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

https://doi.org/10.1016/j.archoralbio.2009.01.018Get rights and content

Abstract

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, ERα and ERβ 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 ERα were observed in hypertrophic and mature layers, while ERβ 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 ERα expression and the 4- and 8-week-old female rats showed a higher ERβ expression than the age-matched male ones in western blot results (P < 0.05).

Conclusions

The results that oestradiol, ERα and ERβ 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.

Introduction

Osteoarthritis (OA) is a degenerative joint disease caused by the degradation of articular cartilage. Most epidemiological studies have shown that OA occurs in a larger number of females than males, and the rates of both prevalence and incidence increase in postmenopausal women. Furthermore, the symptoms of OA are more severe in women than in men.[1], [2], [3], [4] The marked sexual dimorphism attracted investigators to focus on the female reproductive hormones in OA. The temporomandibular joint (TMJ) plays an important role in craniofacial growth and function, and shows a high incidence of OA.5 In literature, the effect of oestrogen on mandibular condylar cartilage has been studied by several researchers. In TMJs from ovariectomized animals the increased condylar cartilage thickness and even degenerative changes were noticed.[6], [7], [8], [9] Studies on the effect of different concentrations of exogenous oestrogen on cultured mandibular condylar cartilage blocks or chondrocytes also suggested that oestrogen played an important role in modulating the morphology and function of condylar cartilages.[10], [11] Additional support was provided by the existence of oestrogen receptors (ERs) in mandibular condylar cartilages reported by several studies. As a nuclear hormone receptor, ER is one member of a family of activated transcription factors that can initiate or enhance the transcription of genes containing specific oestrogen response elements.12 The two subtypes of ERs, ERα and ERβ, are distinct proteins encoded by separate genes located on different chromosomes. As early as middle of 1980s, Sherian et al.13 and Aufdermorte et al.14 found ERs in the mandibular condylar cartilage of female baboons, although Milam SB et al.15 failed to detect ER in condylar cartilages of male baboons by autoradiography. With the methods of immunohistochemistry and in situ hybridization Yamada et al.16 revealed that ERα is distributed in male rat mandibular condylar cartilage. It seemed that oestrogens have the potential to modify the development and function of mandibular condylar cartilage.

It is suggested that oestrogen can be produced not only by gonads but also by a number of extragonadal sites, including cartilage and bone, in which oestrogen can act locally in a paracrine or intracrine fashion.17 In our previous report,18 it was found that mandibular condylar chondrocytes expressed aromatase which is essential in oestrogen synthesis, and oestrogen may be synthesized locally through the presence of aromatase. However, in literature very few reports were found focusing on the local expression of oestrogen in mandibular condylar cartilages in vivo. Whether there is difference in the expression of oestrogen, as well as its receptors, in mandibular condylar cartilages between sexes and between those at different ages is worth investigating for a future exploration of the role of oestrogen in the onset of OA. Thus the aim of the present study was to compare the expression of oestrogen, ERα and ERβ in mandibular condylar cartilages of Sprague–Dawley (SD) rats in different age and sex groups.

Section snippets

Animals and tissue preparation

The study was approved by the Animal Research Committee of the Fourth Military Medical University. One hundred SD rats, at the age of 2, 4, 8 weeks and 4, 12 months in both sexes, 10 in each age and sex group, were provided by the animal centre of the Fourth Military Medical University.

Under deep anaesthesia with intraperitoneal injection of pentobarbital sodium (50 mg/kg body weight), 50 rats, 5 in each age and sex group for immunohistochemistry, were perfused with 200 mL normal saline through

Immunolocalization of oestradiol and ERs

No immunoreactivity was observed in the controls in which the primary antibodies were omitted. Immunocytochemistry for oestradiol and ERs demonstrated intense immunoreaction in rat mandibular condylar cartilage (Fig. 1a–c). Oestradiol and ERα were expressed in the hypertrophic and mature layers, abundant in the hypertrophic layer, while ERβ only in hypertrophic layer (Fig. 1d–f). Oestradiol was primarily localized to cytoplasms of chondrocytes (Fig. 1g), ERβ primarily to nuclei (Fig. 1i) but

Discussion

With the use of immunohistochemistry, ELISA and western blot in the present study, the local expressions of oestradiol, ERα and ERβ were detected in rat mandibular condylar cartilages. Some difference was found in age or sex distribution between the results of immunohistochemistry and western blot or ELISA. That is assumed mainly due to different detection method. The immunohistochemistry method provides information for the density of oestradiol or ERs positive chondrocytes. But the western

Acknowledgments

The authors thank Dr. Wang Tao and Dr. Dai Juan for technical assistance. The present study was supported by the National Nature Science Foundation of China (NSFC) No. 30471909 and No. 30772429.

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