Mandibular growth retardation in corticosteroid-treated juvenile mice

Juvenile mice were treated for up to eight weeks with weekly doses of a synthetic analogue of cortisol:triamcinolone hexacetonide. The mandibular condylar cartilage was studied histologically and histochemically at regular intervals. Morphometric measurements were performed along the mandibular posterior vertical dimension (condylar process and ramus). By the second injection significant morphological changes were noted in the condylar cartilage, followed by retardation of bone growth. The most distinctive feature in the cartilage of triamcinolone-treated mice was a marked increase in the dimension of its mineralized zone concomitant with a significant increase in the number of hypertrophic chondrocytes. The role of condylar cartilage in mandibular growth is discussed.     

Histological and histomorphometric investigation of the condylar cartilage of juvenile pigs after anterior mandibular displacement

The condylar cartilage of the mandible is considered a secondary growth center and represents a joint cartilage different from other cartilage structures regarding its histological structure, its histochemical and immunohistochemical properties and its growth pattern. This study aimed to histologically and histomorphometrically investigate the condylar cartilage after anterior mandibular displacement similar to functional orthopedic treatment. A total of 12 pigs (sus scrofa domesticus) aged 10 weeks were divided into an experimental group and a control group comprising 6 animals each. The experimental animals were provided bilaterally with synthetic occlusal build-ups in the posterior area which induced anterior displacement of the mandible in terminal occlusion. After 4 weeks, the temporomandibular structures were removed en bloc and the condylar cartilage was analyzed histologically and histomorphometrically. As a result, the experimental animals displayed a significantly increased total cartilage thickness of the posterocranial mandibular condyle which was primarily caused by an increase in thickness of the hypertrophic and chondogenic layers. Similarly, the proliferative layer showed a significant increase, whereas significant differences in thickness were absent in the articular layer. Increased cell proliferation was not observed in the experimental animals as compared to the controls. The changes found in the condylar cartilage area suggest that the zonal structure of the condylar cartilage may be modified by an altered spatial relationship between the mandibular condyle and the glenoid fossa.     

Morphological Changes in the Mandible of Male Mice Associated With Aging and Biomechanical Stimulus

Degenerative changes in the temporomandibular joint (TMJ) associated with aging can affect mandibular shape and reduce growth potential when stimulated by functional appliance therapy. This study was designed to evaluate the morphological changes in the mandibles of male mice associated with aging and biomechanical stimulus. Every 3 days over the course of 1 month, the lower incisors were trimmed by 1 mm to induce mandibular advancement (MA) when the animal was feeding. The left mandibles of the 23 experimental and 27 control animals were subsequently dissected, and digital images were obtained to analyze nine linear/angular measurements. Because mandibular morphology depends on the maintenance of condylar cartilage, the surfaces of the condylar cartilage and the ascending ramus of the mandible were also analyzed by scanning electron microscopy (SEM). The linear measurements of the mandible showed changes according to age in the control group and a growth response in the mandibular condyle in 7‐ and 15‐month‐old mice after MA. Moreover, SEM analysis revealed depressions in the anterior region of the condylar cartilage and inclined vascular grooves in the ascending ramus in the 7‐ and 15‐month‐old experimental mice. Although the growth potential is reduced in mice after 6 months of age, the results showed that continuous growth of the mandible occurs after maturation, except in the condyle, and that biomechanical stimulus of the TMJ of male mice leads to condylar growth. These results suggest that mature and old individuals can favorably respond to maxillary functional orthopedic therapy. Anat Rec, 292:431–438, 2009. © 2009 Wiley‐Liss, Inc.     

