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.

Location of the glenoid fossa after a period of unilateral masticatory function in young rabbits.

Changes in glenoid fossa position and skull morphology after a period of unilateral masticatory function were studied. The right-side maxillary and mandibular molars in twenty-seven 10-day-old rabbits were ground down under general anaesthesia. The procedure was repeated twice a week, until the rabbits were 50 days old. Fourteen rabbits were then killed and 13 left to grow to age 100 days. Nine 50-day-old and sixteen 100-day-old rabbits with unmodified occlusions served as controls. Three-dimensional measurements were made using a machine-vision technique and a video-imaging camera. The glenoid fossa position become more anterior in both groups of animals subjected to molar grinding as compared with controls (P < 0.01 in the 50-day-old group and P < 0.05 in 100-day-old group). In the 100-day-old group the right-side fossa was also in a more inferior position (P < 0.01). The glenoid fossa was more anteriorly located on the right than on the left side of individual animals in the group in which the right-side molars had been ground down (P < 0.001).

Distribution and characterization of proliferative cells in the rat mandibular condyle during growth.

Distribution of proliferative cells and localization of types I and II collagen were examined in the rat mandibular condylar cartilage of 36 long-Evans/Turku strain rats during normal postnatal growth using an immunohistochemical method combined with histomorphometry. There were considerable differences in the thickness of the proliferative cell layer in the condylar head, with most mitoses occurring in the postero-superior area. It was found that the extracellular matrix of the proliferative cells does not stain for type II collagen in 20-day-old and older rats, and that besides the subchondral bone, the strongest intensity for type I collagen stain was always localized in the articular surface of the condylar head. Statistically significant overlapping of the proliferative cell layer and the one secreting type II collagen occurred during the earlier stages of development, particularly in the postero-superior area of the condylar head. As type II collagen is considered to be a marker for identification of typical cartilage cells, the findings indicate that, in addition to undifferentiated cells, a portion of the proliferative cells can be characterized as chondroblasts during the early postnatal period in rats, but not in the later stages of development. The developmental phase of the condylar cartilage should therefore be taken into consideration when the effect of various biomechanical and humoral/hormonal factors on growth of the condylar cartilage is examined.

Comparison of amounts and properties of collagen and proteoglycans in condylar, costal and nasal cartilages.

In this study we compared the compositions of extracellular matrices of condylar, costal and nasal cartilages to characterize differences in growth patterns in relation to matrix composition. Condylar, costal and nasal cartilages of 25- and 35-day-old rabbits were extracted and subjected to bio- and histochemical analysis to determine total amounts of collagen and amounts and aggregating properties of proteoglycans. We found that proteoglycan content and aggregate formation were greatest in nasal cartilage, and lower in costal and markedly lower in condylar cartilage. The amount of proteoglycans increased by varying amounts in all samples with age. Collagen content was highest in costal cartilage. In 25-day-old rabbits the quantity of collagen in condylar cartilage exceeded that in nasal cartilage. In 35-day-old rabbits the quantities were nearly the same. It is suggested that collagen does not only provide tensile strength, but counteracts forces responsible for interstitial growth such as osmotic pressure. Based on the results, it seems that the amount of proteoglycans is greater in cartilages, which have greater independent growth potential. Variations in increase in amount of proteoglycans with age could reflect differences in the timing of growth of such cartilages.

Effect of cytochalasin D on articular cartilage cell phenotype and shape in long-term organ culture.

It has been documented, on the basis of cell culture experiments, that cytochalasin treatment promotes a round cell shape in chondroblast cultures by altering the cytoskeleton, and that it simultaneously alters the balance between production of type I and type II collagens. The aim in this study was to monitor the deposition of pro-type-I and type II collagens, and possible changes in articular cartilage layers in the mandibular condyle of the mouse under the influence of Cytochalasin D (CD) when total craniomandibular joints of 5-day-old mice were cultured in one block. The experimental group comprised 20 Balb/c mice of both sexes. Twenty in vitro controls were cultured without the administration of cytochalasin. The mice in the third group were used as in vivo controls. The cells in the prechondroblast layer responded with a rapid change in shape when treated with CD and assumed a rounded morphology. The total thickness of the cell layer was reduced at 7 days. Immunostaining against pro-type-I collagen was intense in the narrow fibrous and prechondroblast layers in the CD-treated group, whereas the stained area was wider and the staining gradually reduced in the deeper cartilage layers in the in vitro controls. Staining against type II collagen became weaker at the end of the culturing period of the CD-treated group, whereas in the in vitro controls the staining against type II collagen was clearly visible at all observation times. These phenomena can be explained by changes in differentiation and the altered cell cycle of the chondroblasts in organ culture under the influence of CD.

