Age-related changes in collagen properties and mineralization in cancellous and cortical bone in the porcine mandibular condyle.

Collagen is an important constituent of bone, and it has been suggested that changes in collagen and mineral properties of bone are interrelated during growth. The aim of this study was to quantify age-related changes in collagen properties and the degree of mineralization of bone (DMB). The DMB in cancellous and cortical bone samples from the mandibular condyle of 35 female pigs aged 0-100 weeks was determined using micro-computed tomography. Subsequently, the amount of collagen and the number of pentosidine (Pen), hydroxylysylpyridinoline (HP), and lysylpyridinoline (LP) cross-links were quantified by means of high-performance liquid chromatography. The amount of collagen increased with age in cancellous bone but remained unchanged in cortical bone. The number of Pen and LP cross-links decreased in both bone types. In contrast, the number of HP cross-links decreased only in cancellous bone. The sum of the number of HP and LP cross-links decreased with age in cancellous bone only. The DMB increased in cancellous and cortical bone. It was concluded that the largest changes in the number of mature collagen cross-links and the mineralization in porcine cancellous and cortical bone take place before the age of 40 weeks. The low number of mature cross-links after this age suggests that the bone turnover rate continues to be high and thereby prevents the development of mature cross-links.

Ae2(a,b)-deficient mice exhibit osteopetrosis of long bones but not of calvaria.

Extracellular acidification by osteoclasts is essential to bone resorption. During proton pumping, intracellular pH (pH(i)) is thought to be kept at a near-neutral level by chloride/bicarbonate exchange. Here we show that the Na(+)-independent chloride/bicarbonate anion exchanger 2 (Ae2) is relevant for this process in the osteoclasts from the long bones of Ae2(a,b)(-/-) mice (deficient in the main isoforms Ae2a, Ae2b(1), and Ae2b(2)). Although the long bones of these mice had normal numbers of multinucleated osteoclasts, these cells lacked a ruffled border and displayed impaired bone resorption activity, resulting in an osteopetrotic phenotype of long bones. Moreover, in vitro osteoclastogenesis assays using long-bone marrow cells from Ae2(a,b)(-/-) mice suggested a role for Ae2 in osteoclast formation, as fusion of preosteoclasts for the generation of active multinucleated osteoclasts was found to be slightly delayed. In contrast to the abnormalities observed in the long bones, the skull of Ae2(a,b)(-/-) mice showed no alterations, indicating that calvaria osteoclasts may display normal resorptive activity. Microfluorimetric analysis of osteoclasts from normal mice showed that, in addition to Ae2 activity, calvaria osteoclasts--but not long-bone osteoclasts--possess a sodium-dependent bicarbonate transporting activity. Possibly, this might compensate for the absence of Ae2 in calvaria osteoclasts of Ae2(a,b)(-/-) mice.

The adaptive response of jaw muscles to varying functional demands.

Jaw muscles are versatile entities that are able to adapt their anatomical characteristics, such as size, cross-sectional area, and fibre properties, to altered functional demands. The dynamic nature of muscle fibres allows them to change their phenotype to optimize the required contractile function while minimizing energy use. Changes in these anatomical parameters are associated with changes in neuromuscular activity as the pattern of muscle activation by the central nervous system plays an important role in the modulation of muscle properties. This review summarizes the adaptive response of jaw muscles to various stimuli or perturbations in the orofacial system and addresses general changes in muscles as they adapt, specific adaptive changes in jaw muscles under various physiologic and pathologic conditions, and their adaptive response to non-surgical and surgical therapeutic interventions. Although the jaw muscles are used concertedly in the masticatory system, their adaptive changes are not always uniform and vary with the nature, intensity, and duration of the stimulus. In general, stretch, increases neuromuscular activity, and resistance training result in hypertrophy, elicits increases in mitochondrial content and cross-sectional area of the fibres, and may change the fibre-type composition of the muscle towards a larger percentage of slow-type fibres. In contrast, changes in the opposite direction occur when neuromuscular activity is reduced, the muscle is immobilized in a shortened position, or paralysed. The broad range of stimuli that affect the properties of jaw muscles might help explain the large variability in the anatomical and physiological characteristics found among individuals, muscles, and muscle portions.

