Skip to main content
NCBI home page
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2014 May 1.
Published in final edited form as: J Bone Miner Res. 2013 May;28(5):1127–1134. doi: 10.1002/jbmr.1835

Increased mandibular condylar growth in mice with estrogen receptor beta deficiency

Kamiya Yosuke, Chen Jing, Xu Manshan, Utreja Achint, Choi Thomas, Drissi Hicham, Wadhwa Sunil
PMCID: PMC3601565  NIHMSID: NIHMS423703  PMID: 23197372

Abstract

Temporomandibular joint (TMJ) disorders predominantly afflict women of childbearing age, suggesting a role for female hormones in the disease process. In long bones, estrogen acting via estrogen receptor beta (ERβ) inhibits axial skeletal growth in female mice. However, the role of ERβ in the mandibular condyle is largely unknown. We hypothesize that female ERβ deficient mice will have increased mandibular condylar growth compared with wild type (WT) female mice. This study examined female 7-, 49- and 120-day-old WT and ERβ knockout (KO) mice. There was a significant increase in mandibular condylar cartilage thickness, due to an increased number of cells, in the 49- and 120-day-old female ERβ KO compared with WT controls. Analysis in 49-day-old female ERβ KO mice revealed a significant increase in collagen type X, Pthrp and osteoprotegrin gene expression and a significant decrease in Rankl and Ihh gene expression, compared with WT controls. Subchondral bone analysis revealed a significant increase in total condylar volume and a decrease in the number of osteoclasts in the 49-day-old ERβ KO compared with WT female mice. There was no difference in cell proliferation in condylar cartilage between the genotypes. However, there were differences in the expression of proteins that regulate the cell cycle; we found a decrease in the expression of Tieg1 and p57 in the mandibular condylar cartilage from ERβ KO mice compared with WT mice. Taken together, our results suggest that ERβ deficiency increases condylar growth in female mice by inhibiting the fibrocartilage turnover.

Introduction

The National Institute of Dental and Craniofacial Research, a division of the National Institutes of Health, reported that temporomandibular joint (TMJ) disease is the second most common musculoskeletal disease in the United States, with 10.8 million people suffering from TMJ problems at any given time. Eighty percent of individuals seeking treatment for TMJ disorders (TMDs) are females of childbearing age. Because of the high prevalence of TMDs in women of reproductive age, it has been postulated that estrogen may make an individual susceptible to TMD (1-9). Estrogen modulates multiple biological processes within the TMJ region including inflammation (10), matrix metalloproteinase activity (11), and pain modulation (12). However, none of these have been able to adequately explain the gender predilection for TMD, suggesting that other estrogen dependent mechanisms are involved.

Estrogen inhibits the growth of the mandibular condyle. Addition of estrogen to rat mandibular condylar organ cultures results in a decrease in condylar cartilage thickness and proliferation (13). Numerous studies have shown that ovariectomy causes an increase in the thickness of the mandibular condylar cartilage in young rats (14,15), newborn mice (16) and young mice (17); and this is reversed by the administration of exogenous estrogen (15). Not only does lack of estrogen (ovariectomy) increase the condylar cartilage thickness but it also increases the width of the mandibular condylar head, which includes both the condylar cartilage and the subchondral bone, in young mice (17). Furthermore, estrogen negatively regulates condylar cartilage differentiation in vivo. In 4-month-old mice, ovariectomy has been shown to cause an increase in collagen type X expression after 1-8 weeks (18). The ability of estrogen to decrease growth and differentiation of the mandibular condylar cartilage is consistent with the literature on growth plate cartilages; however, estrogen is believed to have a chondro-protective effect on articular cartilages. (see for review (19)). The distinct roles of estrogen in the different cartilages might help explain why the peak incidence of TMD occurs when estrogen levels are the highest; as opposed to the peak incidence of osteoarthritis in other joints, which occurs when estrogen levels are at the lowest (e.g., menopausal women).

The classical estrogen receptors, alpha and beta, have distinct roles in estrogen’s regulation of skeletal growth. ERβ deficient mice have normal estrogen levels (20) and skeletal axial growth is affected only in adult female mice. Specifically, ERβ deficient adult female mice (70-240 days old) have increased axial growth (crown-rump length) compared with the matched wild type mice (20,21). On the other hand, estrogen receptor alpha (ERα) seems to promote skeletal linear growth in male mice but its effect on female mice is unclear (20-22). Since ERβ inhibits skeletal growth exclusively in females, the goal of this study was to examine whether a similar mechanism occurs in the TMJ. We hypothesized that female ERβ deficient mice would have larger mandibular condyles than female WT mice. Greater understanding of estrogen signaling within the TMJ is critical to deciphering the gender predilection of the disease process.

Materials and Methods

Mice

Breeding pairs of C57Bl/6 wild type (Cat # 000664) and ERβ KO mice (homozygous male, heterozygous female, Cat # 004745) were purchased from Jackson Labs. For the studies with ERβ KO mice, only homozygous females were used. Seven- to120-day-old female C57Bl/6 WT (total n=42) and ERβ KO mice (total n=43) were used for the study (see Table 1 for details). At either three or sixteen hours prior to euthanasia, mice were injected intraperitoneally with 0.1 mg bromodeoxyuridine (BrdU) per gram body weight. All experiments were performed in accordance with animal welfare based on an approved IACUC protocol #AAAD0950 from Columbia University animal care committee.

