CCN Proteins pp 283-308 | Cite as

Gene Expression Analysis of CCN Protein in Bone Under Mechanical Stress

  • Teruko Takano-Yamamoto
  • Tomohiro Fukunaga
  • Nobuo Takeshita
Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 1489)

Abstract

To investigate mechanical-dependent bone remodeling, we had previously applied various types of mechanical loading onto the teeth of rats and mice. In vitro cultured bone cells were then used to elucidate the mechanisms underlying the specific phenomenon revealed by in vivo experiments. This review describes the techniques used to upregulate CCN2 expression in bone cells produced by different types of mechanical stress, such as fluid shear stress and substrate strain in vitro, and compression or tension force in vivo.

Keywords

Fluid Shear Stress Tooth Movement PDMS Membrane Maxillary Incisor Mandibular Ramus 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.
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Notes

Acknowledgement

This work was supported by grants to T.T.-Y., T.F. and N.T. from Ministry of Education, Culture, Sports, Science and Technology in Japan, Japan Society for the Promotion of Science, and Japan Science and Technology Agency.

References

  1. 1.
    Kruse K, Jülicher F (2005) Oscillations in cell biology. Curr Opin Cell Biol 17:20–26CrossRefPubMedGoogle Scholar
  2. 2.
    Zhou XL, Batiza AF, Loukin SH, Palmer CP, Kung C, Saimi Y (2003) The transient receptor potential channel on the yeast vacuole is mechanosensitive. Proc Natl Acad Sci U S A 100:7105–7110CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Neel PL, Harris RW (1971) Motion-induced inhibition of elongation and induction of dormancy in liquidambar. Science 173:58–59CrossRefPubMedGoogle Scholar
  4. 4.
    Ingber DE (2005) Mechanical control of tissue growth: function follows form. Proc Natl Acad Sci U S A 102:11571–11572CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Tzima E, Irani-Tehrani M, Kiosses WB, Dejana E, Schultz DA, Engelhardt B, Cao G, DeLisser H, Schwartz MA (2005) A mechanosensory complex that mediates the endothelial cell response to fluid shear stress. Nature 437:426–431CrossRefPubMedGoogle Scholar
  6. 6.
    Aikawa R, Nagai T, Kudoh S, Zou Y, Tanaka M, Tamura M, Akazawa H, Takano H, Nagai R, Komuro I (2002) Integrins play a critical role in mechanical stress-induced p38 MAPK activation. Hypertension 39:233–238CrossRefPubMedGoogle Scholar
  7. 7.
    Liu Y, Feng J, Shi L, Niu R, Sun Q, Liu H, Li J, Guo J, Zhu J, Han D (2012) In situ mechanical analysis of cardiomyocytes at nano scales. Nanoscale 4:99–102CrossRefPubMedGoogle Scholar
  8. 8.
    Toms SR, Eberhardt AW (2003) A nonlinear finite element analysis of the periodontal ligament under orthodontic tooth loading. Am J Orthod Dentofacial Orthop 123:657–665CrossRefPubMedGoogle Scholar
  9. 9.
    Fujihara C, Yamada S, Ozaki N, Takeshita N, Kawaki H, Takano-Yamamoto T, Murakami S (2010) Role of mechanical stress-induced glutamate signaling-associated molecules in cytodifferentiation of periodontal ligament cells. J Biol Chem 285:28286–28297CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Hirata H, Tatsumi H, Sokabe M (2008) Mechanical forces facilitate actin polymerization at focal adhesions in a zyxin-dependent manner. J Cell Sci 121:2795–2804CrossRefPubMedGoogle Scholar
  11. 11.
    Takano-Yamamoto T, Soma S, Nakagawa K, Kobayashi Y, Kawakami M, Sakuda M (1991) Comparison of the effects of hydrostatic compressive force on glycosaminoglycan synthesis and proliferation in rabbit chondrocytes from mandibular condylar cartilage, nasal septum, and spheno-occipital synchondrosis in vitro. Am J Orthod Dentofacial Orthop 99:448–455CrossRefPubMedGoogle Scholar