Development of the micro architecture and mineralization of the basilar part of the pig occipital bone.

In this study, the development of the architecture and the degree and distribution of mineralization in the basilar part of the pig occipital bone, one of the contact points between the spine and skull base, was investigated. Multiple regions of the basiocciput of pig specimens of different gestational ages were examined with three-dimensional microcomputed tomography (microCT). The cortex of the basilar part developed from a structure with a large intertrabecular separation into a more compact one, whereas its center maintained a trabecular structure. The cortex displayed a significant increase in bone volume fraction with age, together with an increase in trabecular thickness. In the center no change in bone volume fraction was observed, because of a combined decrease in trabecular number and increase in trabecular separation. During development the degree of mineralization was almost identical in both the center and the cortex and it tended to increase with age. This chance was, however, insignificant. The distribution of mineralization within the trabecular elements of both regions demonstrated that the cores of the elements were more highly mineralized than their surfaces. This difference in mineralization confirms the preexisting notion that trabecular elements grow in size due to surface apposition of new bone that initially is less mineralized.

Relationship between tissue stiffness and degree of mineralization of developing trabecular bone.

It is unknown how the degree of mineralization of bone in individual trabecular elements is related to the corresponding mechanical properties at the bone tissue level. Understanding this relationship is important for the comprehension of the mechanical behavior of bone at both the apparent and tissue level. The purpose of the present study was, therefore, to determine the tissue stiffness and degree of mineralization of the trabecular bone tissue and to establish a relationship between these two variables. A second goal was to assess the change in this relation during development. Mandibular condylar specimens of four fetal and four newborn pigs were used. The tissue stiffness was measured using nanoindentation. A pair of indents was made in the cores of 15 trabecular elements per specimen. Subsequently, the degree of mineralization of these locations was determined from microcomputed tomography. The mean tissue stiffness was 11.2 GPa (+/-0.5 GPa) in the fetal group and 12.0 GPa (+/-0.8 GPa) in the newborn group, which was not significantly different. The degree of mineralization of the fetal trabecular cores was 744 mg/cm3 (+/-28 mg/cm3). The one in the newborn bone measured 719 mg/cm(3) (+/-34 mg/cm3). Again, the difference was statistically insignificant. A significant relationship between tissue stiffness and degree of mineralization was obtained for fetal (R = 0.42, p < 0.001) and newborn (R = 0.72, p < 0.001) groups. It was concluded that woven bone tissue in fetal and newborn trabecular cores resembles adult trabecular bone in terms of tissue properties and is strongly correlated with degree of mineralization.

Jaw-muscle activity changes after the induction of osteoarthrosis in the temporomandibular joint by mechanical loading.

AIMS: To examine the effect of mechanical loading on the induction of temporomandibular joint osteoarthrosis (TMJ OA). METHODS: Mechanical stress was applied to the rat TMJ by forced jaw opening of 3 hours a day for 5 days. The electromyographic (EMG) activity of the masseter and digastric muscles was continuously monitored by radio-telemetry. It was characterized by the total time each muscle was active (duty time), the number of bursts, and the average burst length. For histologic analysis, rats were sacrificed before, immediately after, and 3 weeks after the period of forced jaw opening. RESULTS: The condylar cartilage revealed OA-like lesions with a decrease in the number of chondrocytes immediately after forced jaw opening. Three weeks later, the OA-like lesions were repaired to some extent. After the forced jaw opening, the duty time of the masseter increased, whereas the duty time of the digastric decreased significantly (P < .01) at the 5% activity level. Three weeks later, the masseter duty time had decreased and the digastric duty time had slightly increased, returning to the levels observed before forced jaw opening. CONCLUSION: These results suggest that mechanical overloading of the TMJ induced OA-like lesions with a simultaneous influence on jaw muscle activity, especially at the low activity level. This might imply that muscle activity adapted to reduce the effects of (forced) joint overloading.

Modeling of the effect of friction in the temporomandibular joint on displacement of its disc during prolonged clenching.

