscle stimulation. Tissues were collected postmortem for histology. RESULTS During mastication, buccinator EMG showed regular peaks that preceded those of the jaw closers. Pattern differences clearly distinguished working and balancing sides. The buccinator shortened and thickened when it contracted. Positive pressures were observed at the mandibular attachment of the buccinator, increasing when the muscle was active. Histological evaluation showed a complex interweaving of fibres closely associated with salivary tissue. CONCLUSIONS Buccinator contraction does thicken the cheek, and during mastication this activity takes place just as the closing stroke begins. In addition to controlling the bolus, there may be an effect on salivation. Despite the fact that the muscle pulls on its attachment, the local mechanical environment at the alveolar bone is one of positive pressure.

Cellular and Matrix Response of the Mandibular Condylar Cartilage to Botulinum Toxin

Objectives To evaluate the cellular and matrix effects of botulinum toxin type A (Botox) on mandibular condylar cartilage (MCC) and subchondral bone. Materials and Methods Botox (0.3 unit) was injected into the right masseter of 5-week-old transgenic mice (Col10a1-RFPcherry) at day 1. Left side masseter was used as intra-animal control. The following bone labels were intraperitoneally injected: calcein at day 7, alizarin red at day 14 and calcein at day 21. In addition, EdU was injected 48 and 24 hours before sacrifice. Mice were sacrificed 30 days after Botox injection. Experimental and control side mandibles were dissected and examined by x-ray imaging and micro-CT. Subsequently, MCC along with the subchondral bone was sectioned and stained with tartrate resistant acid phosphatase (TRAP), EdU, TUNEL, alkaline phosphatase, toluidine blue and safranin O. In addition, we performed immunohistochemistry for pSMAD and VEGF. Results Bone volume fraction, tissue density and trabecular thickness were significantly decreased on the right side of the subchondral bone and mineralized cartilage (Botox was injected) when compared to the left side. There was no significant difference in the mandibular length and condylar head length; however, the condylar width was significantly decreased after Botox injection. Our histology showed decreased numbers of Col10a1 expressing cells, decreased cell proliferation and increased cell apoptosis in the subchondral bone and mandibular condylar cartilage, decreased TRAP activity and mineralization of Botox injected side cartilage and subchondral bone. Furthermore, we observed reduced proteoglycan and glycosaminoglycan distribution and decreased expression of pSMAD 1/5/8 and VEGF in the MCC of the Botox injected side in comparison to control side. Conclusion Injection of Botox in masseter muscle leads to decreased mineralization and matrix deposition, reduced chondrocyte proliferation and differentiation and increased cell apoptosis in the MCC and subchondral bone.

The Effect of Altered Loading on Mandibular Condylar Cartilage.

Objective The purpose of this study was to delineate the cellular, mechanical and morphometric effects of altered loading on the mandibular condylar cartilage (MCC) and subchondral bone. We hypothesized that altered loading will induce differentiation of cells by accelerating the lineage progression of the MCC. Materials and Methods Four-week-old male Dkk3 XCol2A1XCol10A1 mice were randomly divided into two groups: (1) Loaded-Altered loading of MCC was induced by forced mouth opening using a custom-made spring; (2) Control-served as an unloaded group. Mice were euthanized and flow cytometery based cell analysis, micro-CT, gene expression analysis, histology and morphometric measurements were done to assess the response. Results Our flow cytometery data showed that altered loading resulted in a significant increase in a number of Col2a1-positive (blue) and Col10a1-positive (red) expressing cells. The gene expression analysis showed significant increase in expression of BMP2, Col10a1 and Sox 9 in the altered loading group. There was a significant increase in the bone volume fraction and trabecular thickness, but a decrease in the trabecular spacing of the subchondral bone with the altered loading. Morphometric measurements revealed increased mandibular length, increased condylar length and increased cartilage width with altered loading. Our histology showed increased mineralization/calcification of the MCC with 5 days of loading. An unexpected observation was an increase in expression of tartrate resistant acid phosphatase activity in the fibrocartilaginous region with loading. Conclusion Altered loading leads to mineralization of fibrocartilage and drives the lineage towards differentiation/maturation.