Table 1.

The numbers of mice.

Mouse
Genotype		 Total
Number		 Age
(days)		 Real
Time PCR		 Histology/micro-CT
3 hr BrdUa 16 hr Brdu
Wild Type 42 7 8
49 6 8 7
120 6 7
ERβ KO 43 7 8
49 7 8 8
120 6 6
Open in a new tab
a

these mice were also used for measuring body weight, serum IGF-1 level in 49 day-old mice and condylar cartilage thickness.

RNA Extraction and PCR Amplification

For each mouse the mandibular condylar heads (left and right), containing both the mandibular condylar cartilage and subchondral bone, were carefully harvested and all the soft tissues were removed under a dissecting microscope. Total RNA was obtained from the condylar head and extracted with TRIzol Reagent (Invitrogen life technologies, Carlsbad, CA) following the manufacturer’s protocol. Total RNA was reverse transcribed into cDNA using the ABI High Capacity cDNA Archive Kit (Applied Biosystems, Foster city, CA) following the manufacturer’s protocol. Real-Time PCR was performed, for expression of different genes, in separate wells (singleplex assay) of 96-well plates with reaction volume of 20 μl. Gapdh was used as an endogenous control. Three replicates of each sample were amplified using Assays-on-Demand Gene Expression for the particular gene of interest, with predesigned unlabeled gene-specific PCR primers and TaqMan MGB FAM dye-labeled probes. The PCR reaction mixture (including 2X TaqMan Universal PCR Master Mix, 20X Assays-on-Demand Gene Expression Assay Mix, 50 ng of cDNA) was run in an Applied Biosystems ABI Prism 7300 Sequence Detection System instrument utilizing universal thermal cycling parameters. For the genes in which the efficiencies of target and endogenous control amplification were approximately equal, relative expression in a test sample was compared to a reference calibrator sample (ΔΔCt Method) and used for data analysis. For the genes that were not amplified with the same efficiency as the endogenous control the Relative Standard Curve method, in which target quantity is determined from the standard curve and divided by the target quantity of the calibrator, was used. Gene expression was performed for Proteoglycan 4 (Prg4), Parathyroid hormone related protein (Pthrp), SRY-box containing gene 9 (Sox9), collagen type II (Col2a1), indian hedgehog (Ihh), collagen type X (Col10a1), vascular endothelial growth factor (Vegf), insulin-like growth factor 1 (Igf-1), receptor activator for nuclear factor κ B ligand (Rankl), and osteoprotegrin (Opg).

Histology and immunohistochemistry

Whole mouse heads were sectioned into halves, fixed in 10% formalin for 4 days at room temperature and decalcified in 14% EDTA (pH 7.1) (Sigma, St Louis, MO) for 10 days and 14 days in 49-day and 120-day samples, respectively.. Subsequently, the samples were processed through progressive concentrations of ethanol, cleared in xylene and embedded in paraffin. The TMJ was sagitally serially sectioned into 5μm sections by Microm HM 355s microtome (Thermo Fisher Scientific, Waltham, MA), and every 5th section was stained with H&E.

Histomorphometry measurements were made in a blinded, nonbiased manner using the BioQuant computerized image analysis system (BioQuant, Nashville, TN). Mandibular condylar cartilage thickness measurements were performed on H&E sagittal sections corresponding to the mid-coronal portion of the mandibular condylar head. The entire mandibular condylar cartilage area was divided into 3 layers, superfacial layer (layer S), flattened layer (layer F) and Hypertrophic layer (layer H) based on the cartilage cell size, shape and staining (Figure 2A).The entire anterior-posterior condylar cartilage area was measured for each section and the average of at least three sections was calculated as the cartilage thickness for that mouse. Measurements included cell number, thickness and area of each layer. Six to eight mice from each age and genotype group were analyzed.

Figure 2.

Figure 2

Open in a new tab

H & E staining of the mandibular condylar cartilage (A) from female WT and ERβ KO mice at 49 days and 120 days of age. B, C-Cell counts and thickness measurements were performed for the Superficial layer (S, Articular and Polymorphic zones), Flattened Zone (F) and Upper Hypertrophic zone (H). n=6-8 for each age and genotype, * = significant difference p<0.05. Bars = 100μm.

