Mei-Qing Wang

Subchondral bone loss following orthodontically induced cartilage degradation in the mandibular condyles of rats

Osteoarthritis (OA) is a degenerative joint disease generally characterized by progressive cartilage degradation and subchondral bone changes. Subchondral bone changes have been proposed to initiate or accompany with cartilage degradation in OA. The purpose of this study was to characterize cartilage damage, subchondral bone remodeling, and the possible mechanism involved in these morphological changes in our reported rat model with OA-like lesions in the mandibular condyle. In experimental groups, the dental occlusion was orthodontically disturbed. By histological analysis, transmission electron microscopy (TEM), micro-CT scanning and serum tests, changes in condylar cartilage and subchondral bone were analyzed at 8 and 12 weeks after treatment. The mRNA and protein levels of bone pro-resorptive and pro-formative factors by chondrocytes were investigated. Increased degraded cartilage areas and obvious cartilage calcification were observed in 8- and 12-week treated (EXP) groups compared to the age-matched controls. Subchondral bone loss, characterized as decreased bone mineral density (BMD), bone volume fraction (BV/TV) and trabecular thickness (Tb.Th), but increased trabecular separation (Tb.Sp), was observed in the 12-week but not the 8-week EXP group, respectively, versus their age-matched controls. The subchondral bone loss in the 12-week EXP group was accompanied with decreased new bone formation rate, but increased serum carboxy terminal telopeptides (CTXs), and increased osteoclast numbers and proportion of surface area in the subchondral bone regions. Increased mRNA and protein levels of M-CSF, VEGF, RUNX and RANKL/OPG ratio, but decreased OPG, were found in condylar cartilage in the 12-week EXP group versus its age-matched controls, and those of RANKL/OPG ratios were significantly higher in the 12-week EXP group than the 8-week EXP. In addition, increased mRNA levels of VEGF, RUNX and RANKL/OPG ratio, but decreased OPG, were also found in condylar cartilage in the 8-week EXP group versus its age-matched controls (All P<0.05). This study demonstrated that obvious subchondral bone loss followed cartilage degradation in the mandibular condyles in the present rat models and suggested that the imbalance of chondrocyte-secreted regulatory factors within the degraded cartilage may play a role in the osteoclastogenesis, and thus leading to the subchondral bone loss in OA.

Occlusal Effects on Longitudinal Bone Alterations of the Temporomandibular Joint

The pathological changes of subchondral bone during osteoarthritis (OA) development in the temporomandibular joint (TMJ) are poorly understood. In the present study, we investigated the longitudinal alterations of subchondral bone using a rat TMJ-OA model developed in our laboratory. Changes in bone mass were examined by micro-CT, and changes in osteoblast and osteoclast activities were analyzed by real-time PCR, immunohistochemistry, and TRAP staining. Subchondral bone loss was detected from 8 weeks after dental occlusion alteration and reached the maximum at 12 weeks, followed by a repair phase until 32 weeks. Although bone mass increased at late stages, poor mechanical structure and lower bone mineral density (BMD) were found in these rats. The numbers of TRAP-positive cells were increased at 12 weeks, while the numbers of osteocalcin-expressing cells were increased at both 12 and 32 weeks. Levels of mRNA expression of TRAP and cathepsin K were increased at 12 weeks, while levels of ALP and osteocalcin were increased at both 12 and 32 weeks. These findings demonstrated that there is an active bone remodeling in subchondral bone in TMJs in response to alteration in occlusion, although new bone was formed with lower BMD and poor mechanical properties.