Mandibular coronoid process in parathyroid hormone-related protein-deficient mice shows ectopic cartilage formation accompanied by abnormal bone modeling

Parathyroid hormone-related protein (PTHrP) null mutant mice were analyzed to investigate an additional role for PTHrP in cell differentiation. We found ectopic cartilage formation in the mandibular coronoid process in newborn mice. While many previous studies involving PTHrP gene knockout mouse have shown that the cartilage in various regions becomes smaller, this is the first report showing an "increase" of cartilage volume. Investigations of mandibular growth using normal mice indicated that coronoid secondary cartilage never formed from E 15 to d 4, but small amount of cartilage temporally formed at d 7, and this also applies to PTHrP-wild type mice. Therefore, PTHrP deficiency consequently advanced the secondary cartilage formation, which is a novel role of PTHrP in chondrocyte differentiation. In situ hybridization of matrix proteins showed that this coronoid cartilage had characteristics of the lower hypertrophic cell zone usually present at the site of endochondral bone formation and/or "chondroid bone" occasionally found in distraction osteogenesis. In addition, the coronoid process in the PTHrP-deficient mouse also showed abnormal expansion of bone marrow and an increase in the number of multinucleated osteoclasts, an indication of abnormal bone modeling. These results indicate that PTHrP is involved in bone modeling as well as in chondrocyte differentiation. In situ hybridization of matrix protein mRNAs in the abnormal mandibular condylar cartilage revealed that this cartilage was proportionally smaller, supporting previous immunohistochemical results.     

Age-related changes in the localization of glycosaminoglycans in condylar cartilage of the mandible in rats

There is little information available regarding the morphological and biomolecular characteristics of mandibular condylar cartilage. The purpose of this study was to determine the age-related changes in the morphology and immunolocalization of glycosaminoglycans (GAGs) in mandibular condyles. The mandibular condylar cartilages from 4-, 8-, 16-, 32-, and 64-week-old Wistar male rats were examined to verify the localization of chondroitin-4-sulfate (Ch-4S), chondroitin-6-sulfate (Ch-6S) and keratan sulfate (KS) using an indirect immunofluorescent technique with three monoclonal antibodies for glycosaminoglycans, 2-B-6, 3-B-3 and 5-D-4, respectively. Morphologically, the condylar cartilage was a growth cartilage during growing periods, began to differentiate into articular cartilage from the central area of 16-week-old condyles, and became mature articular cartilage at 32 weeks of age. A regional difference was found in the morphological features and distribution of GAGs between the anterior, central, postero-superior and posterior areas of the condyles at each age. The immunohistochemical localizations of these three glycosaminoglycans showed age-related, morphology-dependent changes, from growth cartilage to articular cartilage-like cartilage. Immunoreactions for all of the antibodies decreased progressively with age in the interterritorial matrix, while the pericellular and territorial matrix in the condylar cartilage of the mandible maintained relatively higher immunoreactivity. In conclusion, age-related and regional differences in the localization of glycosaminoglycans Ch-4S, Ch-6S, and KS were found in the mandibular condyles in rats, and these changes are believed to be related to functional and developmental requirements.     

功能性下颌偏斜青春期模型大鼠髁突软骨中血管内皮细胞生长因子的表达

背景：对于功能性下颌偏斜是否会导致骨性的下颌偏斜以及发生功能性下颌偏斜后下颌骨的生长方式是否会发生改变,目前尚存在争议。血管内皮细胞生长因子能够使内皮细胞的渗透性增加、刺激血管内皮细胞的有丝分裂、促进血管生成以及调节骨的形成。 目的：探索大鼠功能性下颌偏斜对其髁突软骨中血管内皮细胞生长因子表达的影响及意义。 方法：选4周龄雄性SD大鼠60只,随机分为对照组(n=20)和实验组(n=40)。实验组大鼠佩戴作者所在课题组自制镍铬合金上前牙冠套,使下颌发生功能性左偏(2.1±0.3) mm,模拟功能性下颌偏斜。 结果与结论：造模后7-28 d功能性下颌偏斜模型大鼠髁突软骨矢状向后部区域偏斜侧血管内皮细胞生长因子的阳性细胞数量较非偏斜侧减少;造模后14-28 d功能性下颌偏斜模型大鼠髁突软骨矢状向中部及冠状向中部区域偏斜侧血管内皮细胞生长因子的阳性细胞数量较非偏斜侧减少,但与对照组接近;造模后14-28 d功能性下颌偏斜模型大鼠髁突软骨冠状向外部区域偏斜侧血管内皮细胞生长因子的阳性细胞数量较非偏斜侧和对照组减少。说明青春期大鼠发生功能性的下颌偏斜后,双侧髁突软骨中的血管内皮细胞生长因子表达存在差异,导致软骨内的成骨活动发生变化,进而可能使大鼠下颌骨的正常生长发育受到影响,提示应尽早矫治功能性下颌偏斜。     