For immunohistochemistry, tissue sections were deparaffinized with xylene and rehydrated with decreasing concentrations of ethanol. Following rehydration, the sections were treated with 3 % peroxide to block endogenous peroxidase activity and digested for 60 minutes with pepsin for unmasking (Lab Vision, Cat # AP-9007-006, Fremont, CA). All sections were blocked with Protein Block Serum-Free (DakoCytomation, code # X0909, Carpinteria, CA). Immunohistochemical staining was performed using the LSAB + System-HRP Kit (DakoCytomation, code # K0690) following the procedure recommended by the manufacturer. Primary antibodies used in this study were collagen type II antigen (Millipore, MAB8887, 1:200 dilution), Tieg1 (Santa Cruz, sc-23159, 1:100 dilution), p57 (Santa Cruz, sc-8298, 1:200 dilution) and IGF-1 (Abcam, ab40657, 1:100 dilution). BrdU immunohistochemical analysis was completed using a BrdU staining kit following the manufacturer’s instructions (Zymed Laboratories-Invitrogen Corporation, Carlsbad, CA, USA). Negative controls were prepared by omitting the primary antibody step and incubating with blocking solution. To quantify BrdU, Tieg1 and p57 staining, the labeling index (number of BrdU, p57 or Tieg1 positive cells divided by the total number of cells) was calculated. Three to five sections, corresponding to the same anatomical area (mid sagittal), were counted for each animal and the average of these sections was used for the labeling index.

Osteoclastic cell measurement were performed on Tartrate-Resistant Acid Phosphatase (TRAP) stained sections by TRAP kit (Sigma Aldrich, St. Luois, MO). The measurements were made in the bone region beneath the mandibular condylar cartilage. The thickness of the measured region was designated to the approximate cartilage thickness of the correlated region. Osteoclast number, osteoclastic surface, and bone surface in this region were analyzed with BioQuant computerized image analysis system.

micro-CT Analysis

The subchondral bones of the mandibular condyles from C57Bl/6 WT and ERβ KO female mice were analyzed. The three-dimensional morphology of the subchondral bone was evaluated by the micro-CT facility at University of Connecticut Health Center, Farmington CT. The analysis included bone volume, total volume, trabecular number, trabecular spacing, and trabecular thickness.

Serum IGF-1

Mouse serum was collected from 49-day-old WT and ERβ KO female mice. Quantitative assay was performed with IGF1 Mouse ELISA Kit (Abcam, ab100695) following the manufacturer’s protocol. The serum was diluted 1:250 in Assay Diluent A (from the Kit).

Statistical Analyses

Statistical significance of differences among means was determined by one way analysis of variance (ANOVA) with post-hoc comparison of more than two means by the Bonferroni method using GraphPad Prism (San Diego, Ca).

Results

Size measurements

At 49 days of age both WT and ERβ KO female mice weighed the same, but by 120 days of age, ERβ KO mice weighed significantly more than age-matched WT mice (Figure 1A). There was a significant increase in the female ERβ KO mandibular condylar cartilage thickness at 49 and 120 days of age compared with age-matched female WT mice (Figure 1B).

Figure 1.

Figure 1

Open in a new tab

Body weight (A) and mandibular condylar cartilage thickness (B) of female WT and ERβ KO mice at 7, 49 and 120 days of age. n= 6-8 for each age and genotype. * = significant difference p<0.05. C. Serum IGF-1 level of 49-day-old female WT and ERβ KO mice. p=0.052.

Histological analysis

Sagittal sections of the mandibular condylar cartilage from 49- and 120-day-old female ERβ KO and WT mice were stained with H&E (Figure 2A). The mandibular condylar cartilage can be divided into 4 zones: articular, polymorphic, flattened and hypertrophic. In the mandibular condylar cartilage from the 49-day-old ERβ KO mice cells in the polymorphic and flattened zones were vertically stacked, reminiscent of the proliferating zone of the growth plate cartilage. The vertically stacked cells were not apparent in the mandibular condylar cartilage from WT mice of the same age. Due to the difficulty in delineating articular from polymorphic zones in 49- and 120-day-old mice, cell counts were done for the layer S, which includes both the articular and polymorphic zones; the layer F, which includes cells in the flattened zone; and the layer H, which includes hypertrophic chondrocytes not embedded in the subchondral bone. We found significant increases in the total number of cells and in the number of cells in the flattened layer in 49- and 120-day-old ERβ KO mice compared with the WT mice (Figure 2B and 2C). Thickness of the F layer was also significantly increased in 49- and 120-day-old KO mice (Figure 2B and 2C).

Gene expression

Gene expression analysis revealed significant increases in Prg4, Pthrp, Col10a1 and Opg mRNA expression and significant decreases in Sox9, Rankl, Vegf, and Ihh mRNA expression in the 49-day-old ERβ KO compared with age-matched WT mice (Figure 3B). Immunohistochemistry of collagen type 2 expression revealed similar levels of expression between ERβ KO and WT mice (Figure 3A), which was consistent with the gene expression analysis results. However, when gene expression assay was performed with 120-day-old WT and ERβ KO mice, there were no significant differences in any of the genes examined (Figure 3C).

Figure 3.

Figure 3

Open in a new tab

Col2 Immunohistochemistry (A) and Quantification of Gene Expression (B, C) by Q-PCR was performed on the mandibular condyles of female 49- (B) and 120 day-old (C) WT and ERβ KO mice. n=6-8 for each age and genotype, * = significant difference p<0.05. Bars = 100μm.