Dental Anomalies Associated with Craniometaphyseal Dysplasia

Craniometaphyseal dysplasia (CMD) is a rare genetic disorder encompassing hyperostosis of craniofacial bones and metaphyseal widening of tubular bones. Dental abnormalities are features of CMD that have been little discussed in the literature. We performed dentofacial examination of patients with CMD and evaluated consequences of orthodontic movement in a mouse model carrying a CMD knock-in (KI) mutation (Phe377del) in the Ank gene. All patients have a history of delayed eruption of permanent teeth. Analysis of data obtained by cone-beam computed tomography showed significant bucco-lingual expansion of jawbones, more pronounced in mandibles than in maxillae. There was no measurable increase in bone density compared with that in unaffected individuals. Orthodontic cephalometric analysis showed that patients with CMD tend to have a short anterior cranial base, short upper facial height, and short maxillary length. Microcomputed tomography (micro-CT) analysis in homozygous AnkKI/KI mice, a model for CMD, showed that molars can be moved by orthodontic force without ankylosis, however, at a slower rate compared with those in wild-type Ank+/+ mice (p < .05). Histological analysis of molars in AnkKI/KI mice revealed decreased numbers of TRAP+ osteoclasts on the bone surface of pressure sides. Based on these findings, recommendations for the dental treatment of patients with CMD are provided.

Dental Abnormalities in a Mouse Model for Craniometaphyseal Dysplasia

Mice carrying a knock-in mutation (Phe377del) in the Ank gene replicate many skeletal characteristics of human craniometaphyseal dysplasia, including hyperostotic mandibles. AnkKI/KI mice have normal morphology of erupted molars and incisors but excessive cementum deposition with increased numbers of Ibsp- and Dmp1-positive cells on root surfaces. The cervical loops of adult AnkKI/KI lower incisors are at the level of the third molars, while they are close to the mandibular foramen in Ank+/+ mice. Furthermore, AnkKI/KI incisors show decreased eruption rates, decreased proliferation of odontoblast precursors, and increased cell apoptosis in the stellate reticulum. However, their capability for continuous elongation is not compromised. Quantification of TRAP-positive cells in the apical ends of AnkKI/KI incisors revealed decreased osteoclast numbers and osteoclast surfaces. Bisphosphonate injections in Ank+/+ mice replicate the AnkKI/KI incisor phenotype. These results and a comparison with the dental phenotype of Ank loss-of-function mouse models suggest that increased cementum thickness may be caused by decreased extracellular PPi levels and that the incisor phenotype is likely due to hyperostosis of mandibles, which distinguishes AnkKI/KI mice from the other Ank mouse models.

A Morphometric and Cellular Analysis Method for the Murine Mandibular Condyle

The temporomandibular joint (TMJ) has the capacity to adapt to external stimuli, and loading changes can affect the position of condyles, as well as the structural and cellular components of the mandibular condylar cartilage (MCC). This manuscript describes methods for analyzing these changes and a method for altering the loading of the TMJ in mice (i.e., compressive static TMJ loading). The structural evaluation illustrated here is a simple morphometric approach that uses the Digimizer software and is performed in radiographs of small bones. In addition, the analysis of cellular changes leading to alterations in collagen expression, bone remodeling, cell division, and proteoglycan distribution in the MCC is described. The quantification of these changes in histological sections - by counting the positive fluorescent pixels using image software and measuring the distance mapping and stained area with Digimizer - is also demonstrated. The methods shown here are not limited to the murine TMJ, but could be used on additional bones of small experimental animals and in other regions of endochondral ossification.

