Spring 2007

Scientific Article

Genomic Elements in
Orthodontic Tooth Movement
Philippe Sebrechts and Ana Paula Ferraz

Introduction

Bone remodeling in healthy people is a dynamic process in which bone mass homeostasis is maintained throughout life. Two constantly functioning simultaneous events define bone remodeling: osteoclast-mediated bone resorption and bone deposition by osteoblasts.[1, 2] Orthodontic Tooth Movement (OTM) disrupts homeostasis. Osteoblasts and osteoclasts are sensitive environment-to genome-to-environment communicators capable of restoring equilibrium. In response to applied force, the body has a highly sophisticated mechanism involving hundreds of genes and thousands of proteins which convert mechanical force to biological events.[3]

Application of orthodontic force for even one day enhances osteoclast attachment to and resorption of alveolar bone.[4] Bone cell proliferation, differentiation, function, and apoptosis are carefully regulated by a wide variety of biologic mediators (hormones, cytokines, and growth factors) to influence the homeostasis of bone tissue.[7, 8] Growth Factors (GFs) are proteins that stimulate cell proliferation and differentiation; they function by binding to specific receptors on target cells. GFs trigger autophosphorylation of tyrosine kinase. Once phosphorylated, the enzyme has a high affinity for proteins that initiate a cascade of reactions associated with cell transformation.[1] Cytokines and growth factors such as bone morphogenic proteins (BMPs), fibroblast growth factor (FGF), platelet-derived growth factor (PDGF), insulin-like growth factor, and transforming growth factor-ß (TGF-ß) regulate bone cells. [7, 8] The bone-specific transcriptional regulator Cbfa1 (core binding factor) is a target of mechanical signals in osteoblastic cells. Low level mechanical deformation (stretching) of human osteoblastic cells directly up-regulates the expression and DNA binding activity of Cbfa1. [18] Genomic differences contribute to individual variances in these regulators. [7, 8] Bone is a dynamic organ that is continuously molded, shaped and repaired.[9]

Gene Mutation and
Compromised Bone Cell Function

Certain hormones, cytokines and humoral factors produced in distant organs can influence bone density and calcium homeostasis locally by inducing NF-kB ligand (RANKL) expression within bone cells. [9] Connective tissue disorders are among the most common genetic maladies. Of these, Osteogenesis Imperfecta (OI) has been intensively studied.[6] Osteopetrosis is a heterogeneous group of heritable conditions in which there is a defect in bone resorption by osteoclasts. The disease is associated with increased skeletal mass due to abnormally dense bone. Remodeling is impacted resulting in irregular bone shapes and structures. Three mutations which impair bone acidification have been linked to OI.[2] FGF mutations produce localized skeletal defects. Skull bone defects are caused by a failure in specification or proliferation of skeletogenic mesenchyme.[10]

Collagen fibrils are degraded and resynthesized as part of continual bone remodeling or turnover. Both a deficit and an excess of collagen can result in disease. Single amino-acid substitutions in the helical portion of type I collagen disrupts bone organization resulting in illness.[6]

Recent discovery of the gene Sclerostin (SOST) in chromosome 17q12-21, provided the first evidence for a role of BMP in bone mass regulation. SOST is the locus for sclerosteosis, an autosomal recessive High Bone Mass (HBM) trait initially found in people with excessive bone formation. Preliminary data suggest that SOST decreases bone formation by suppressing BMP activity in bone. [22]

Low-density lipoprotein receptor-related protein 5 gene (LPR5) encodes a protein responsible for HBM and strikingly dense bone. Gain of function in LPR5 leads to an autosomal dominant variety of HBM, while loss of function leads to osteoporosis. [6]

Genes, Molecules,
and Bone Cell Differentiation

Cell differentiation and specialization are key components of bone remodeling. Genetic studies have identified several genes that control osteoblast development.[14] Transcription factors are cytoskeletal proteins that migrate into the nuclear matrix to bind with specific DNA nucleotide sequences in the promoter regions of genes preceding genomic expression. The osteoblast is the bone-forming cell that originates from mesenchymal stem cells. The “master” regulator of osteoblast differentiation is the transcription factor Cbfa1, the earliest and most specific marker of osteogenesis. Cbfa1 binds to the osteoblast-specific cis-acting element 2 (OSEA) found in the promoter regions of all major osteoblast- specific genes; its function is to control their expression. Mitogen-activated protein kinase (MAPK) signaling is involved in the stimulation of osteoblast-related gene expression by extracellular matrix integrin receptor interaction as well as mechanostressing. [18]