PURPOSE: The effect of the frictional coefficient in the temporomandibular joint on the disc during prolonged clenching was examined. MATERIALS AND METHODS: For this purpose, a finite element model of the temporomandibular joint based on magnetic resonance images from a volunteer subject was used. Muscle forces applied for clenching were used as a loading condition for stress analysis during 10 minutes. With respect to the frictional coefficient between articular surfaces, 3 different values ranging from micro = 0.001 to micro = 0.1 were established. RESULTS: At the onset of clenching, large stresses were found in the central and lateral part of the intermediate zone in the disc, and its stress distribution was not markedly changed during 10 minutes of clenching. In the retrodiscal tissue, stress relaxation occurred during the first 2 minutes of clenching. When the frictional coefficient between articular surfaces increased, the anterior, lateral, and central points in the disc moved further forward. At the end of 10 minutes of clenching, the disc showed a more anterior position as the frictional coefficient increased. CONCLUSION: This result indicates that increase of the frictional coefficient between the articular surfaces may be a major cause for the onset of the disc displacement.

Intratrabecular distribution of tissue stiffness and mineralization in developing trabecular bone.

The purpose of this study was to investigate the relation between bone tissue stiffness and degree of mineralization distribution and to examine possible changes during prenatal development. Understanding this may provide insight into adaptation processes and into deformation mechanisms of the bone microstructure. Mandibular condyles from four fetal and newborn pigs were used. Tissue stiffness was measured using nanoindentation, the degree of mineralization with microCT. Eight indents were made over the trabecular width of 15 trabeculae in each specimen, leading to a total of 960 indents. Subsequently, the degree of mineralization of these locations was determined. Intratrabecular variations in bone tissue stiffness and degree of mineralization showed a similar pattern; low at trabecular surfaces and higher in the cores. A strong correlation was found between the two variables, which remained unchanged during development. It was concluded that bone tissue in fetal and newborn trabecular cores resembles adult trabecular bone tissue properties and is distributed in a regular radial pattern in trabeculae. For the first time, it was shown that the intratrabecular tissue stiffness develops along the same path as the degree of mineralization. Knowledge regarding intratrabecular tissue stiffness and mineralization results in a better understanding of trabecular bone mechanical behavior on a structural and tissue level.

Biomechanical consequences of developmental changes in trabecular architecture and mineralization of the pig mandibular condyle.

The purpose of the present study was to examine the changes in apparent mechanical properties of trabecular bone in the mandibular condyle during fetal development and to investigate the contributions of altering architecture, and degree and distribution of mineralization to this change. Three-dimensional, high-resolution micro-computed tomography (microCT) reconstructions were utilized to assess the altering architecture and mineralization during development. From the reconstructions, inhomogeneous finite element models were constructed, in which the tissue moduli were scaled to the local degree of mineralization of bone (DMB). In addition, homogeneous models were devised to study the separate influence of architectural and DMB changes on apparent mechanical properties. It was found that the bone structure became stiffer with age. Both the mechanical and structural anisotropies pointed to a rod-like structure that was predominantly oriented from anteroinferior to posterosuperior. Resistance against shear, also increasing with age, was highest in the sagittal plane. The reorganization of trabecular elements, which occurred without a change in bone volume fraction, contributed to the increase in apparent stiffness. The increase in DMB, however, contributed more dominantly. Incorporating the observed inhomogeneous distribution of mineralization decreased the apparent stiffness, but increased the mechanical anisotropy. This denotes that there might be a directional dependency of the DMB of trabecular elements, i.e. differently orientated trabecular elements might have different DMBs. In conclusion, the changes in DMB and its distribution are important to consider when studying mechanical properties during development and should be considered in other situations where differences in DMB are expected.

Dynamic shear properties of the porcine molar periodontal ligament.

The role of the periodontal ligament (PDL) is to support the tooth during function and resist external forces applied to it. The dominant vertical component of these forces is associated with shear in the PDL. Little information, however, is available on the dynamic behavior of the PDL in shear. Therefore, the present study was designed to determine the dynamic shear properties of the PDL in the porcine molar (n=10). From dissected mandibles transverse sections of the mesial root of the first molar were obtained at the apical and coronal levels and used for dynamic shear tests. Shear strain (0.5%, 1.0%, and 1.5%) was applied in superoinferior direction parallel to the root axis with a wide range of frequencies (0.01-100 Hz). The dynamic complex and storage moduli increased significantly with the loading frequency, the dynamic loss modulus showed only a small increase. The dynamic elasticity was significantly larger in the coronal region than in the apical region although the dynamic viscosity was similar in both regions. The present results suggest that non-linearities, compression/shear coupling, and intrinsic viscoelasticity affect the shear material behavior of the PDL, which might have important implications for load transmission from tooth to bone and vice versa.