Cell cycle exit

Proliferation was measured by BrdU immunohistochemistry in the mandibular condylar cartilage from female WT and ERβ KO mice injected with BrdU three hours prior to sacrifice. We found no significant difference in the number of BrdU labeled cells/ total number of cells in the mandibular condylar cartilage from 49 – (Figure 3B) and 120 day-old (data not shown) female ERβ KO compared to aged matched female WT mice. In order to determine if the increase in cell numbers in the ERβ KO mice was due to a delay in the number of cells exiting the proliferative pool, we examined markers of cell cycle exit. Immunohistochemistry for BrdU was performed on the mandibular condylar cartilage of mice that received BrdU either 3 hours or 16 hours prior to sacrifice. There was a significant decrease in the percentage of BrdU labeled cells after 16 hours compared with 3 hours in WT mice; but, no significant difference was evident in the ERβ KO mice (Figure 4). This decrease in BrdU labeling represents cells that have exited the proliferative pool of the mandibular condylar cartilage (23). Immunohistochemistry was also performed for two regulators of cell cycle exit, Tieg1 and p57. Both were predominantly localized to the area of transition between flattened and hypertrophic zones in the mandibular condylar cartilage. We also found that there was a significant decrease in p57 and Tieg1 in the female ERβ KO compared with age-matched WT mice (Figure 5).

Figure 4.

Figure 4

Open in a new tab

Immunohistochemistry for BrdU in the mandibular condyle was performed on female 49-day-old WT and ERβ KO mice (A). Mice were injected with BrdU at either 3 hours or 16 hours prior to sacrifice. B. Quantification of BrdU labeling. n=6-8 for each age and genotype, * = significant difference p<0.05. Bars = 100μm.

Figure 5.

Figure 5

Open in a new tab

A. Immunohistochemistry for Tieg1 and p57 was performed on the mandibular condyles of female 49-day-old WT and ERβ KO mice. B. Quantification of Tieg1 and p57 labeling. n=6-8 for each age and genotype, * = significant difference p<0.05. Bars = 100μm.

Local and serum IGF-1

ELISA assay showed a tendency toward increased serum IGF-1 levels in 49-day-old ERβ KO mice compared with WT. However, the difference is not statistically significant (Figure 1C, p=0.052). Real-time PCR detected no significant difference in condylar head mRNA levels (Figure 3C) between wild type and ERβ KO mice, in either the 49- or 120-day-old groups. Similarly, immunohistochemistry showed no difference in the IGF-1 protein distribution pattern in the condylar cartilage of wild type and ERβ KO female mice (data not shown).

Subchondral bone

Micro-Ct analysis revealed a significant increase in the total volume and a significant decrease in the bone density (BV/TV) and trabecular thickness in 49-day-old ERβ KO compared with age-matched WT mice (Figure 6). Micro-CT of 120-day-old WT and ERβ KO mice revealed no significant differences in any of the measurements. TRAP staining was performed to quantitate the number of osteoclasts and the ratio of osteoclast surface/bone surface (OcS/BS) in 49- and 120-day-old groups. We found that in 49-day-old mice, there were significant decreases in both the number of osteoclasts and OcS/BS in the subchondral bone of ERβ KO mice compared with age-matched WT mice (Figure 7).

Figure 6.

Figure 6

Open in a new tab

micro-Ct analysis of the subchondral bone from female WT and ERβ KO at 49 and 120 days of age on: Total Volume, Bone Volume, Bone Volume Fraction (BV/TV), Trabecular Thickness, Trabecular Number, and Trabecular Spacing. n=6-8 for each age and genotype, * = significant difference p<0.05.

Figure 7.

Figure 7

Open in a new tab

A. TRAP staining of the mandibular condyles of female 49 day- and 120-day-old WT and ERβ KO mice. B. Quantification of osteoclast surface/bone surface (OcS/BS). C. Quantification of osteoclast number, n=6-8 for each age and genotype, * = significant difference p<0.05. Bars = 200μm.

Discussion

The mandibular condylar cartilage is unique compared with other cartilages in the body in a number of ways. For example, other articular cartilages are composed of hyaline cartilage, whereas the mandibular condylar cartilage is composed of fibrocartilage (reviewed in (24)) displaying distinct layers of cells at various stages of differentiation. Furthermore, the mandibular condylar cartilage undergoes endochondral ossification and is derived from periosteum (25). Interestingly, estrogen acting via the ERβ pathway has been shown to inhibit endochondral ossification of the femur (26), mechanical load-induced periosteal bone formation (27) and fibrocartilage maturation of the fracture callus (28).

In our study we found that the female ERβKO mice had increased mandibular condylar cartilage thickness and an increased number of cells in the flattened zone at 49 and 120 days of age compared with age-matched female WT mice. We believe that this increase may be due to a delay in cell cycle exit in the cells of the flattened zone. In support of this claim, we found a significant decrease in the number of BrdU labeled cells in female WT mice that were injected with BrdU 16 hours prior to sacrifice when compared with those injected with BrdU 3 hours prior to sacrifice. This reduction in BrdU labeling signifies cells have exited the flattened zone and undergone hypertrophic maturation. In contrast, ERβ KO female mice exhibited no significant differences in BrdU labeling when BrdU was injected 16 hours prior to sacrifice as compared to 3 hours prior to sacrifice. A significant decrease in the expression of the cell cycle regulators, p57 and Tieg1, was found in the mandibular condyle of female ERβ KO compared with female WT mice. In growth plate cartilage, p57 has been shown to inhibit cyclin-dependent kinases, and its activation is associated with cells exiting the proliferative pool and undergoing hypertrophic maturation (29,30). Tieg1 has been shown to act downstream of estrogen receptor beta signaling (31). Furthermore, in tenocytes (32) and myoblasts (33) Tieg1 deficiency caused a delay in cell cycle exit.