Changes in proteoglycan and collagen content in the mandibular condylar cartilage of the rabbit caused by an altered relationship between the condyle and glenoid fossa.

Twenty 5-day-old New Zealand rabbits underwent surgery to induce premature synostosis of the cranial sutures, resulting in posterior displacement of the glenoid fossa. Twenty sham-operated rabbits served as controls. The animals were killed at age 15 days for histochemical and biochemical analyses. The collagen content of the superior region of the condyle determined biochemically was lower in treated animals than in controls. Biochemical and histochemical analyses revealed the proteoglycan content to be significantly reduced in the superior region of the condyle (P < or = 0.001). Low levels of aggregating proteoglycans were seen. Since levels of aggregating proteoglycans decreased, catabolism must have exceeded their synthesis or the monomers must have been unable to aggregate and escaped from the tissue. It is concluded that an experiment in which the location of the mandibular condyle in the glenoid fossa is changed, while causing marked reductions in amounts of both collagen and proteoglycans in the cartilage tissue of the mandibular condyle, will also induce changes resembling those observed in animal models of arthritis. It is possible that the two phenomena have similar mechanisms.

Tissue-separating capacity of growth cartilages.

The tissue-separating capacity of chondral structures has been debated for more than 30 years, and one aspect that has particularly been questioned is whether the secondary cartilage of the mandibular condyle is comparable to primary growth cartilage, e.g. the epiphyseal growth plate. The present report summarizes information gained by using a specific interosseal transplantation method. These findings lead to the conclusion that all the structures examined, i.e. the proximal epiphyseal cartilage of the tibia, the cartilage of the costochondral junction of the ribs, the basicranial synchondroses, the medial cartilage of the clavicle and the mandibular condyle, have the capacity to separate adjoining skeletal structures. The changes induced by the transplanted structures in the recipient area vary, however, suggesting a hierarchial arrangement of cartilages with regard to their tissue-separating capacity. It is suggested that the tissue-separating capacity is a basic phenomenon in the function of growth, not only of primary growth cartilages, but of secondary cartilages as well.

Craniofacial growth after a period of unilateral masticatory function in young rabbits.

Changes in craniofacial growth after a period of unilateral masticatory function were studied in rabbits. 10-day-old animals were divided into 3 groups. In Group I, mandibular and maxillary molars were ground down 2x a week on the right side under general anesthesia until age 50 days, and were thereafter left to grow without grinding. In Group II, the right-side molars were ground until age 40 days on. Between days 40 and 60, grinding was performed on the left side. The animals were thereafter left to grow. Group III consisted of unoperated control animals. All of the animals were fed whole pellets and water ad libitum, and were sacrificed at age 100 days. There were measurable differences in growth after periods of unilateral masticatory function. The mandibular ramus was higher, the condylar processus was larger sagittally, and angles between the anterior or posterior borders of the condylar process and inferior border of mandible were smaller in the treated than in the control animals, and there were differences between right and left sides of the same animal in the maxilla and mandible. The inclination of the articular surface of the glenoid fossa was steeper anteriorly on both sides in the treated than in the controls. It was concluded that growth after a period of unilateral masticatory function in young rabbits does not rectify all of the asymmetric changes in the maxilla, mandible and glenoid fossa resulting from the asymmetric function.

Combined effects of errors in frontal-view asymmetry diagnosis.

The aim of the present investigation was to determine the relative extent of geometric error and errors in point identification in postero-anterior roentgenography. In one series of tests a group of dry human skulls was used, and the same cephalometric landmarks were identified twice by two orthodontists, using postero-anterior roentgenographs, first using the dry skulls as such, and then the same skulls with metal markers inserted to show the exact locations of the cephalometric points. Consistency and normal variation in the reproducibility of head position in the cephalostat between repeated roentgenographs were studied by a photographic technique in a group of young healthy adults, measuring the extent of minor head movements. Geometric error was calculated using a computer-aided design program (CAD) by rotating the three-dimensional co-ordinates of the cephalometric landmarks and thus obtaining projection error in the frontal view. Accuracy in cephalometric point identification was best in dental landmarks and vertical orientation of superior orbital margins. Geometric error was least when landmarks near the anterior midsagittal plane, such as upper and lower dental midlines or point gonion were compared with each other. Width measurements from frontal-view cephalograms are most sensitive to minor movements in head posture. Due to combined errors, the use of width measurements in facial asymmetry diagnosis should not be used since variance in errors in landmark identification can be larger than that in actual landmark location.