Bone morphogenetic protein rescues the lack of secondary cartilage in Runx2-deficient mice

Secondary cartilages including mandibular condylar cartilage have unique characteristics. They originate from alkaline phosphatase (ALP)-positive progenitor cells of the periosteum, and exhibit characteristic modes of differentiation. They also have a unique extracellular matrix, and coexpress type I, II and X collagens. We have previously shown that there is a total absence of secondary cartilages in Runx2-deficient (Runx2-/-) mice. To clarify whether Runx2 is essential for chondrocytic differentiation of secondary cartilages, we performed an organ culture system using mandibular explants derived from Runx2-/- mice at embryonic day 18.0. Since mRNA for bone morphogenetic protein 2 (BMP2) was strongly expressed in osteoblasts of condylar anlagen in wild-type mice, and was down-regulated in those of Runx2-/- mice, we chose to investigate BMP2 effects on secondary cartilage formation. Condensed mesenchymal cells of mandibular condylar anlagen in precultured explants were ALP-positive and expressed type I collagen and Sox9. After culture with recombinant human (rh) BMP2, chondrocytic cells showing ALP activity and expressing Sox5, Sox9, and type I and II collagens, appeared from mesenchymal condensation. This expression profile was comparable with the reported pattern of chondrocytes in mouse secondary cartilages. However, chondrocyte hypertrophy was not observed in the explants. These findings indicate that BMP2 partially rescued chondrocyte differentiation but not chondrocyte hypertrophy in secondary cartilage formation in Runx2-/- mice. Runx2 is required for chondrocyte hypertrophy in secondary cartilage formation, and it is likely that BMP2, which is abundantly secreted by osteoblasts in condylar anlagen, contributes to the early process of secondary cartilage formation.     

Unilateral masticatory function changes the proteoglycan content of mandibular condylar cartilage in rabbit

Unilateral masticatory function was induced in 10-day-old rabbits by grinding the right side molars out of occlusion under general anesthesia. The grinding procedure was repeated twice a week thereafter. They were killed at the ages of 25 and 35 days. Rabbits of the same age with uninterfered occlusions served as controls. The changes in articular cartilage of the mandibular condyle were studied biochemically and histochemically. There was a significant decrease in the proteoglycan content of the condylar cartilage in both groups of animals subjected to molar grinding compared to the animals with untouched occlusions. Especially the amount of aggregating proteoglycans was reduced. The condylar cartilage matrix synthesis is sensitive to loading produced by masticatory function during growth. The mechanical properties of the articular cartilage after a period of unilateral mastication will be impaired and it is possible that this makes the joint cartilage more susceptible to pathological events.     

An ultrastructural study of osteoclasts and chondroclasts in poorly calcified mandible induced by high doses of strontium diet to fetal mice

A high dose strontium diet was fed to fetal mice from day 1 of gestation to birth in order to investigate the ultrastructural changes of osteoclasts/chondroclasts when associated with poorly calcified bone/cartilage. Calcification in the mandibular bone and condylar cartilage was extensively inhibited by this diet. Multinucleated osteoclasts and chondroclasts were observed on the mandibular alveolar bone and in the resorption area of the condylar cartilage, respectively. However, both cell types never formed ruffled borders and clear zones at the cell surfaces facing the matrices indicative of bone resorption, although they had well-developed organelles and vacuoles. Furthermore, they revealed signs of phagocytosis of the matrix vesicles. These results indicate that osteoclasts/chondroclasts can exhibit phagocytotic activity in response to requirements.     

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.     