Another possible explanation for the increased mandibular condylar cartilage thickness is that ERβ KO mice have been shown to have increased serum IGF-1 levels(20), and exogenous IGF-1 treatment causes increased mandibular condylar cartilage thickness (34). However, we do not believe this is the major mechanism involved as we did not find any significant changes in serum IGF-1 levels or in the local expression of IGF-1 mRNA within the mandibular condylar cartilage of female ERβ KO compared with wild-type mice. Additionally, IGF-1 treatment was recently shown to cause increased mandibular condylar cartilage proliferation (35), which was not part of our ERβKO mandibular condylar cartilage phenotype.

A more pronounced phenotype was present in the mandibular condylar cartilage of ERβ KO female mice at 49 days of age than at 120 days of age. For example, 49-day-old female ERβ KO mice presented with decreased osteoclast numbers, increased chondrocyte maturation markers and a larger mandibular condylar total volume compared with the age-matched WT mice. At 2-4 months of age, the long bones of ERβKO female mice exhibit an increased femur length compared with age-matched WT mice; this difference is not evident in 8-month-old mice (21,26). Unlike the femur, the majority of growth of the mandibular condyle occurs before puberty (36-38), which may explain the more pronounced female ERβ KO mandibular condylar phenotype observed at 49 days of age.

There were decreased osteoclast numbers in the subchondral bone of the mandibular condylar cartilage in female ERβ KO mice, which is consistent to what has been reported in ERβ KO long bones(39). The decrease in the number of osteoclasts could be due to either decreased RANKL expression in the hypertrophic chondrocytes of the mandibular condylar cartilage (40) or decreased Tieg1 expression, which has been shown to repress OPG promoter activity in female ERβ KO mice (41). We speculate that the increase in Col X expression in the mandibular condylar cartilage of ERβ KO mice is due to a decrease in osteoclasts (as seen in Figure 7). In support, accumulation of hypertrophic chondrocytes in the mandibular condyle has been reported in models of defective osteoclasts (42).

Decreased levels of estrogen by ovariectomy resulted in increased mandibular condylar cartilage thickness and increased osteoclast numbers compared with female sham operated controls (16,43). The fact that we see decreased osteoclast numbers in the mandibular condyles of the ERβ KO mice, suggests that other estrogen mediated pathways are involved. For example, estrogen receptor alpha may be also involved; estrogen through the estrogen receptor alpha has been shown to cause increased osteoclast apoptosis (44).

In this study we show that deletion of estrogen receptor beta reduces osteoclast surface/ bone surface and delays the number of cells exiting the flattened zone, resulting in a larger mandibular condyle at 49 days of age. In the growth plate cartilage, there is believed to be a finite number of cells capable of proliferation (45). We have previously shown that mechanical loading also causes increased mandibular condylar cartilage proliferation (46). Therefore, it is possible that high estrogen levels during puberty may deplete the number of proliferative cells in the mandibular condylar cartilage, making post-pubertal females less responsive to mechanical load-induced TMJ remodeling. A similar phenomenon occurs in periosteal bones, where pre-pubertal females exhibit a greater increase in load-induced periosteal bone formation than post-pubertal females (47). In our future studies we plan to investigate the effects of estrogen receptor beta deficiency in decreased versus increased TMJ loading models.

ACKNOWLEDGEMENTS

This work was supported by 1R01DE020097 (SW, HD) from the NIH. We thank Dr. Ilona Polur (Columbia University College of Dental Medicine) for proofreading the manuscript.

Authors’ roles: Study design: HD and SW. Study conduct, data collection, and analysis: YK, JC, AU, MX, and TC. Data interpretation: JC, HD and SW. Drafting manuscript: JC, HD and SW. Revising manuscript content: JC, HD and SW. YK, JC, MX, and SW take responsibility for the integrity of the data analysis.