The expression of types X and VI collagen and fibrillin in rat mandibular condylar cartilage. Response to mastication forces.

Types X and VI collagen and fibrillin were localized by in situ hybridization and immunohistochemical methods in the mandibular condyles of rats, and the response of these molecules to post-weaning diets of soft food, ordinary pellets, or hardened pellets was studied. Type X collagen was synthesized, particularly in conditions of soft food consistency, by cells in the perichondrium-periosteum and in the bone and by cells at the erosion front between cartilage and bone. Type X collagen synthesis diminished under higher compression forces due to mastication and with increasing age. Type VI collagen and fibrillin were synthesized by cells in the perichondrium-periosteum and by chondrocytes and by stromal osteoblasts and were not modified by higher mechanical forces. In contrast to previous findings in the growth plate of long bones, type X collagen in the mandibular condyle was not synthesized by hypertrophic chondrocytes but was associated with cells of the osteoblastic rather than the chondroblastic phenotype.

Electrical stimulation of masseter muscles maintains condylar cartilage in long-term organ culture.

When condylar cartilage is maintained under nonfunctional organ culture conditions, its phenotypic expression is altered to a premature form with less expression of the type II collagen characteristic of mature chondroblasts. The aim of this study was to examine whether, by electrical stimulation of the major masticatory muscle, the masseter muscle, chondrogenic expression could be maintained under organ culture conditions in which the jaws with the craniomandibular joint were cultured in one block. Sixty BALB/c mice of both sexes were divided randomly into three groups of equal size. Two groups were decapitated at the age of 5 days. The cranial base and mandible were dissected out in one block, and the explant was placed on its cut surface on a culture dish. The masseter muscles of the explants in one group were stimulated with an electric pulsing device delivering an AC current of a frequency of 0.7 Hz and an amplitude of 5V with hourly active and silent periods. Five experimental and five control explants were fixed after culture periods of 1, 3, 7, and 14 days. The mice in the third group were used as in vivo controls. By electrical stimulation of the masseter muscle, the phenotypic characteristics of the condylar chondroblasts, such as the deposition of type II collagen and the thickness of the cartilage layers, closely resembled the situation in vivo, while the controls in a non-functional environment gradually lost their characteristic form.

Differences in biologic response of the mandibular condyle to forward traction or opening of the mandible. An experimental study in the rat.

The effect of different mandibular postures on the growth of the mandible was studied. A total of 60 female Wistar rats were divided into 3 groups, and all the animals were anesthetized for 6 h daily from the age of 30 days onwards. The first group served as controls. The second had the mandible maintained in a protracted position, and the third had the mandible in an open position during the anesthesia. Macroscopic measurements showed the growth of the mandibular condyle to be increased in a posteroinferior direction and also in a superior direction in the animals with the mandible in an open position. The cartilage layer containing collagen type II was significantly thinned, whereas cell proliferation had significantly increased in the posterosuperior region. In association with mandibular protrusion the thickness of the cell layer containing collagen type II had increased, and cell proliferation in the posterosuperior region had significantly decreased. Significant differences in the growth of the condylar process were observed histologically and histochemically between the experimental animals, implying that the most rewarding aspect of the regulation of condylar growth seems to be the possibility to regulate the maturation rate of the cartilage cells.

Growth and type-II collagen expression in the glenoid fossa of the temporomandibular joint during altered loading: a study in the rat.

The aim of this study was to measure changes in growth of the glenoid fossa and its articular eminence after decreased loading. A further aim was to evaluate the role of mechanical forces in relation to the existence of a cartilage layer, by determining type-II collagen secretion. A total of 99 Wistar rats were used: 48 animals were fed whole pellets and 51 were fed ground pellets. At age 21 days, after weaning, the upper and lower incisors of the soft-diet group were shortened by cutting them, twice a week. Ten animals fed whole pellets and 10 fed ground pellets were injected i. p. with Alizarin red (200 mg/kg) at ages 22, 30 and 40 days, and killed at ages 30, 40 and 50 days respectively. The heads were freed from the soft tissue and the zygomatic process cut sagittally at the deepest point of the greatest transversal concavity of the eminence. Bone apposition was measured. The other animals were used for studies involving collagen II immunostaining. Bone growth decreased in the group fed ground pellets except in the anterior-most part of the glenoid fossa at 50 days. Immunohistochemical analysis revealed larger areas of anti-collagen II staining in the group fed whole pellets, most markedly in the posterior part of the glenoid fossa. Growth of the articulationg surface of the temporal component of the temporomandibular joint appears to depend on mechanical factors, such as the condyle. The underlying mechanics seem likely to be different. The presence of type-II collagen is obviously not regulated only by compressive forces but probably also by tension loading.