Heterogeneous postnatal transitions in myosin heavy chain isoforms within the rabbit temporalis muscle.

Postnatal changes in the fiber type composition and fiber cross-sectional area were investigated in the superficial (TEM1) and deep (TEM23) temporalis of male rabbits. It was hypothesized that, due to the transition from suckling to chewing during early postnatal development, the proportion of fast fiber types would decrease, while the proportion of fibers positive for myosin heavy chain (MyHC) cardiac alpha would increase, and that, due to the influence of testosterone during late postnatal development, the proportion of these alpha fibers would decrease again. Classification of the fibers types was performed by immunohistochemistry according to their MyHC content. The proportion of alpha fiber types significantly increased in both muscle portions from 2% and 8% for TEM1 and TEM23 at week 1 to 29% and 54% at week 8, respectively,. While in TEM1 the proportion of this fiber type did not change thereafter, it decreased again to 27% in TEM23 at week 20. The change for the fast fiber types was opposite to that of the alpha fiber types. Significantly more MyHC IIX fibers were found in TEM1 than in TEM23 in adult rabbits. In the first 8 weeks, the cross-sectional areas of all fibers increased. After this period, only MyHC cardiac alpha + I fibers continued to increase significantly. It was concluded that there are developmental differences in the myosin heavy chain transitions of the two portions of the temporalis muscle.

Regional differences in architecture and mineralization of developing mandibular bone.

The goal of this study was to investigate the mutual relationship between architecture and mineralization during early development of the pig mandible. These factors are considered to define the balance between the requirements for bone growth on the one hand and for load bearing on the other. Architecture and mineralization were examined using micro-CT, whereas the mineral composition was assessed spectrophotometrically in groups of fetal and newborn pigs. The development of the condyle coincided with a reorganization of bone elements without an increase in bone volume fraction, but with an increase in mineralization and a change in mineral composition. In the corpus, the bone volume fraction and mineralization increased simultaneously with a restructuring of the bone elements and a change in mineral composition. The growth of the condyle was reflected by regional differences in architecture and mineralization. The anterior and inferior regions were characterized by a more dense bone structure and a higher mineralization as compared to posterior and superior regions, respectively. In the corpus, growth was mainly indicated by differences between buccal and lingual plates as well as between anterior, middle, and posterior regions characterized by a more compact structure and higher mineralization in the lingual and middle regions. In conclusion, the architecture and mineralization in the condyle and corpus started to deviate early during development toward their destiny as trabecular and cortical bone, respectively. These results were compatible with those obtained with mineral composition analysis. Regional differences within condyle and corpus reflected known developmental growth directions.

Architecture and mineralization of developing cortical and trabecular bone of the mandible.

Ossification of the presumptive trabecular bone in the mandibular condyle and the presumptive cortical bone in the mandibular corpus of the pig mandible was investigated during development, using micro-computed tomography (microCT). Three-dimensional architecture and mineralization characteristics were assessed from ten pigs of different developmental ages. In the condyle, increases in trabecular thickness and separation and a decrease in the trabecular number, led to an unchanged bone volume fraction. A conversion from rod-like into plate-like trabeculae was observed. Bone volume and trabecular thickness were always higher in the corpus, where an increase in bone volume fraction was caused by an increase in the trabecular thickness and a decrease in separation. A transition from a plate-like structure into a more compact structure took place. The average degree of mineralization in the condyle and the corpus increased with age. In the corpus, the degrees of mineralization were higher than in the condyle. The differences between the condyle and corpus and the changes with age could be explained by differences in the distribution of mineralization within the trabecular elements. Generally, the degrees of mineralization increased from the surface toward the centers of the trabecular elements, indicating growth of the trabecular elements by the surface apposition of new mineral.