TGF-Beta1 has been shown to prevent osteoblast differentiation, while BMP-2, -4, -6 and -7 can induce osteoblast differentiation.[1] Researchers found that blocking fibronectin receptors with fibronectin antibody did not affect exogenous osteoclast alveolar bone resorption. They concluded fibronectin does not play a significant role in osteoclast adhesion and subsequent resorption.[4] Differentiation (maturation) and/or bone-forming activity of osteoblasts are inversely associated with apoptosis. This suggests higher bone densities and bone formation rates are linked to lower rates of apoptosis.[16]

The RANK pathway in osteoclasts integrates diverse signals that regulate bone resorption and calcium homeostasis. Expression of RANK-ligand (RANKL) by osteoblasts upregulates synthesis of osteoprotegrin (OPN), a decoy molecule that inhibits RANK/RANKL binding, osteoclast differentiation and osteoclastogenesis. The cytokine thrombopoietin, which regulates platelet levels, has also been shown to induce OPG expression, leading to abnormal increases in bone density. Estrogens are humoral factors which reduce bone resorption and augment bone density. Estrogen mediation involves increased OPG expression and/or decreased RANKL expression; the result is decreased RANK activation and subsequently decreased osteoclast activation.[9]

New Definitions of Oral Health,
Disease and Treatment

Bone receptor-ligand docking is actively being studied in a quest to augment health with new pharmaceuticals. Understanding genes which regulate osteoclast bone-matrix acidification, chloride channel function, and osteoblast- derived mineral and protein matrices, will permit gene therapy to restore normal matrix and protein synthesis and function.

There is a molecular link between mechanical stimulation and osteoblast differentiation through the induction of expression and DNA binding potential of Cbfa1. Because Cbfa1 directly controls the rate of bone formation by differentiated osteoblasts, identifying stimuli that increase the expression and/or potency of Cbfa1 in these cells may lead to novel therapeutics to prevent or treat bone loss diseases. The bone generating capacity of small physical signals, such as low amplitude mechanical strain, suggests that biomechanical intervention might help to strengthen bone in degenerative diseases such as osteoporosis, without the side effects associated with pharmacological treatment. In addition to being noninvasive and inducing a therapeutic response from bone tissue itself, low intensity mechanical signals incorporate all aspects of a complex remodeling cycle ultimately improving bone quantity and quality. [18]

Diabetes enhances periodontal bone loss by causing a more persistent inflammatory response, greater loss of attachment, more alveolar bone resorption, and impaired new bone formation. Inhibition of cytokines Tumor Necrosis Factor (TNF) and IL-1B has been shown to reduce the deleterious effects of diabetes regarding bone loss. TNF and IL-1B disregulation and advanced glycation end products are two potential etiologic factors in more persistent inflammatory responses of diabetics. [21]

Elevation of RANKL and decrease in OPG in gingival crevicular fluid of patients with periodontal disease suggests RANKL and OPG are important factors in bone and joint destruction in rheumatoid arthritis and periodontal disease. OPG has a protective role in bone destructive diseases. The RANKL:OPG ratio also increases in multiple myeloma.Treatment with OPG and /or anti-RANKL antibody in patients with TMJ internal derangement seems promising as a therapeutic strategy to inhibit bone destruction. [20]

Osteopontin (OPN) is an integrin-binding bone protein over expressed by many human cancers. In particular, it is elevated in the blood and primary tumors of patients with breast cancer, and in some cases, has been correlated with poor prognosis. Clarification of the molecular requirements for metastasis to lymph nodes and translation of this knowledge into treatment could have important implications for patient management and prognosis.[19] For the practice lifetimes of today’s dental students, clinical reality will include establishing standardized concentrations of regulatory molecules affecting skeletal growth and homeostasis, measuring these individually and adjusting abnormal levels through tissue, cell, and genomically specific interactions.

Conclusions

Inter-patient variability to mechanical application is common in OTM. Reasons for variability lie in genetic differences between people in PDL and alveolar bone cell numbers and genomes, including genes for signaling proteins. The ENCODE (ENCyclopedia of DNA elements) Project has started identifying structural and functional elements of the human genome. Research may eventually permit a correlation of patient genotype with clinical presentation and laboratory-derived protein profiles. This will allow orthodontists to identify biological promoters and inhibitors of OTM and plan molecular intervention to maximize adaptive response. Understanding the communication pathways between PDL and alveolar bone cells and extracellular environments will enhance prospects for therapeutic interventions designed to control the bone remodeling process. [3]

References

1. Short, F., R. M., and M. M., The Role of Growth Factors in Controlling Osteoblast Differentiation. Today’s FDA, 2002: p. 23-25.