Effect of reduced articular function on deposition of type I and type II collagens in the mandibular condylar cartilage of the rat.

A group of rats was fed a soft diet after weaning and the incisors shortened regularly to keep them out of occlusion. The controls were fed a hard diet. Immunohistochemical techniques and image analysis were employed to investigate deposition of pro-type I collagen and type II collagen, and the thickness of articular cartilage layers in the mandibular condyle. The immunostaining against pro-type I collagen was most intense intracellularly in the fibrous and upper chondroblast layers in 30- and 50-day-old rats fed a hard diet. In the rats fed a soft diet, marked intra- and extracellular staining against pro-type I collagen was visible in the upper chondroblast and upper hypertrophic layers but also in the lower hypertrophic layer. The intensity of staining against type II collagen was weak in animals on a soft diet, while in the animals fed a hard diet the staining was intense in the superior layers of mature chondroblasts. The total number of chondroblasts recorded was reduced by 35 percent at the age of 50 days in the soft-diet compared to the hard-diet animals. The results show that the deposition of type I and II collagens, the thickness of the cartilage cell layers and the number of chondrocytes are sensitive to alterations in loading.

Effect of unilateral masticatory function on craniofacial growth in the rabbit.

In this study the main purpose was to investigate the effect of altered masticatory function on the shape and inclination of the articular surface of the glenoid fossa, and on the growth of the mandible in the rabbit. The maxillary and mandibular molars of 14, 10-d-old rabbits were ground down to the gingiva on the right side twice a week; 9 were controls. At age 50 d all rabbits were killed, soft tissue of the heads was removed, and the measurements were made. The inclination of the articular surface of the glenoid fossa was shallower on both sides in the experimental group than in the controls; the difference was greater on the right side. There were dimensional and angular differences between right and left sides of the mandible and maxilla in the experimental animals. In conclusion, unilateral masticatory function was found to cause an asymmetric craniofacial growth. Furthermore, function has an effect on the shape of the glenoid fossa, especially on the balancing side, in conjunction with asymmetric function.

Reactions of the mandible to experimentally induced asymmetrical growth of the maxilla in the rat.

The purpose of this investigation was to examine the interrelation in growth and morphogenesis between the maxilla and the mandible. Asymmetrical growth was induced in the maxilla of 10-day-old LE/T rats by means of unilateral artificial synostosis of the frontonasal and frontopremaxillary sutures. Untreated rats were used as controls. Biometric and microscopic observations were made at the ages of 30, 50, and 100 days. The arrest of sutural growth was followed by reduced sagittal growth of the maxilla, significantly more so on the treated side. Its anterior part became bent towards the treated side and rotated around the sagittal axis. In response to the primary alteration in maxillary shape, the mandible adapted by developing a secondary asymmetry in that it was shorter on the treated side than on the contralateral side throughout the experiment and shorter on the untreated side than in the untreated controls at 30 days. When the distances were adjusted for body weight, the mandibular length on the untreated side appeared to be virtually unaffected by the experimental procedure or even exceeded the control value at 50 days. The mandibular ramus was higher on the treated side than on the contralateral side at 30 days, and became bilaterally higher than in the untreated controls with increasing age. The results imply that the growth in length of the mandible follows that of the maxilla to some extent and are indicative of a restraining effect of the shortened maxilla on mandibular growth. The existence of a mechanism responsible for keeping the height of the ramus the same on both sides is suggested.

Effect of mechanical forces on chondrocyte maturation and differentiation in the mandibular condyle of the rat.