Circadian variation and intermuscular correlation of rabbit jaw muscle activity.

The activation of jaw muscles varies with different tasks and must be coordinated to ensure proper function of the masticatory system. The activation patterns might differ in various muscles or over the time course. In order to evaluate the activation patterns and the intermuscular correlation during normal daily activity the electromyograms (EMG) of the superficial and deep masseter, medial pterygoid and digastric muscles were continuously recorded in rabbits and related to activity levels. Muscle use was assessed as the relative time per hour (duty time) during which predefined levels of the peak-EMG of the day were exceeded. Pearson's correlation of duty times was calculated for 6 muscle pairs at various activity levels. The duty times of the muscles differed significantly at levels exceeding 50% of the peak-EMG. The animals exhibited apparent intraday variations of duty times revealing a circadian covariant pattern of muscle use. These variations, however, were different in each individual animal. The activation of pairs of jaw-closing muscles was more highly correlated than that of pairs consisting of a jaw-closing and a jaw-opening muscle. The mutual dependence of hourly muscle activity among jaw-closers and among jaw-closers and jaw-openers varied with the activity level suggesting that those muscle groups might be independently controlled during non-powerful and powerful motor behaviors.

Architecture and mineralization of developing trabecular bone in the pig mandibular condyle.

Architecture and mineralization are important determinants of trabecular bone quality. To date, no quantitative information is available on changes in trabecular bone architecture and mineralization of newly formed bone during development. Three-dimensional architecture and mineralization of the trabecular bone in the mandibular condyle from six pigs of different developmental ages were investigated with micro-CT. Anteriorly in the condyle, a more advanced state of remodeling was observed than posteriorly, where more active growth takes place. Posteriorly, the bone volume fraction increased with age (r=0.87; P<0.05) by an increase of trabecular thickness (r=0.88; P<0.05), while the number of trabeculae declined (r=-0.86; P<0.05). Anteriorly, despite an increase in trabecular thickness (r=0.97; P<0.001), there was no change in bone volume fraction due to a simultaneous decline in trabecular number (r=-0.84; P<0.05) and increase in trabecular separation (r=0.95; P<0.01). Posteriorly, rods were remodeled into plates as expressed by the structure model index (r=-0.97; P<0.001), whereas anteriorly, a plate-like structure was already present in early stages. The trabecular structure had a clear orientation throughout the developmental process. The global degree of mineralization increased both anteriorly (r=0.86; P<0.05) and posteriorly (r=0.89; P<0.05). We suggest that the degree of mineralization does not depend on the bone volume, but on the thickness of the trabeculae as the mineralized centers of trabeculae were getting larger and more highly mineralized with age compared to their appositional layers. This indicates that besides apposition of new bone material on the surface of trabeculae, the mineralized tissue in their centers still changes and matures.

The Relationship between molar dentoalveolar and craniofacial heights.

Excessive vertical growth of the posterior dentoalveolar region has been implicated in the etiology of the so-called long-face syndrome. In this study, we tested the hypothesis that molar dentoalveolar heights are positively related to vertical craniofacial features. Cephalometric measurements obtained from 82 adult subjects were entered as independent variables in a multiple regression model. Maxillary and mandibular molar dentoalveolar heights were entered as dependent variables. Approximately 70% of the total variance was explained by anterior lower facial height (ANS-Me) and the mandibular palatal plane angle (PP-MP). Increases of ANS-Me and PP-MP had opposite effects on the amount of molar dentoalveolar heights. The lowest values of molar dentoalveolar heights were found in subjects with a small ANS-Me distance but with a wide PP-MP angle. The findings suggest that individuals with a marked divergence of the jaws may also have a reduced molar dentoalveolar vertical development.

Biomechanical response of condylar cartilage-on-bone to dynamic shear.

