Elsevier

Archives of Oral Biology

Volume 57, Issue 10, October 2012, Pages 1377-1384
Archives of Oral Biology

Osteoblasts stimulate osteoclastogenesis via RANKL expression more strongly than periodontal ligament cells do in response to PGE2

https://doi.org/10.1016/j.archoralbio.2012.07.009Get rights and content

Abstract

Objective

Periodontal ligament cells (PDLs) produce prostaglandin E2 (PGE2) in response to orthodontic force. PGE2 is a potent osteoclast-inducing factor that induces the receptor activator of nuclear factor-κB ligand (RANKL). Some studies reported that PDLs express RANKL in response to mechanical stress, whereas another study reported that they do not. Based on an immunohistochemical study, RANKL expression is localized around the alveolar bone surface 3 days after tooth movement. However, ankylosed teeth cannot be moved by therapeutic mechanical stress, suggesting that PDLs play a major role in alveolar bone resorption. In this study, we compared the functional difference in osteoclastogenesis between human PDLs (HPDLs) and normal human osteoblasts (HOBs) as a direct effect of PGE2 exposure.

Design

We examined the expression of RANKL, osteoprotegerin, and macrophage colony-stimulating factor after 48-h culture with or without PGE2 (10−11 to 10−5 M) in HPDLs and HOBs. Then to confirm whether RANKL produced by PGE2 treatment induces osteoclastogenesis or not, RAW264.7 cells were co-cultured on HPDLs or HOBs pretreated with 10−6 M of PGE2.

Result

PGE2 exposure increased significantly RANKL expression in HOBs compared with HPDLs. PGE2 exposure significantly decreased osteoprotegerin expression in HPDLs compared with HOBs. The number of tartrate-resistant acid phosphatase staining osteoclast-like cells from RAW264.7 cells increased significantly by PGE2 pretreatment in HOBs and was reduced by small interfering RNA knockdown of RANKL.

Conclusion

These results suggest that osteoblasts strongly influence the stimulation of osteoclastogenesis via RANKL, induced by PGE2 in periodontal tissues, compared with PDLs.

Introduction

Orthodontic tooth movement occurs during sequential bone remodelling induced by therapeutic mechanical stress.1 Mechanical stress induces bone resorption on the compression side and bone formation on the tension side.2, 3, 4 On the compression side, osteoclast formation and differentiation are regulated by the balance among the receptor activator of nuclear factor-κB ligand (RANKL), osteoprotegerin (OPG), and macrophage colony-stimulating factor (M-CSF).5, 6 Many studies have shown that bone resorption via osteoclasts is mediated by prostaglandins, especially prostaglandin E2 (PGE2),7, 8 which is produced by periodontal ligament cells (PDLs) in response to mechanical stress in vivo9 and in vitro.7, 10, 11 PGE2 also stimulates RANKL expression.12, 13 Some studies reported that PDLs express RANKL in response to mechanical stress,7, 14 whereas another study reported that they do not.15 However, ankylosed teeth cannot be moved by therapeutic mechanical stress, suggesting that PDLs play a major role in alveolar bone resorption. Based on an immunohistochemical study, RANKL is expressed consistently around the alveolar bone surface 3 days after tooth movement.16 Consequently, PGE2 produced in response to mechanical stress in PDLs may stimulate RANKL expression directly in PDLs and in surrounding cells, such as osteoblasts in alveolar bone.

In this study, we compared the functional difference in osteoclastogenesis between human PDLs (HPDLs) and normal human osteoblasts (HOBs) on the application of PGE2 produced by HPDLs in response to mechanical stress.7, 10

Section snippets

Cell cultures

The protocol for this experiment was reviewed and approved by the Nihon University Department of Dentistry Ethics Committee. HPDLs were obtained from the healthy premolars of 14 patients aged 11–50 years in the course of orthodontic treatment. HPDLs were isolated and cultured according to Somerman et al.17 The cells were cultured in α-minimum essential medium (α-MEM) supplemented with 10% foetal bovine serum (FBS), 100 U/mL penicillin-G sodium, 100 μg/mL streptomycin sulfate, and 0.25 μg/mL

Effect of compressive force on RANKL and COX-2 mRNA expression

RANKL and COX-2 expression in HPDLs in response to 24 h of compressive force (comp) are shown in Fig. 1A. Although COX-2 expression increased significantly in response to the compressive force in all 14 HPDLs compared with the non-compression controls, RANKL expression was almost undetected. Marked RANKL expression was observed in HPDLs 2, 12, and 13, but it did not depend on the compressive force (Fig. 1B).

Effect of PGE2 on RANKL, OPG, and M-CSF mRNA expression

RANKL expression in HPDLs and HOBs after a 48-h culture with 0–10−5 M PGE2 is shown in

Discussion

PGE2 is a potent osteoclast-inducing factor20, 21, 22 that is produced in PDLs in response to mechanical stress in vivo9 and in vitro.7, 10, 11 PGE2 induces RANKL expression in osteoblasts5 and HPDLs.7 RANKL has been identified as a key cytokine that regulates osteoclastogenesis and bone resorption.23 During tooth movement, alveolar bone resorption by osteoclasts is regulated by the balance among RANKL, OPG, and M-CSF.24

Some studies have reported that PDLs express RANKL in response to

Funding

Grant-in-Aid for Scientific Research (c) (22592298) and the “Strategic Research Base Development” Program for Private Universities (S1001024) from the Ministry of Education, Science, Culture, Sports, Science and Technology. A Grant from the Dental Research Centre and Sato Fund, Nihon University School of Dentistry.

Competing interests

None declared.

Ethical approval statement

The protocol for this experiment was reviewed and approved by the Nihon University Department of Dentistry and Ethics committee (Ref. no. 2).

Acknowledgments

This study was supported by a Grant-in-Aid for Scientific Research (c) (22592298) and the “Strategic Research Base Development” Program for Private Universities (S1001024) from the Ministry of Education, Science, Culture, Sports, Science and Technology, a Grant from the Dental Research Centre and Sato Fund, Nihon University School of Dentistry, and The Promotion and Mutual Aid Corporation for Private Schools of Japan.

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