References

  • 1.Abubaker AO, Raslan WF, Sotereanos GC. Estrogen and progesterone receptors in temporomandibular joint discs of symptomatic and asymptomatic persons: a preliminary study. J Oral Maxillofac Surg. 1993 Oct;51(10):1096–100. doi: 10.1016/s0278-2391(10)80448-3. [DOI] [PubMed] [Google Scholar]
  • 2.Aufdemorte TB, Van Sickels JE, Dolwick MF, Sheridan PJ, Holt GR, Aragon SB, Gates GA. Estrogen receptors in the temporomandibular joint of the baboon (Papio cynocephalus): an autoradiographic study. Oral Surg Oral Med Oral Pathol. 1986 Apr;61(4):307–14. doi: 10.1016/0030-4220(86)90407-x. [DOI] [PubMed] [Google Scholar]
  • 3.Milam SB, Aufdemorte TB, Sheridan PJ, Triplett RG, Van Sickels JE, Holt GR. Sexual dimorphism in the distribution of estrogen receptors in the temporomandibular joint complex of the baboon. Oral Surg Oral Med Oral Pathol. 1987 Nov;64(5):527–32. doi: 10.1016/0030-4220(87)90025-9. [DOI] [PubMed] [Google Scholar]
  • 4.Yamada K, Nozawa-Inoue K, Kawano Y, Kohno S, Amizuka N, Iwanaga T, Maeda T. Expression of estrogen receptor alpha (ER alpha) in the rat temporomandibular joint. Anat Rec A Discov Mol Cell Evol Biol. 2003 Oct;274(2):934–41. doi: 10.1002/ar.a.10107. [DOI] [PubMed] [Google Scholar]
  • 5.Wang YH, Liu Y, Maye P, Rowe DW. Examination of mineralized nodule formation in living osteoblastic cultures using fluorescent dyes. Biotechnol Prog. 2006 Nov-Dec;22(6):1697–701. doi: 10.1021/bp060274b. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.LeResche L, Saunders K, Von Korff MR, Barlow W, Dworkin SF. Use of exogenous hormones and risk of temporomandibular disorder pain. Pain. 1997 Jan;69(1-2):153–60. doi: 10.1016/s0304-3959(96)03230-7. [DOI] [PubMed] [Google Scholar]
  • 7.Meisler JG. Chronic pain conditions in women. J Womens Health. 1999 Apr;8(3):313–20. doi: 10.1089/jwh.1999.8.313. [DOI] [PubMed] [Google Scholar]
  • 8.Landi N, Lombardi I, Manfredini D, Casarosa E, Biondi K, Gabbanini M, Bosco M. Sexual hormone serum levels and temporomandibular disorders. A preliminary study. Gynecol Endocrinol. 2005 Feb;20(2):99–103. doi: 10.1080/09513590400021136. [DOI] [PubMed] [Google Scholar]
  • 9.Kang SC, Lee DG, Choi JH, Kim ST, Kim YK, Ahn HJ. Association between estrogen receptor polymorphism and pain susceptibility in female temporomandibular joint osteoarthritis patients. Int J Oral Maxillofac Surg. 2007 May;36(5):391–4. doi: 10.1016/j.ijom.2006.12.004. [DOI] [PubMed] [Google Scholar]
  • 10.Galal N, El Beialy W, Deyama Y, Yoshimura Y, Yoshikawa T, Suzuki K, Totsuka Y. Effect of estrogen on bone resorption and inflammation in the temporomandibular joint cellular elements. Int J Mol Med. 2008 Jun;21(6):785–90. [PubMed] [Google Scholar]
  • 11.Wadhwa S, Kapila S. TMJ disorders: future innovations in diagnostics and therapeutics. J Dent Educ. 2008 Aug;72(8):930–47. [PMC free article] [PubMed] [Google Scholar]
  • 12.Craft RM. Modulation of pain by estrogens. Pain. 2007 Nov;132(Suppl 1):S3–12. doi: 10.1016/j.pain.2007.09.028. [DOI] [PubMed] [Google Scholar]
  • 13.Talwar RM, Wong BS, Svoboda K, Harper RP. Effects of estrogen on chondrocyte proliferation and collagen synthesis in skeletally mature articular cartilage. J Oral Maxillofac Surg. 2006 Apr;64(4):600–9. doi: 10.1016/j.joms.2005.12.006. [DOI] [PubMed] [Google Scholar]
  • 14.Okuda T, Yasuoka T, Nakashima M, Oka N. The effect of ovariectomy on the temporomandibular joints of growing rats. J Oral Maxillofac Surg. 1996 Oct;54(10):1201–10. doi: 10.1016/s0278-2391(96)90352-3. discussion 1210-1. [DOI] [PubMed] [Google Scholar]
  • 15.Yasuoka T, Nakashima M, Okuda T, Tatematsu N. Effect of estrogen replacement on temporomandibular joint remodeling in ovariectomized rats. J Oral Maxillofac Surg. 2000 Feb;58(2):189–96. doi: 10.1016/s0278-2391(00)90337-9. discussion 196-7. [DOI] [PubMed] [Google Scholar]
  • 16.Fujita T, Ohtani J, Shigekawa M, Kawata T, Kaku M, Kohno S, Motokawa M, Tohma Y, Tanne K. Influence of sex hormone disturbances on the internal structure of the mandible in newborn mice. Eur J Orthod. 2006 Apr;28(2):190–4. doi: 10.1093/ejo/cji093. [DOI] [PubMed] [Google Scholar]
  • 17.Fujita T, Kawata T, Tokimasa C, Kohno S, Kaku M, Tanne K. Breadth of the mandibular condyle affected by disturbances of the sex hormones in ovariectomized and orchiectomized mice. Clin Orthod Res. 2001 Aug;4(3):172–6. doi: 10.1034/j.1600-0544.2001.040307.x. [DOI] [PubMed] [Google Scholar]