  12. 12.
    Fanning PJ, Emkey G, Smith RJ, Grodzinsky AJ, Szasz N, Trippel SB (2003) Mechanical regulation of mitogen-activated protein kinase signaling in articular cartilage. J Biol Chem 278:50940–50948CrossRefPubMedGoogle Scholar
  13. 13.
    Klein-Nulend J, Nijweide PJ, Burger EH (2003) Osteocyte and bone structure. Curr Osteoporos Rep 1:5–10CrossRefPubMedGoogle Scholar
  14. 14.
    Bonewald LF, Johnson ML (2008) Osteocytes, mechanosensing and Wnt signaling. Bone 42:606–615CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Simons M, Walz G (2006) Polycystic kidney disease: cell division without a c(l)ue? Kidney Int 70:854–864CrossRefPubMedGoogle Scholar
  16. 16.
    Yamamoto K, Takahashi T, Asahara T, Ohura N, Sokabe T, Kamiya A, Ando J (2003) Proliferation, differentiation, and tube formation by endothelial progenitor cells in response to shear stress. J Appl Physiol 95:2081–2088CrossRefPubMedGoogle Scholar
  17. 17.
    Wong M, Siegrist M, Goodwin K (2003) Cyclic tensile strain and cyclic hydrostatic pressure differentially regulate expression of hypertrophic markers in primary chondrocytes. Bone 33:685–693CrossRefPubMedGoogle Scholar
  18. 18.
    Ozcivici E, Luu YK, Adler B, Qin YX, Rubin J, Judex S, Rubin CT (2010) Mechanical signals as anabolic agents in bone. Nat Rev Rheumatol 6:50–59CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Burger EH, Klein-Nulend J (1999) Mechanotransduction in bone—role of the lacuno-canalicular network. FASEB J 13:S101–S112PubMedGoogle Scholar
  20. 20.
    Huiskes R, Ruimerman R, van Lenthe GH, Janssen JD (2000) Effects of mechanical forces on maintenance and adaptation of form in trabecular bone. Nature 405:704–706CrossRefPubMedGoogle Scholar
  21. 21.
    Lanyon LE, Goodship AE, Pye CJ, MacFie JH (1982) Mechanically adaptive bone remodelling. J Biomech 15:141–154CrossRefPubMedGoogle Scholar
  22. 22.
    Lanyon LE (1987) Functional strain in bone tissue as an objective, and controlling stimulus for adaptive bone remodelling. J Biomech 20:1083–1093CrossRefPubMedGoogle Scholar
  23. 23.
    Frost HM (1965) Bone biodynamics. Am J Med Sci 249:614CrossRefGoogle Scholar
  24. 24.
    Terai K, Takano-Yamamoto T, Ohba Y, Hiura K, Sugimoto M, Sato M, Kawahata H, Inaguma N, Kitamura Y, Nomura S (1999) Role of osteopontin in bone remodeling caused by mechanical stress. J Bone Miner Res 14:839–849CrossRefPubMedGoogle Scholar
  25. 25.
    Gerstenfeld LC (1999) Osteopontin in skeletal tissue homeostasis: An emerging picture of the autocrine/paracrine functions of the extracellular matrix. J Bone Miner Res 14:850–855CrossRefPubMedGoogle Scholar
  26. 26.
    Jun JI, Lau LF (2011) Taking aim at the extracellular matrix: CCN proteins as emerging therapeutic targets. Nat Rev Drug Discov 10:945–963CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Kubota S, Takigawa M (2007) Role of CCN2/CTGF/Hcs24 in bone growth. Int Rev Cytol 257:1–41CrossRefPubMedGoogle Scholar
  28. 28.
    Jiang CG, Lv L, Liu FR, Wang ZN, Na D, Li F, Li JB, Sun Z, Xu HM (2013) Connective tissue growth factor is a positive regulator of epithelial-mesenchymal transition and promotes the adhesion with gastric cancer cells in human peritoneal mesothelial cells. Cytokine 61:173–180CrossRefPubMedGoogle Scholar
  29. 29.
    Kawaki H, Kubota S, Suzuki A, Suzuki M, Kohsaka K, Hoshi K, Fujii T, Lazar N, Ohgawara T, Maeda T, Perbal B, Takano-Yamamoto T, Takigawa M (2011) Differential roles of CCN family proteins during osteoblast differentiation: Involvement of Smad and MAPK signaling pathways. Bone 49:975–989CrossRefPubMedGoogle Scholar
  30. 30.