Changes in the expression of MMP-3, MMP-9, TIMP-1 and aggrecan in the condylar cartilage of rats induced by experimentally created disordered occlusion

OBJECTIVE To investigate the effects of experimentally created disordered occlusion on the mandibular condylar cartilage in terms of histological morphology and expression of MMP-3, MMP-9, TIMP-1 and aggrecan. MATERIALS AND METHODS Eighty 8-week-old Sprague-Dawley rats were randomly divided into two experimental (Exp) and two control (Con) groups, with equal sex and number distribution as subgroups. In the Exp group, the disordered occlusion was created by orthodontically moving the first and third molars 0.8mm away. Hematoxylin-eosin and immunohistochemical staining were performed on the mandibular condyles at the end of the 8th or 12th week. Gene expression was analysed by real-time PCR. RESULTS Osteoarthritis-like lesions, typically seen as a cell-free area, were detected in the Exp group, predominantly in females. In the cell-free area, the immunopositive expression of MMP-3, MMP-9, TIMP-1 and aggrecan were absent. Hyper-proliferation changes, typically seen as conjunctive invaginations of chondrocytes, were also observed where immunopositive expression of the tested materials was strong. There were sex and time point related differences in gene expression. In the 8-week subgroup, the expression of MMP-3 decreased, while aggrecan increased in males; however, both MMP-9 and TIMP increased in the female group (P<0.05). In the 12-week subgroup, the expression of MMP-3 increased, while TIMP, MMP-9 (male only) and aggrecan (female only) decreased (P<0.05). CONCLUSIONS The present results indicate that the experimentally created disordered occlusion led to osteoarthritis-like lesions accompanied by changes in the expression of MMP-3, MMP-9, TIMP-1 and aggrecan in mandibular condyle cartilage with gender differences.

Age- and sex-related changes of mandibular condylar cartilage and subchondral bone: A histomorphometric and micro-CT study in rats

OBJECTIVE To quantify the age- and sex-related changes in the rat condylar cartilage and subchondral bone. METHODS SD rats were obtained at the ages of 2, 3, 4, 5, 6 and 7 months. For each sex, the temporomandibular joints tissue blocks from four rats were subjected to histological assessment of cartilage thickness and subchondral bone architecture; for the remaining three rats, the mandibular condyles were delivered for gross measurement and evaluation of the mineralization and architecture properties of the subchondral bone by means of micro-CT. RESULTS Rapid decrease of cartilage thickness but increase of subchondral bone density occurred respectively from 2 to 3 and 3 to 4 months old in female and 2 to 4 and 3 to 5 months old in male (P<0.05), whereas rapid changes of subchondral bone architecture occurred from 3 to 4 months old in both sexes (P<0.05). The significant enlargement of condyle size occurred at 4 or 5 months old in female but at 5 or 6 months in male (P<0.05). CONCLUSION This study demonstrated that the rapid developmental changes of rat condylar cartilage and subchondral bone primarily occurred before 4 months of age, resulting in thinner cartilage but larger and thicker subchondral bone, and they were followed by rapid growth in condylar size. Sex differences were identified that the endochondral ossification of fibrocartilage and formation of subchondral bone were faster in female than in male rats, leading to the earlier enlargement of condyle in female than in male.

Mandibular condylar cartilage response to moving 2 molars in rats.

INTRODUCTION The purpose of this study was to investigate the responses of mandibular condylar cartilage to moving 2 molars in different combinations. METHODS Rats were assigned to male and female control and experimental groups (each, n = 5). Elastic rubber bands were used to move medially the maxillary left and the mandibular right first molars in experimental group I. The same method was used to distally move the maxillary left and the mandibular right third molars, 2 mandibular third molars, and 2 maxillary third molars in experimental groups II, III, and IV, respectively. At the end of the eighth week, all condyles were examined histologically. The areas of histologic change as a percentage of total cartilage area were compared by using the Mann-Whitney U test. RESULTS Cartilage degenerative remodeling was observed in experimental groups II, III, and IV. The percentage areas of degenerative remodeling were higher in female experimental groups II and III than in the female control group, and in female experimental group II than in female experimental group IV and male experimental group II (all, P <0.05). CONCLUSIONS The mandibular condylar cartilage of female rats responded variously to different combinations of molar movement; the most obvious remodeling was observed in groups in which the maxillary left and mandibular right third molars were moved.