  • 18.Min HJ, Lee MJ, Kim JY, Cho SW, Park HD, Lee SI, Kim HJ, Jung HS. Alteration of BMP-4 and Runx2 expression patterns in mouse temporomandibular joint after ovariectomy. Oral Dis. 2007 Mar;13(2):220–7. doi: 10.1111/j.1601-0825.2006.01270.x. [DOI] [PubMed] [Google Scholar]
  • 19.Tanko LB, Sondergaard BC, Oestergaard S, Karsdal MA, Christiansen C. An update review of cellular mechanisms conferring the indirect and direct effects of estrogen on articular cartilage. Climacteric. 2008 Feb;11(1):4–16. doi: 10.1080/13697130701857639. [DOI] [PubMed] [Google Scholar]
  • 20.Lindberg MK, Alatalo SL, Halleen JM, Mohan S, Gustafsson JA, Ohlsson C. Estrogen receptor specificity in the regulation of the skeleton in female mice. J Endocrinol. 2001 Nov;171(2):229–36. doi: 10.1677/joe.0.1710229. [DOI] [PubMed] [Google Scholar]
  • 21.Chagin AS, Lindberg MK, Andersson N, Moverare S, Gustafsson JA, Savendahl L, Ohlsson C. Estrogen receptor-beta inhibits skeletal growth and has the capacity to mediate growth plate fusion in female mice. J Bone Miner Res. 2004 Jan;19(1):72–7. doi: 10.1359/JBMR.0301203. [DOI] [PubMed] [Google Scholar]
  • 22.Vidal O, Lindberg MK, Hollberg K, Baylink DJ, Andersson G, Lubahn DB, Mohan S, Gustafsson JA, Ohlsson C. Estrogen receptor specificity in the regulation of skeletal growth and maturation in male mice. Proc Natl Acad Sci U S A. 2000 May;97(10):5474–9. doi: 10.1073/pnas.97.10.5474. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Rabie AB, Tang GH, Xiong H, Hagg U. PTHrP regulates chondrocyte maturation in condylar cartilage. J Dent Res. 2003 Aug;82(8):627–31. doi: 10.1177/154405910308200811. [DOI] [PubMed] [Google Scholar]
  • 24.Benjamin M, Ralphs JR. Biology of fibrocartilage cells. Int Rev Cytol. 2004;233:1–45. doi: 10.1016/S0074-7696(04)33001-9. [DOI] [PubMed] [Google Scholar]
  • 25.Meikle MC. Remodeling the dentofacial skeleton: the biological basis of orthodontics and dentofacial orthopedics. J Dent Res. 2007 Jan;86(1):12–24. doi: 10.1177/154405910708600103. [DOI] [PubMed] [Google Scholar]
  • 26.Windahl SH, Vidal O, Andersson G, Gustafsson JA, Ohlsson C. Increased cortical bone mineral content but unchanged trabecular bone mineral density in female ERbeta(-/-) mice. J Clin Invest. 1999 Oct;104(7):895–901. doi: 10.1172/JCI6730. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 27.Saxon LK, Robling AG, Castillo AB, Mohan S, Turner CH. The skeletal responsiveness to mechanical loading is enhanced in mice with a null mutation in estrogen receptor-beta. Am J Physiol Endocrinol Metab. 2007 Aug;293(2):E484–91. doi: 10.1152/ajpendo.00189.2007. [DOI] [PubMed] [Google Scholar]
  • 28.He YX, Liu Z, Pan XH, Tang T, Guo BS, Zheng LZ, Xie XH, Wang XL, Lee KM, Li G, Cao YP, Wei L, Chen Y, Yang ZJ, Hung LK, Qin L, Zhang G. Deletion of estrogen receptor beta accelerates early stage of bone healing in a mouse osteotomy model. Osteoporos Int. 2012 Jan;23(1):377–89. doi: 10.1007/s00198-011-1812-x. [DOI] [PubMed] [Google Scholar]
  • 29.Kronenberg HM. PTHrP and skeletal development. Ann N Y Acad Sci. 2006 Apr;1068:1–13. doi: 10.1196/annals.1346.002. [DOI] [PubMed] [Google Scholar]
  • 30.Zhang P, Liegeois NJ, Wong C, Finegold M, Hou H, Thompson JC, Silverman A, Harper JW, DePinho RA, Elledge SJ. Altered cell differentiation and proliferation in mice lacking p57KIP2 indicates a role in Beckwith-Wiedemann syndrome. Nature. 1997 May;387(6629):151–8. doi: 10.1038/387151a0. [DOI] [PubMed] [Google Scholar]
  • 31.Hawse JR, Subramaniam M, Monroe DG, Hemmingsen AH, Ingle JN, Khosla S, Oursler MJ, Spelsberg TC. Estrogen receptor beta isoform-specific induction of transforming growth factor beta-inducible early gene-1 in human osteoblast cells: an essential role for the activation function 1 domain. Mol Endocrinol. 2008 Jul;22(7):1579–95. doi: 10.1210/me.2007-0253. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 32.Haddad O, Gumez L, Hawse JR, Subramaniam M, Spelsberg TC, Bensamoun SF. TIEG1-null tenocytes display age-dependent differences in their gene expression, adhesion, spreading and proliferation properties. Exp Cell Res. 2011 Jul;317(12):1726–35. doi: 10.1016/j.yexcr.2011.05.007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33.Miyake M, Hayashi S, Iwasaki S, Uchida T, Watanabe K, Ohwada S, Aso H, Yamaguchi T. TIEG1 negatively controls the myoblast pool indispensable for fusion during myogenic differentiation of C2C12 cells. J Cell Physiol. 2011 Apr;226(4):1128–36. doi: 10.1002/jcp.22434. [DOI] [PubMed] [Google Scholar]