2. Tolar, J., et al., Osteopetrosis. New England Journal of Medicine, 2004. 351 (27): p. 2839-49.

3. Masella, R.S., et al., Current concepts in the biology of orthodontic tooth movement. American Journal of Orthodontics & Dentofacial Orthopedics, 2006. 129 (4): p. 458-68.

4. Copus, D.T.T., T. P. Oyen, O. Veis, A., SEM Investigation of the Role of Fibronectin in Osteoclastic Bone Resorption Upon Orthodontic Force Application. Northwestern Dental Research, 1999. 9: p. 5-12.

5. Young, M.F. and M.F. Young, Bone matrix proteins: their function, regulation, and relationship to osteoporosis. Osteoporosis International, 2003. 14 Suppl 3: p. S35-42.

6. Prockop, D.J. and L. Ala-Kokko, Harrison’s Principles of Internal Medicine, 16th Edition Chapter 342. Inherited Disorders of Connective Tissue. 2006, McGraw-Hill.

7. Rosenberg, A.E., Robbins and Cotran Pathologic Basis of Disease 7th Edition CHAPTER 26 Bones, Joints, and Soft Tissue Tumors, V. Kumar, A. Abbas, and N. Fausto, Editors. 2005, Elsevier Saunders: Philadelphia.

8. Aida, Y., et al., The effect of IL-1[beta] on the expression of inflammatory cytokines and their receptors in human chondrocytes. Life Sciences, 2006. 79(8): p. 764-771.

9. Boyle, W.J., et al., Osteoclast differentiation and activation. Nature, 2003. 423(6937): p. 337-42.

10. Helms, J.A., et al., Cranial skeletal biology. Nature, 2003. 423(6937): p. 326-31.

11. Ferrari, S.L., et al., LRP5 gene polymorphisms and idiopathic osteoporosis in men. Bone, 2005. 37(6): p. 770-5.

12. Basch, R.S. Human Hematopoietic Cells Growing in Mouse Bone Marrow. 2006 [cited; Available from: http://www.med. nyu.edu/research/baschr01.html.

13. Oursler, M.J., et al. ASBMR Bone Curriculum. 2006 [cited 17 December 2006]; Available from: http://depts.washington.edu/bonebio/ ASBMRed/
ASBMRed.html.

14. Bringhurst, F.R., et al., Harrison’s Principles of Internal Medicine, 16th Edition Chapter 331. Bone and Mineral Metabolism in Health and Disease. 2006, McGraw-Hill.

15. Rutherford, R.B., et al., Extracellular phosphate alters cementoblast gene expression. Journal of Dental Research, 2006. 85(6): p. 505-9.

16. Sheng, M.H., et al., High osteoblastic activity in C3H/HeJ mice compared to C57BL/6J mice is associated with low apoptosis in C3H/HeJ osteoblasts. Calcified Tissue International, 2006. 78(5): p. 293-301.

17. Kollet, O., et al., Osteoclasts degrade endosteal components and promote mobilization of hematopoietic progenitor cells.[see comment]. Nature Medicine, 2006. 12(6): p. 657-64.

18. Ziros, P., et. al., The Bone Specific Transcriptional Regulator Cbfa1 Is a Target of Mechanical Signals in Osteoblastic Cells. The Journal of Biological Chemistry, 2002. 277 (26): pp. 23934-23941.

19. Allan, Alison., et al., Role of the Integrin-Binding Protein Osteopontin in Lymphatic Metastasis of Breast Cancer. American Journal of Pathology, 2006. 169 (1): 233-246.

20. Wakita, T., et al., Increase in RANKL: OG Ratio in Synovia of Patients with Temporomandibular Joint Disorder. Journal Dent Res.2006. 85 (7): 627-632.

21. Liu, R., et al., Diabetes Enhances Periodontal Bone Loss through Enhanced Resoption and Diminished Bone Formation. Journal Dent Res.2006. 85 (6): 510-514.

22. Harada, Shun-ichi and Rodan, Gideon. Control of osteoblast function and regulation of bone mass. Nature. 2003. 423: 349-354.

We would like to extend our thanks to
Dr. Richard Masella for his encouragement
and the inspiration for this paper.

About the Authors:

Ana Paula Ferraz is a third year dental student at NOVA Southeastern University College of Dental Medicine and Secretary of the Nu Sigma Chapter of Psi Omega. She would like to thank Dr. Masella for his expertise in the field of Orthodontics and his dedication to Psi Omega. Phillipe Sebrechts is pursuing his dental education at NOVA Southeastern University (NSU), Health Professions Division, College of Dental Medicine, and his Masters of Public Health at NSU’s College of Osteopoathic Medicine, both in Fort Lauderdale, Florida.

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