The effects of age and sex on the expression of oestrogen and its receptors in rat mandibular condylar cartilages

OBJECTIVE Oestrogen expression may indicate a difference in resistance potential to mechanical strain. The purpose of this study was to investigate the expression of oestrogen and oestrogen receptors in mandibular condylar cartilages in male and female Sprague-Dawley rats at different ages. MATERIALS AND METHODS One-hundred SD rats at the age of 2, 4, 8 weeks and 4, 12 months in both sexes, 10 in each age-sex group, were enrolled in this study. The expression of oestradiol, ERalpha and ERbeta was detected in mandibular condylar cartilages by the method of immunohistochemistry, and enzyme-linked immunosorbent assay or western blot. RESULTS Oestradiol and ERs immunoreactivity were obvious in mandibular condylar cartilages of SD rats. Oestradiol and ERalpha were observed in hypertrophic and mature layers, while ERbeta only in hypertrophic layer. There was no sex difference of same age (except 8-week age group) in the expression of oestradiol. The expression of both ERs, however, was usually higher in male than in age-matched female rats (P<0.05), except that the 8-week-old female rats showed a higher ERalpha expression and the 4- and 8-week-old female rats showed a higher ERbeta expression than the age-matched male ones in western blot results (P<0.05). CONCLUSIONS The results that oestradiol, ERalpha and ERbeta are co-expressed in rat mandibular condylar cartilage, indicate that mandibular condylar cartilage is a target for oestrogen. The age and sex related differences in ERs expression may indicate a difference in potential to resist mechanical loading between genders at different ages.

Osteochondral angiogenesis in rat mandibular condyles with osteoarthritis-like changes

OBJECTIVE To investigate angiogenesis at the osteochondral junction and changes in expression of pro- and anti-angiogenic factors in rat mandibular condyles with osteoarthritis-like changes. METHODS In order to evoke osteoarthritis-like lesions in mandibular condyles, disordered occlusion was created experimentally in rats. Osteochondral vascularity was assessed histologically at 20 and 24 weeks. Protein and mRNA levels of pro-angiogenic factors including vascular endothelial growth factor (VEGF), connective tissue growth factor (CTGF) and matrix metalloproteases 9 (MMP9), and anti-angiogenic factor chondromodulin-I (CHM-I) were investigated by means of immunohistochemical staining and real-time PCR. RESULTS Osteochondral angiogenesis was demonstrated as increased numbers of vascular channels terminating in the calcified cartilage and non-calcified cartilage in 20- and 24-week experimental groups compared with controls (all P<0.05). In the experimental groups, VEGF, CTGF and MMP9 were highly expressed in the tissues adjacent to the osteochondral junction. However, CHM-I was more expressed in the superior but not deep hypertrophic chondrocytes. Compared to their age-matched controls, the protein levels of VEGF and CTGF were higher in 20-week experimental group, and the protein and mRNA levels of CTGF, MMP-9, and CHM-I increased in the 24-week experimental group (all P<0.05). CONCLUSION In the present rat model, osteochondral angiogenesis was observed in mandibular condyles with osteoarthritis-like changes, accompanied with local upregulation of VEGF, CTGF and MMP9. Although the increase in CHM-I may moderate pro-angiogenic factors effects in the superior cartilage, the deficiency of deep hypertrophic chondrocytes to express CHM-I may permit vascular invasion into condylar cartilage.