  • 34.Suzuki S, Itoh K, Ohyama K. Local administration of IGF-I stimulates the growth of mandibular condyle in mature rats. J Orthod. 2004 Jun;31(2):138–43. doi: 10.1179/146531204225020436. [DOI] [PubMed] [Google Scholar]
  • 35.Chen Y, Ke J, Long X, Meng Q, Deng M, Fang W, Li J, Cai H, Chen S. Insulin-like growth factor-1 boosts the developing process of condylar hyperplasia by stimulating chondrocytes proliferation. Osteoarthritis Cartilage. 2012 Apr;20(4):279–87. doi: 10.1016/j.joca.2011.12.013. [DOI] [PubMed] [Google Scholar]
  • 36.Festing M. A multivariate analysis of subline divergence in the shape of the mandible in C57BL-Gr mice. Genet Res. 1973 Apr;21(2):121–32. doi: 10.1017/s0016672300013306. [DOI] [PubMed] [Google Scholar]
  • 37.Lovell DP, Totman P, Johnson FM. Variation in the shape of the mouse mandible. 1. Effect of age and sex on the results obtained from the discriminant functions used for genetic monitoring. Genet Res. 1984 Feb;43(1):65–73. doi: 10.1017/s0016672300025726. [DOI] [PubMed] [Google Scholar]
  • 38.Chen J, Sorensen KP, Gupta T, Kilts T, Young M, Wadhwa S. Altered functional loading causes differential effects in the subchondral bone and condylar cartilage in the temporomandibular joint from young mice. Osteoarthritis Cartilage. 2009 Mar;17(3):354–61. doi: 10.1016/j.joca.2008.05.021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Sims NA, Dupont S, Krust A, Clement-Lacroix P, Minet D, Resche-Rigon M, Gaillard-Kelly M, Baron R. Deletion of estrogen receptors reveals a regulatory role for estrogen receptors-beta in bone remodeling in females but not in males. Bone. 2002 Jan;30(1):18–25. doi: 10.1016/s8756-3282(01)00643-3. [DOI] [PubMed] [Google Scholar]
  • 40.Xiong J, Onal M, Jilka RL, Weinstein RS, Manolagas SC, O’Brien CA. Matrix-embedded cells control osteoclast formation. Nat Med. 2011 Sep;17(10):1235–41. doi: 10.1038/nm.2448. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 41.Subramaniam M, Hawse JR, Bruinsma ES, Grygo SB, Cicek M, Oursler MJ, Spelsberg TC. TGFbeta inducible early gene-1 directly binds to, and represses, the OPG promoter in osteoblasts. Biochem Biophys Res Commun. 2010 Jan;392(1):72–6. doi: 10.1016/j.bbrc.2009.12.171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 42.Shibata S, Oda T, Abe T, Yamashita Y, Takano Y. Structural features of incremental line-like striations in mandibular condylar cartilage of c-src-deficient mice. Arch Oral Biol. 2006 Nov;51(11):951–9. doi: 10.1016/j.archoralbio.2006.05.009. [DOI] [PubMed] [Google Scholar]
  • 43.Fujita T, Kawata T, Tokimasa C, Kaku M, Kawasoko S, Tanne K. Influences of ovariectomy and orchiectomy on the remodeling of mandibular condyle in mice. J Craniofac Genet Dev Biol. 1998 Jul-Sep;18(3):164–70. [PubMed] [Google Scholar]
  • 44.Nakamura T, Imai Y, Matsumoto T, Sato S, Takeuchi K, Igarashi K, Harada Y, Azuma Y, Krust A, Yamamoto Y, Nishina H, Takeda S, Takayanagi H, Metzger D, Kanno J, Takaoka K, Martin TJ, Chambon P, Kato S. Estrogen prevents bone loss via estrogen receptor alpha and induction of Fas ligand in osteoclasts. Cell. 2007 Sep;130(5):811–23. doi: 10.1016/j.cell.2007.07.025. [DOI] [PubMed] [Google Scholar]
  • 45.Schrier L, Ferns SP, Barnes KM, Emons JA, Newman EI, Nilsson O, Baron J. Depletion of resting zone chondrocytes during growth plate senescence. J Endocrinol. 2006 Apr;189(1):27–36. doi: 10.1677/joe.1.06489. [DOI] [PubMed] [Google Scholar]
  • 46.Sobue T, Yeh WC, Chhibber A, Utreja A, Diaz-Doran V, Adams D, Kalajzic Z, Chen J, Wadhwa S. Murine TMJ loading causes increased proliferation and chondrocyte maturation. J Dent Res. 2011 Apr;90(4):512–6. doi: 10.1177/0022034510390810. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 47.Daly RM. The effect of exercise on bone mass and structural geometry during growth. Med Sport Sci. 2007;51:33–49. doi: 10.1159/000103003. [DOI] [PubMed] [Google Scholar]

RESOURCES