Shear stress can result in fatigue, damage, and irreversible deformation of the mandibular condylar cartilage. However, little information is available on its dynamic properties in shear. We tested the hypothesis that the dynamic shear properties of the condylar cartilage depend on the frequency and amplitude of shear strain. Ten porcine mandibular condyles were used for dynamic shear tests. Two cartilage-bone plugs were dissected from each condyle and tested in a simple shear sandwich configuration under a compressive strain of 10%. Sinusoidal shear strain was applied with an amplitude of 1.0, 2.0, and 3.0% and a frequency range between 0.01 and 10 Hz. The magnitudes of the shear dynamic moduli were found to be dependent on the frequency and the shear strain amplitude. They increased with shear strain. tan delta ranged from 0.2 to 0.4, which means that the cartilage is primarily elastic in nature and has a small but not negligible viscosity. In conclusion, the present results show that the shear behavior of the mandibular condylar cartilage is dependent on the frequency and amplitude of the applied shear strain. The observed shear characteristics suggest a significant role of shear strain on the interstitial fluid flow within the cartilage.

The influence of mineralization on intratrabecular stress and strain distribution in developing trabecular bone.

The load-transfer pathway in trabecular bone is largely determined by its architecture. However, the influence of variations in mineralization is not known. The goal of this study was to examine the influence of inhomogeneously distributed degrees of mineralization (DMB) on intratrabecular stresses and strains. Cubic mandibular condylar bone specimens from fetal and newborn pigs were used. Finite element models were constructed, in which the element tissue moduli were scaled to the local DMB. Disregarding the observed distribution of mineralization was associated with an overestimation of average equivalent strain and underestimation of von Mises equivalent stress. From the surface of trabecular elements towards their core the strain decreased irrespective of tissue stiffness distribution. This indicates that the trabecular elements were bent during the compression experiment. Inhomogeneously distributed tissue stiffness resulted in a low stress at the surface that increased towards the core. In contrast, disregarding this tissue stiffness distribution resulted in high stress at the surface which decreased towards the core. It was concluded that the increased DMB, together with concurring alterations in architecture, during development leads to a structure which is able to resist increasing loads without an increase in average deformation, which may lead to damage.

Age-related changes in microarchitecture and mineralization of cancellous bone in the porcine mandibular condyle.

The mandibular condyle is considered a good model for developing cancellous bone because of its rapid growth and high rate of remodeling. The aim of the present study was to analyze the simultaneous changes in microarchitecture and mineralization of cancellous bone during development in a three-dimensional fashion. Eight mandibular condyles of pigs aged 8 weeks prepartum to 108 weeks postpartum were scanned using microCT with an isotropic spatial resolution of 10 microm. The number of trabeculae decreased during development, whereas both the trabecular thickness and the distance between the trabeculae increased. The bone surface to volume ratio decreased during development, possibly limiting the amount of (re)modeling. Both the mean degree of mineralization and intratrabecular differences in mineralization between the surfaces and cores of trabecular elements increased during development. The trabecular surfaces were more highly mineralized in the older condyles compared to the younger ones. Together with the observed decrease in the relative size of trabecular surface, this finding suggests a decrease in (re)modeling activity during development. In accordance with the general growth and development of the pig, it was concluded that most developmental changes in cancellous bone occur until the age of 40 weeks postpartum.

Effect of food consistency on the degree of mineralization in the rat mandible.

A switch to a soft diet, associated with reduced forces applied to the mandible during mastication, may result in an alteration of the degree of mineralization in the mandible. This alteration may be regionally different. The aim of this study was to analyze this alteration by examination of the degree of mineralization in the mandible of growing rats fed with a hard or soft diet. Fifteen Wistar male rats were used in this investigation. After weaning, six rats were fed with a hard diet and the remaining nine rats with a soft diet. After 9 weeks, three-dimensional reconstructions of the cortical and trabecular bone of their mandibles were obtained using a microCT system. The degree of mineralization was determined for the trabecular bone in the condyle and for the cortical bone in the anterior and posterior areas of the mandibular body. In both diet groups the degree of mineralization was significantly (p < 0.01) lower in the trabecular than in the cortical bone. In the mandibular body, the anterior area showed a significantly (p < 0.01) higher degree of mineralization than the posterior area in both diet groups. In both areas the soft diet group had a significantly (p < 0.05 or 0.01) higher degree of mineralization than the hard diet group. The trabecular bone in the condyle of the hard diet group showed a significantly (p < 0.01) higher degree of mineralization than in the soft diet group. The present results indicate the importance of proper masticatory muscle function for craniofacial growth and development.