Functional analysis of CTRP3/cartducin in Meckel's cartilage and developing condylar cartilage in the fetal mouse mandible

CTRP3/cartducin, a novel C1q family protein, is expressed in proliferating chondrocytes in the growth plate and has an important role in regulating the growth of both chondrogenic precursors and chondrocytes in vitro. We examined the expression of CTRP3/cartducin mRNA in Meckel's cartilage and in condylar cartilage of the fetal mouse mandible. Based on in situ hybridization studies, CTRP3/cartducin mRNA was not expressed in the anlagen of Meckel's cartilage at embryonic day (E)11.5, but it was strongly expressed in Meckel's cartilage at E14.0, and then reduced in the hypertrophic chondrocytes at E16.0. CTRP3/cartducin mRNA was not expressed in the condylar anlagen at E14.0, but was expressed in the upper part of newly formed condylar cartilage at E15.0. At E16.0, CTRP3/cartducin mRNA was expressed from the polymorphic cell zone to the upper part of the hypertrophic cell zone, but was reduced in the lower part of the hypertrophic cell zone. CTRP3/cartducin-antisense oligodeoxynucleotide (AS-ODN) treatment of Meckel's cartilage and condylar anlagen from E14.0 using an organ culture system indicated that, after 4-day culture, CTRP3/cartducin abrogation induced curvature deformation of Meckel's cartilage with loss of the perichondrium and new cartilage formation. Aggrecan, type I collagen, and tenascin-C were simultaneously immunostained in this newly formed cartilage, indicating possible transformation from the perichondrium into cartilage. Further, addition of recombinant mouse CTRP3/cartducin protein to the organ culture medium with AS-ODN tended to reverse the deformation. These results suggest a novel function for CTRP3/cartducin in maintaining the perichondrium. Moreover, AS-ODN induced a deformation of the shape, loss of the perichondrium/fibrous cell zone, and disorder of the distinct architecture of zones in the mandibular condylar cartilage. Additionally, AS-ODN-treated condylar cartilage showed reduced levels of mRNA expression of aggrecan, collagen types I and X, and reduced BrdU-incorporation. These results suggest that CTRP3/cartducin is not only involved in the proliferation and differentiation of chondrocytes, but also contributes to the regulation of mandibular condylar cartilage.     

Tissue engineering the mandibular condyle

Tissue engineering provides the revolutionary possibility for curing temporomandibular joint (TMJ) disorders. Although characterization of the mandibular condyle has been extensively studied, tissue engineering of the mandibular condyle is still in an inchoate stage. The purpose of this review is to provide a summary of advances relevant to tissue engineering of mandibular cartilage and bone, and to serve as a reference for future research in this field. A concise anatomical overview of the mandibular condyle is provided, and the structure and function of the mandibular condyle are reviewed, including the cell types, extracellular matrix (ECM) composition, and biomechanical properties. Collagens and proteoglycans are distributed heterogeneously (topographically and zonally). The complexity of collagen types (including types I, II, III, and X) and cell types (including fibroblast-like cells, mesenchymal cells, and differentiated chondrocytes) indicates that mandibular cartilage is an intermediate between fibrocartilage and hyaline cartilage. The fibrocartilaginous fibrous zone at the surface is separated from hyaline-like mature and hypertrophic zones below by a thin and highly cellular proliferative zone. Mechanically, the mandibular condylar cartilage is anisotropic under tension (stiffer anteroposteriorly) and heterogeneous under compression (anterior region stiffer than posterior). Tissue engineering of mandibular condylar cartilage and bone is reviewed, consisting of cell culture, growth factors, scaffolds, and bioreactors. Ideal engineered constructs for mandibular condyle regeneration must involve two distinct yet integrated stratified layers in a single osteochondral construct to meet the different demands for the regeneration of cartilage and bone tissues. We conclude this review with a brief discussion of tissue engineering strategies, along with future directions for tissue engineering the mandibular condyle.     