The effects of alveolar decortications on orthodontic tooth movement and bone remodelling in rats

Objectives Alveolar decortication (AD) is a minimally invasive procedure that can be performed in the orthodontic office as an intervention to accelerate tooth movement. There is a gap in the literature evaluating the earlier and delayed responses after AD using lighter orthodontic forces in a rat model. Therefore, the aim of this study was to determine the effects of AD in the amount of orthodontic tooth movement and on alveolar bone remodelling in a rodent model, after 7 or 14 days. Materials and methods A total of 32 15-week-old male Wistar rats were used in four treatment groups: (1) orthodontic spring only (7 days), (2) orthodontic spring only + AD (7 days), (3) orthodontic spring only (14 days), and (4) orthodontic spring only + AD (14 days). A closed coil nickel-titanium spring delivering 8-10 g of force was used to move the molar mesially. Alveolar decortication was done using a high speed, quarter round bur adjacent to the left first maxillary molar, on the palatal alveolar bone. At each endpoint, rats were sacrificed and microfocus computed tomography and histological analysis were performed. Results The spring + AD group presented with a significant increase in the rate of tooth movement when compared with spring only group, 7 and 14 days after the beginning of the experiments. In addition, the spring + AD group had a significant decrease in bone volume and tissue density and a significant increase in the trabecular spacing and the number of osteoclasts at 7 and 14 days. Furthermore, a fibrous tissue was found to replace the alveolar bone in the spring + AD group at day 14. Conclusion Alveolar decortications enhanced bone remodelling around the tooth movement region and could be used as an adjunct surgical procedure to accelerate the rate of tooth movement.

Bone Response of Loaded Periodontal Ligament

The tooth-periodontal ligament-alveolar bone complex acts symbiotically to dissipate the mechanical loads incurred during mastication and/or orthodontic tooth movement. The periodontal ligament functions both in the tension and compression. At the molecular and celleular levels, the loads in the periodontal ligament trigger mechanobiological events in the alveolar bone, which leads to bone modeling and remodeling. The current review focuses on the bone response to mechanical loading of the periodontal ligament on the tension and pressure sides. Understanding the bone response has major implications for dentistry, including a better understanding of the different types of orthodontic tooth movement.

Temporomandibular Joint Disorders in Older Adults: Temporomandibular Disorders in Older Adults

To review the literature and summarize the evidence of temporomandibualar joint (TMJ) disorders (TMDs) in older adults, focusing on clinical manifestations of TMDs in older adults, highlighting the incidence and sexual dimorphism of TMJ degeneration and the role of sex hormones in this process, and providing potential treatment options for TMD in older adults.Objective To review the literature and summarize the evidence of Temporomandibualar Joint Disorders (TMD) in elderly and older individuals. The goal of this review is to focus on clinical manifestations of TMD in the elderly by highlighting the increased incidence of Temporomandibular Joint (TMJ) degeneration in the elderly, the sexual dimorphism and the role of sex hormones in this process. The review concludes with potential treatment options of TMD in elderly.

PTH [1–34] induced differentiation and mineralization of mandibular condylar cartilage

Intermittent Parathyroid Hormone (I-PTH) is the only FDA approved anabolic drug therapy available for the treatment of osteoporosis in males and postmenopausal females. The effects of I-PTH on the chondrogenic lineage of the mandibular condylar cartilage (MCC) are not well understood. To investigate the role of I-PTH on the MCC and subchondral bone, we carried out our studies using 4 to 5 week old triple transgenic mice (Col1a1XCol2a1XCol10a1). The experimental group was injected with PTH (80 μg/kg) daily for 2 weeks, while control group was injected with saline. Our histology showed that the I-PTH treatment led to an increased number of cells expressing Col1a1, Col2a1 and Col10a1. Additionally, there was an increase in cellular proliferation, increased proteoglycan distribution, increased cartilage thickness, increased TRAP activity, and mineralization. Immunohistochemical staining showed increased expression of pSMAD158 and VEGF in the MCC and subchondral bone. Furthermore our microCT data showed that I-PTH treatment led to an increased bone volume fraction, tissue density and trabecular thickness, with a decrease in trabecular spacing. Morphometric measurements showed increased mandibular length and condyle head length following I-PTH treatment. In conclusion, our study suggests that I-PTH plays a critical role in cellular proliferation, proteoglycan distribution, and mineralization of the MCC.