    Nakanishi T, Nishida T, Shimo T, Kobayashi K, Kubo T, Tamatani T, Tezuka K, Takigawa M (2000) Effects of CTGF/Hcs24, a product of a hypertrophic chondrocyte-specific gene, on the proliferation and differentiation of chondrocytes in culture. Endocrinology 141:264–273PubMedGoogle Scholar
  31. 31.
    Kawaki H, Kubota S, Suzuki A, Lazar N, Yamada T, Matsumura T, Ohgawara T, Maeda T, Perbal B, Lyons KM, Takigawa M (2008) Cooperative regulation of chondrocyte differentiation by CCN2 and CCN3 shown by a comprehensive analysis of the CCN family proteins in cartilage. J Bone Miner Res 23:1751–1764CrossRefPubMedGoogle Scholar
  32. 32.
    Kubota S, Takigawa M (2011) The role of CCN2 in cartilage and bone development. J Cell Commun Signal 5:209–217CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Hishikawa K, Oemar BS, Nakaki T (2001) Static pressure regulates connective tissue growth factor expression in human mesangial cells. J Biol Chem 276:16797–16803CrossRefPubMedGoogle Scholar
  34. 34.
    Schild C, Trueb B (2002) Mechanical stress is required for high-level expression of connective tissue growth factor. Exp Cell Res 274:83–91CrossRefPubMedGoogle Scholar
  35. 35.
    Hoshi K, Kawaki H, Takahashi I, Takeshita N, Seiryu M, Murshid SA, Masuda T, Anada T, Kato R, Kitaura H, Suzuki O, Takano-Yamamoto T (2014) Compressive force-produced CCN2 induces osteocyte apoptosis through ERK1/2 pathway. J Bone Miner Res 29:1244–1257CrossRefPubMedGoogle Scholar
  36. 36.
    Honjo T, Kubota S, Kamioka H, Sugawara Y, Ishihara Y, Yamashiro T, Takigawa M, Takano-Yamamoto T (2012) Promotion of Ccn2 expression and osteoblastic differentiation by actin polymerization, which is induced by laminar fluid flow stress. J Cell Commun Signal 6:225–232CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Yamashiro T, Fukunaga T, Kobashi N, Kamioka H, Nakanishi T, Takigawa M, Takano-Yamamoto T (2001) Mechanical stimulation induces CTGF expression in rat osteocytes. J Dent Res 80:461–465CrossRefPubMedGoogle Scholar
  38. 38.
    Kuroda S, Balam TA, Sakai Y, Tamamura N, Takano-Yamamoto T (2005) Expression of osteopontin mRNA in odontoclasts revealed by in situ hybridization during experimental tooth movement in mice. J Bone Miner Metab 23:110–113CrossRefPubMedGoogle Scholar
  39. 39.
    Sakai Y, Balam TA, Kuroda S, Tamamura N, Fukunaga T, Takigawa M, Takano-Yamamoto T (2009) CTGF and apoptosis in mouse osteocytes induced by tooth movement. J Dent Res 88:345–350CrossRefPubMedGoogle Scholar
  40. 40.
    Soma S, Iwamoto M, Higuchi Y, Kurisu K (1999) Effects of continuous infusion of PTH on experimental tooth movement in rats. J Bone Miner Res 14:546–554CrossRefPubMedGoogle Scholar
  41. 41.
    Soma S, Matsumoto S, Higuchi Y, Takano-Yamamoto T, Yamashita K, Kurisu K, Iwamoto M (2000) Local and chronic application of PTH accelerates tooth movement in rats. J Dent Res 79:1717–1724CrossRefPubMedGoogle Scholar
  42. 42.
    Hakami Z, Kitaura H, Kimura K, Ishida M, Sugisawa H, Ida H, Jafari S, Takano-Yamamoto T (2015) Effect of interleukin-4 on orthodontic tooth movement and associated root resorption. Eur J Orthod 37:87–94CrossRefPubMedGoogle Scholar
  43. 43.
    Takeshita N, Hasegawa M, Seki D, Seiryu M, Miyashita S, Takano I, Oyanagi T, Miyajima Y, Takano-Yamamoto T (2016) In vivo expression and regulation of genes associated with vascularization during early response of sutures to tensile force. J Bone Miner Metab 9:26825658Google Scholar
  44. 44.
    Masuda T, Takahashi I, Anada T, Arai F, Fukuda T, Takano-Yamamoto T, Suzuki O (2008) Development of a cell culture system loading cyclic mechanical strain to chondrogenic cells. J Biotechnol 133:231–238CrossRefPubMedGoogle Scholar
  45. 45.