An in situ hybridization study of the Syndecan family in the developing condylar cartilage of fetal mouse mandible

Mandibular condylar cartilage is a representative secondary cartilage, differing from primary cartilage in various ways. Syndecan is a cell-surface heparan sulfate proteoglycan and speculated to be involved in chondrogenesis and osteogenesis. This study aimed to investigate the expression patterns of the syndecan family in the developing mouse mandibular condylar cartilage. At embryonic day (E)13.0 and E14.0, syndecan-1 and -2 mRNAs were expressed in the mesenchymal cell condensation of the condylar anlage. When condylar cartilage was formed at E15.0, syndecan-1 mRNA was expressed in the embryonic zone, wherein the mesenchymal cell condensation is located. Syndecan-2 mRNA was mainly expressed in the perichondrium. At E16.0, syndecan-1 was expressed from fibrous to flattened cell zones and syndecans-2 was expressed in the lower hypertrophic cell zone. Syndecan-3 mRNA was expressed in the condylar anlage at E13.0 and E13.5 but was not expressed in the condylar cartilage at E15.0. It was later expressed in the lower hypertrophic cell zone at E16.0. Syndecan-4 mRNA was expressed in the condylar anlage at E14.0 and the condylar cartilage at E15.0 and E16.0. These findings indicated that syndecans-1 and -2 could be involved in the formation from mesenchymal cell condensation to condylar cartilage. The different expression patterns of the syndecan family in the condylar and limb bud cartilage suggest the functional heterogeneity of chondrocytes in the primary and secondary cartilage.     

Differential responses to parathyroid hormone-related protein (PTHrP) deficiency in the various craniofacial cartilages.

PTHrP null mutant mice exhibit skeletal abnormalities both in the craniofacial region and limbs. In the growth plate cartilage of the null mutant, a diminished number of proliferating chondrocytes and accelerated chondrocytic differentiation are observed. In order to examine the effect of PTHrP deficiency on the craniofacial morphology and highlight the differential feature of the composing cartilages, we examined the various cartilages in the craniofacial region of neonatal PTHrP deficient mice. The major part of the cartilaginous anterior cranial base appeared to be normal in the homozygous PTHrP deficient mice. However, acceleration of chondrocytic differentiation and endochondral bone formation was observed in the posterior part of the anterior cranial base and in the cranial base synchondroses. Ectopic bone formation was observed in the soft tissue-running mid-portion of the Meckel's cartilage, where the cartilage degenerates and converts to ligament in the course of normal development. The zonal structure of the mandibular condylar cartilage was scarcely affected, but the whole condyle was reduced in size. These results suggest the effect of PTHrP deficiency varies widely between the craniofacial cartilages, according to the differential features of each cartilage.     

Distribution of Matrix Proteins in Perichondrium and Periosteum During the Incorporation of Meckel's Cartilage into Ossifying Mandible in Midterm Human Fetuses: An Immunohistochemical Study

Immunohistochemical localization of versican and tenascin‐C were performed; the periosteum of ossifying mandible and the perichondrium of Meckel's cartilage, of vertebral cartilage, and of mandibular condylar cartilage were examined in midterm human fetuses. Versican immunoreactivity was restricted and evident only in perichondrium of Meckel's cartilage and vertebral cartilage; conversely, tenascin‐C immunoreactivity was only evident in periosteum. Therefore, versican and tenascin‐C can be used as molecular markers for human fetal perichondrium and fetal periosteum, respectively. Meckel's cartilage underwent endochondral ossification when it was incorporated into the ossifying mandible at the deciduous lateral incisor region. Versican immunoreactivity in the perichondrium gradually became weak toward the anterior primary bone marrow. Tenascin‐C immunoreactivity in the primary bone marrow was also weak, but tenascin‐C positive areas did not overlap with versican‐positive areas; therefore, degradation of the perichondrium probably progressed slowly. Meanwhile, versican‐positive perichondrium and tenascin‐C‐positive periosteum around the bone collar in vertebral cartilage were clearly discriminated. Therefore, the degradation of Meckel's cartilage perichondrium during endochondral ossification occurred at a different rate than did degradation of vertebral cartilage perichondrium. Additionally, the perichondrium of mandibular condylar cartilage showed tenascin‐C immunoreactivity, but not versican immunoreactivity. That perichondrium of mandibular condylar cartilage has immunoreactivity characteristic of other periosteum tissues may indicate that this cartilage is actually distinct from primary cartilage and representative of secondary cartilage. Anat Rec, 297:1208–1217, 2014. © 2014 Wiley Periodicals, Inc.     