Stable tooth contacts in intercuspal occlusion makes for utilities of the jaw elevators during maximal voluntary clenching

Data are inconsistent concerning whether the level of the surface electromyographic (SEMG) activity of jaw-closing muscles increases when biting forces elevated during maximal voluntary clenching (MVC). In this study, T-Scan III system and BioEMG III system were used to record bite force, occlusal contacts and SEMG activity of the anterior temporalis (TA) and of the masseter muscles (MM) simultaneously. Recordings were obtained from 16 healthy young adult males during different conditions: (i) a fast MVC from resting position to intercuspal position (ICP); (ii) mandibular movements from ICP to protrusive or lateral edge-to-edge positions with teeth in contact with biting; (iii) a fast MVC in protrusive and lateral edge-to-edge positions. A higher level of SEMG activity was associated with a higher bite force during occluding movements (P < 0.05). However, during fast MVC from rest to ICP, the largest number of occlusal contacts was achieved and distributed more symmetrically, the highest level of biting force was obtained, but the SEMG activity of the jaw elevator muscles was reduced compared with its maximum level (P < 0.05). This phenomenon was not observed during the fast MVC in protrusive or lateral edge-to-edge positions. The present results that a lower SEMG activity was associated with the largest number of occlusal contacts and the highest level of bite force during centric MVC demonstrated a complex integration of jaw-closing muscles when a stable occlusion is present.

Experimentally created unilateral anterior crossbite induces a degenerative ossification phenotype in mandibular condyle of growing Sprague-Dawley rats

The effect of unilateral anterior crossbite on the remodelling of mandibular condyle needs to be investigated. This study aimed to investigate the effects of experimentally created unilateral anterior crossbite on the remodelling of mandibular condyle and explore the changes in the expression of relevant transcription factors and growth factors. The experimental unilateral anterior crossbite was created in 6-week-old female growing rats by bonding metal tubes to the left pairs of incisors. Remodelings of mandibular condylar cartilage was assessed histologically at 2, 4 and 8 weeks. Protein and mRNA levels of Sox9, runt-related transcription factor 2 (Runx2), Osterix (Osx), transforming growth factor beta 1 (TGFβ1), transforming growth factor beta receptor 2 (TGFβr2) and type X collagen (ColX) were investigated by immunohistochemistry and real-time PCR, while alkaline phosphatise (ALP) by histochemistry and real-time PCR. Decreased ratio of hypertrophic cartilage layer was noticed in the 4w experimental group versus controls. At all the time points, the expression of Sox9 and ALP increased but that of TGFβ1 and TGFβr2 decreased in experimental groups (P < 0·05). The expression of Runx2, Osx and Col X increased at 2w, but decrease at 4w (P < 0·05). The results that obvious cartilage degradation and altered expression of related transcription factors and growth factors were detected in the mandibular condyles of the experimental group suggested that the present unilateral anterior crossbite plays an adverse role in the TMJ, and thus leading to the degenerative endochondral ossification.

Excess genistein suppresses the synthesis of extracellular matrix in female rat mandibular condylar cartilage

Aim:To investigate the effect of excess genistein on the extracellular matrix in mandibular condylar cartilage of female rats in vivo.Methods:Female SD rats were administered through oral gavage with genistein (50 mg/kg) or placebo daily for 6 weeks. The morphological changes of temporomandibular joints were studied with HE staining. The expression of cartilage matrix compounds (aggrecan and collagen type II), estrogen-related molecules (aromatase, estradiol, ERα and ERβ) and proliferating cell nuclear antigen (PCNA) in mandibular condylar cartilage was detected using immunohistochemistry, ELISA and real-time PCR.Results:The genistein treatment significantly reduced the thickness of the posterior and middle regions of mandibular condylar cartilage, and decreased the expression of collagen type II, aggrecan and PCNA. Compared with the control group, the estradiol content and expression levels of the key estradiol-synthesizing enzyme aromatase in the genistein-treatment group were significantly decreased. The genistein treatment significantly increased the expression of ERβ, but decreased the expression of ERα.Conclusion:Excess genistein suppresses extracellular matrix synthesis and chondrocytes proliferation, resulting in thinner mandibular condylar cartilage. These effects may be detrimental to the ability of mandibular condylar cartilage to adapt to mechanical loads.