    Yamashiro T, Takano-Yamamoto T (2001) Influences of ovariectomy on experimental tooth movement in the rat. J Dent Res 80:1858–1861CrossRefPubMedGoogle Scholar
  46. 46.
    Takano-Yamamoto T, Kawakami M, Yamashiro T (1992) Effect of age on the rate of tooth movement in combination with local use of 1,25(OH)2D3 and mechanical force in the rat. J Dent Res 71:1487–1492CrossRefPubMedGoogle Scholar
  47. 47.
    Waldo CM, Rothblatt JM (1954) Histologic response to tooth movement in the laboratory rat; procedure and preliminary observations. J Dent Res 33:481–486CrossRefPubMedGoogle Scholar
  48. 48.
    Takano-Yamamoto T, Kawakami M, Kobayashi Y, Yamashiro T, Sakuda M (1992) The effect of local application of 1,25-dihydroxycholecalciferol on osteoclast numbers in orthodontically treated rats. J Dent Res 71:53–59CrossRefPubMedGoogle Scholar
  49. 49.
    Takimoto A, Kawatsu M, Yoshimoto Y, Kawamoto T, Seiryu M, Takano-Yamamoto T, Hiraki Y, Shukunami C (2015) Scleraxis and osterix antagonistically regulate tensile force-responsive remodeling of the periodontal ligament and alveolar bone. Development 142:787–796CrossRefPubMedGoogle Scholar
  50. 50.
    Fukunaga T, Yamashiro T, Oya S, Takeshita N, Takigawa M, Takano-Yamamoto T (2003) Connective tissue growth factor mRNA expression pattern in cartilages is associated with their type I collagen expression. Bone 33:911–918CrossRefPubMedGoogle Scholar
  51. 51.
    Zheng L, Yamashiro T, Fukunaga T, Balam TA, Takano-Yamamoto T (2005) Bone morphogenetic protein 3 expression pattern in rat condylar cartilage, femoral cartilage and mandibular fracture callus. Eur J Oral Sci 113:318–325CrossRefPubMedGoogle Scholar
  52. 52.
    Sugawara Y, Ando R, Kamioka H, Ishihara Y, Murshid SA, Hashimoto K, Kataoka N, Tsujioka K, Kajiya F, Yamashiro T, Takano-Yamamoto T (2008) The alteration of a mechanical property of bone cells during the process of changing from osteoblasts to osteocytes. Bone 43:19–24CrossRefPubMedGoogle Scholar
  53. 53.
    Fujisawa T, Hattori T, Takahashi K, Kuboki T, Yamashita A, Takigawa M (1999) Cyclic mechanical stress induces extracellular matrix degradation in cultured chondrocytes via gene expression of matrix metalloproteinases and interleukin-1. J Biochem 125:966–975CrossRefPubMedGoogle Scholar
  54. 54.
    Kessler D, Dethlefsen S, Haase I, Plomann M, Hirche F, Krieg T, Eckes B (2001) Fibroblasts in mechanically stressed collagen lattices assume a “synthetic” phenotype. J Biol Chem 276:36575–36585CrossRefPubMedGoogle Scholar
  55. 55.
    Takano-Yamamoto T, Takemura T, Kitamura Y, Nomura S (1994) Site-specific expression of mRNAs for osteonectin, osteocalcin, and osteopontin revealed by in situ hybridization in rat periodontal ligament during physiological tooth movement. J Histochem Cytochem 42:885–896CrossRefPubMedGoogle Scholar
  56. 56.
    Miyawaki S, Forbes DP (1987) The morphologic and biochemical effects of tensile force application to the interparietal suture of the Sprague-Dawley rat. Am J Orthod Dentofacial Orthop 92:123–133CrossRefPubMedGoogle Scholar
  57. 57.
    Steenvoorden GP, van de Velde JP, Prahl-Andersen B (1990) The effect of duration and magnitude of tensile mechanical forces on sutural tissue in vivo. Eur J Orthod 12:330–339CrossRefPubMedGoogle Scholar
  58. 58.
    Weinbaum S, Cowin SC, Zeng Y (1994) A model for the excitation of osteocytes by mechanical loading-induced bone fluid shear stresses. J Biomech 27:339–360CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Teruko Takano-Yamamoto
    • 1
  • Tomohiro Fukunaga
    • 1
  • Nobuo Takeshita
    • 1
  1. 1.Division of Orthodontics and Dentofacial OrthopedicsTohoku University Graduate School of DentistrySendaiJapan

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