Morphological influence of ascorbic acid deficiency on endochondral ossification in osteogenic disorder Shionogi rat

The influences of chronic deficiency of L-ascorbic acid (AsA) on the differentiation of osteo-chondrogenic cells and the process of endochondral ossification were examined in the mandibular condyle and the tibial epiphysis and metaphysis by using Osteogenic Disorder Shionogi (ODS) rats that bear an inborn deficiency of L-gulonolactone oxidase. Weanling male rats were kept on an AsA-free diet for up to 4 weeks, until the symptoms of scurvy became evident. The tibiae and condylar processes of scorbutic rats displayed undersized and distorted profiles with thin cortical and scanty cancellous bones. In these scorbutic bones, the osteoblasts showed characteristic expanded round profiles of rough endoplasmic reticulum, and lay on the bone surface where the osteoid layer was missing. Trabeculae formation was deadlocked, although calcification of the cartilage matrix proceeded in both types of bone. Scorbutic condylar cartilage showed severe disorganization of cell zones, such as unusual thickening of the calcification zone, whereas the tibial cartilage showed no particular alterations (except for a moderately decreased population of chondrocytes). In condylar cartilage, hypertrophic chondrocytes were encased in a thickened calcification zone, and groups of nonhypertrophic chondrocytes occasionally formed cell nests surrounded by a metachromatic matrix in the hypertrophic cell zone. These results indicate that during endochondral ossification, chronic AsA deficiency depresses osteoblast function and disturbs the differentiation pathway of chondrocytes. The influence of scurvy on mandibular condyle cartilage is different from that on articular and epiphyseal cartilage of the tibia, suggesting that AsA plays different roles in endochondral ossification in the mandibular condyle and long bones.     

In situ hybridization and immunohistochemistry of bone sialoprotein and secreted phosphoprotein 1 (osteopontin) in the developing mouse mandibular condylar cartilage compared with limb bud cartilage

Mandibular condylar cartilage is often classified as a secondary cartilage, differing from the primary cartilaginous skeleton in its rapid progress from progenitor cells to hypertrophic chondrocytes. In this study we used in situ hybridization and immunohistochemistry to investigate whether the formation of primary (tibial) and secondary (condylar) cartilage also differs with respect to the expression of two major non‐collagenous glycoproteins of bone matrix, bone sialoprotein (BSP) and secreted phosphoprotein 1 (Spp1, osteopontin). The mRNAs for both molecules were never expressed until hypertrophic chondrocytes appeared. In the tibial cartilage, hypertrophic chondrocytes first appeared at E14 and the expression of BSP and Spp1 mRNAs was detected in the lower hypertrophic cell zone, but the expression of BSP mRNA was very weak. In the condylar cartilage, hypertrophic chondrocytes appeared at E15 as soon as cartilage tissue appeared. The mRNAs for both molecules were expressed in the newly formed condylar cartilage, although the proteins were not detected by immunostaining; BSP mRNA in the condylar cartilage was more extensively expressed than that in the tibial cartilage at the corresponding stage (first appearance of hypertrophic cell zone). Endochondral bone formation started at E15 in the tibial cartilage and at E16 in the condylar cartilage. At this stage (first appearance of endochondral bone formation), BSP mRNA was also more extensively expressed in the condylar cartilage than in the tibial cartilage. The hypertrophic cell zone in the condylar cartilage rapidly extended during E15–16. These results indicate that the formation process of the mandibular condylar cartilage differs from that of limb bud cartilage with respect to the extensive expression of BSP mRNA and the rapid extension of the hypertrophic cell zone at early stages of cartilage formation. Furthermore, these results support the hypothesis that, in vivo, BSP promotes the initiation of mineralization.