The effects of mechanical factors on the growth of the mandibular condyle were studied by monitoring the maturation of the mesenchymal cells in 55 rats. Thirty-five animals were fed normal pellet food, and 20 were fed a soft diet and their incisors were cut regularly. 3H-thymidine was injected intraperitoneally three days before death at 18, 23, or 33 days. Histologic sections showed the most advanced 3H-thymidine-labeled cells to occur deep in the cartilage, in the lower hypertrophic cell layer in anterior and posterior regions of the condyle, and in the upper hypertrophic cell layer in the superior region at the age of 18 days. A distinct difference in the maturation state of the labeled cells could also be observed between these regions. In animals fed a soft diet, maturation was slower in the superior region of the condyle and faster in the posterior region than in the normal rats. The rate at which cells stepped out of the proliferating cell pool was measured by use of monoclonal antibodies against proliferating cell nuclear antigen. The ratio between labeled cells in the proliferating cell layer and the number of labeled cells beneath it was greater in control animals than in the soft-diet animals. The rate of differentiation and maturation of mesenchymal cells into chondrocytes seems to be controlled by mechanical factors.

Effect of altered loading on condylar growth in the rat.

The aim of this investigation was to ascertain whether lessening of function by feeding rats a soft diet and cutting their incisors can change the growth of the condyle. Sixty Wistar rats were divided into two groups; a control group fed whole pellets and a soft-diet group fed ground pellets. At the age of 21 days the upper and lower incisors of the soft-diet group were shortened by cutting with a wire cutter twice a week. Ten control rats and 10 soft-diet rats were injected with alizarin red (200 mg/kg) intraperitoneally at the age of 22, 30, or 40 days and killed at the age of 30, 40, or 50 days, respectively. The heads were freed of soft tissues, and the growth of the condylar process and the mandible was measured. The height growth of the condylar process was significantly greater in the soft-diet group by 30 and 40 days, and its length growth was greater in the soft-diet group by 50 days. It is concluded that the change in the amount of chewing force and the place of articulation of the condyle alters the growth of the condylar process. Lessening of the load increases condylar growth until a new balance is achieved.

Articular disc and eminence modeling after experimental relocation of the glenoid fossa in growing rabbits.

The articular surface of the glenoid fossa shows some analogy to the mandibular condyle, since the surface is covered by secondary cartilage, which makes the process more elastic than purely bony structures. The condylar cartilage has been shown to be responsive to alterations in load pressures, and this secondary type of cartilage is also able to increase its proliferative activity to a limited extent when the load pressure is altered. The aim here was to measure changes in proliferative activity and type II collagen secretion in the articular surface of the glenoid fossa after steady experimental posterior relocation of the fossa in the rabbit without actively interfering with normal masticatory action. The shape of the articular disc and interrelations of the joint components were measured macroscopically. Twenty-four five-day-old rabbits underwent gluing of the interparietal, temporoparietal, and lambdoidal sutures. Three experimental and 3 control rabbits were injected with tritiated thymidine at 10, 15, 20, and 30 days and were killed after 2 h for histological, autoradiographic, and immunohistochemical examination. The total number of labeled cells in the proliferative layer near the articular eminence was higher in the experimental group, the difference being greatest in the 15- and 20-day-old rabbits. Immunohistochemical examination revealed less staining for type II collagen on the postero-inferior side of the eminence in the experimental group. The articular disc was flattened in the experimental group, and the elastic tissue bundle connecting the articular eminence and the anterior border of the disc was significantly narrower and longer.

Glycosaminoglycan synthesis in the mandibular condyle during growth adaptation.

Condylar growth was studied after an operation simulating functional orthodontic appliances. Twenty-five rabbits underwent a surgical operation for the induction of premature synostosis to displace the glenoid fossa posteriorly during growth. Twenty-five control rabbits underwent sham operations. At the age of 15 days, 10 experimental and 10 control animals and, at the age of 20 days, 5 experimental and 5 control animals were killed. Their mandibular condyles were organ-cultured for 3 h in the presence of radiolabelled sulphur. The condyles were used for autoradiographic purposes. Digital image analysis of autoradiograms of histological sections showed synthesis of glycosaminoglycans to have increased from the anterior to the posterior direction. This increase was more marked in experimental animals than in the condyles of control animals. Ten experimental and 10 control animals were killed at the age of 15 days, and mandibular condyles were organ-cultured for 1, 4 and 7 days. Differentiation of proliferating prechondroblasts into hypertrophied chondrocytes continued under organ culture conditions. A marked decrease in the proliferating cell layer was noticed, especially in control condyles. Hypertrophy was faster and came closer to the surface of the condyle in the anterior region of the condyle. This was most marked in the condyles of experimental animals. The results indicate that a procedure carried out on the glenoid fossa with the same effect as functional appliances increases the synthesis of extracellular matrix in the posterosuperior region of the mandibular condyle.