Calcified Tissue International

, Volume 85, Issue 2, pp 119–126 | Cite as

Lysyl Oxidase (Lox) Gene Deficiency Affects Osteoblastic Phenotype

  • N. Pischon
  • J. M. Mäki
  • P. Weisshaupt
  • N. Heng
  • A. H. Palamakumbura
  • P. N’Guessan
  • A. Ding
  • R. Radlanski
  • H. Renz
  • T. A. L. J. J. Bronckers
  • J. Myllyharju
  • A. M. Kielbassa
  • B. M. Kleber
  • J.-P. Bernimoulin
  • P. C. Trackman
Article

Abstract

Lysyl oxidase (LOX) catalyzes cross-linking of elastin and collagen, which is essential for the structural integrity and function of bone tissue. The present study examined the role of Lox gene deficiency for the osteoblast phenotype in primary calvarial osteoblasts from E18.5 Lox knockout (Lox−/−) and wild type (wt) (C57BL/6) mice. Next to Lox gene depletion, mRNA expression of Lox isoforms, LOXL1–4, was significantly downregulated in Lox−/− bone tissue. A significant decrease of DNA synthesis of Lox−/− osteoblasts compared to wt was found. Early stages of osteoblastic apoptosis studied by annexin-V binding as well as later stages of DNA fragmentation were not affected. However, mineral nodule formation and osteoblastic differentiation were markedly decreased, as revealed by significant downregulation of osteoblastic markers, type I collagen, bone sialoprotein, and Runx2/Cbfa1.

Keywords

Lysyl oxidase LOXL1–4 Knockout Osteoblast 

References

  1. 1.
    Boskey AL (1996) Matrix protein and mineralization: an overview. Connect Tissue Res 35:357–363PubMedCrossRefGoogle Scholar
  2. 2.
    Fratzl P, Gupta HS, Paschalis EP, Roschger P (2004) Structure and mechanical quality of the collagen-mineral nano-composite in bone. J Mater Chem 14:2115–2123CrossRefGoogle Scholar
  3. 3.
    Kagan H, Trackman P (1991) Properties and function of lysyl oxidase. Am J Respir Cell Mol Biol 5:206–210PubMedGoogle Scholar
  4. 4.
    Prockop D, Kivirikko K (1995) Collagens: molecular biology, diseases, and potentials for therapy. Annu Rev Biochem 64:403–434PubMedCrossRefGoogle Scholar
  5. 5.
    Yamauchi M (1996) Collagen: the major matrix molecule in mineralized tissues. In: Anderson JJB, Garner S (eds) Calcium and phosphorus in health and disease. CRC Press, New York, pp 127–141Google Scholar
  6. 6.
    Knott L, Bailey A (1998) Collagen cross-links in mineralizing tissues: a review of their chemistry, function, and clinical relevance. Bone 22:181–187PubMedCrossRefGoogle Scholar
  7. 7.
    Kagan H (1986) Biology and regulation of extracellular matrix: a series. In: Mecham RP (ed) Regulation of matrix accumulation, vol 1. Academic Press, Orlando, pp 321–398Google Scholar
  8. 8.
    Mäki JM, Kivirikko KI (2001) Cloning and characterization of a fourth human lysyl oxidase isoenzyme. J Biochem 355:381–387CrossRefGoogle Scholar
  9. 9.
    Mäki JM, Tikkanen H, Kivirikko KI (2001) Cloning and characterization of a fifth human lysyl oxidase isoenzyme: the third member of the lysyl oxidase-related subfamily with four scavenger receptor cysteine-rich domains. Matrix Biol 20:493–496PubMedCrossRefGoogle Scholar
  10. 10.
    Kenyon K, Modi WS, Contente S, Friedman RM (1993) A novel human cDNA with a predicted protein similar to lysyl oxidase maps to chromosome 15q24–q25. J Biol Chem 268:18435–18437PubMedGoogle Scholar
  11. 11.
    Ito H, Akiyama H, Iguchi H, Iyama K, Miyamoto M, Ohsawa K et al (2001) Molecular cloning and biological activity of a novel lysyl oxidase-related gene expressed in cartilage. J Biol Chem 276:24023–24029PubMedCrossRefGoogle Scholar
  12. 12.
    Jourdan-Le Saux C, Le Saux O, Donlon T, Boyd CD, Csiszar K (1998) The human lysyl oxidase-related gene (LOXL2) maps between markers D8S280 and D8S278 on chromosome 8p21.2–p21.3. Genomics 51:305–307PubMedCrossRefGoogle Scholar
  13. 13.
    Huang Y, Dai J, Tang R, Zhao W, Zhou Z, Wang W et al (2001) Cloning and characterization of a human lysyl oxidase-like 3 gene (hLOXL3). Matrix Biol 20:153–157PubMedCrossRefGoogle Scholar
  14. 14.
    Borel A, Eichenberger D, Farjanel J, Kessler E, Gleyzal C, Hulmes DJ et al (2001) Lysyl oxidase-like protein from bovine aorta. Isolation and maturation to an active form by bone morphogenetic protein-1. J Biol Chem 276:48944–48949PubMedCrossRefGoogle Scholar
  15. 15.
    Kim MS, Kim SS, Jung ST, Park JY, Yoon HW, Ko J et al (2003) Expression and purification of enzymatically active forms of the human lysyl oxidase-like protein 4. J Biol Chem 278:52071–52074PubMedCrossRefGoogle Scholar
  16. 16.
    Knott L, Whitehead C, Fleming R, Bailey A (1995) Biochemical changes in the collagenous matrix of osteoporotic avian bone. J Biochem 310:1045–1051Google Scholar
  17. 17.
    Eyre DR, Dickson IR, Van Ness K (1988) Collagen crosslinking in human bone and articular cartilage: age-related changes in the content of mature hydroxypyridinium residues. J Biochem 252:495–500Google Scholar
  18. 18.
    Geiger BJ, Steenbock H, Parsons HT (1933) Lathyrism in the rat. J Nutr 6:427–442Google Scholar
  19. 19.
    Shim H, Harris ZL (2003) Genetic defects in copper metabolism. J Nutr 133:1527S–1531SPubMedGoogle Scholar
  20. 20.
    Uzel M, Scott I, Babakhanlou-Chase H, Palamakumbura A, Pappano W, Hong H et al (2001) Multiple bone morphogenic protein-1 related mammal metalloproteinases process pro-lysyl oxidase at the correct physiological site and control lysyl oxidase activation in mouse embryo fibroblast cultures. J Biol Chem 276:22537–22543PubMedCrossRefGoogle Scholar
  21. 21.
    Guo Y, Pischon N, Palamakumbura AH, Trackman P (2007) Intracellular distribution of the lysyl oxidase propeptide in osteoblastic cells. Am J Physiol Cell Physiol 292:2095–2102CrossRefGoogle Scholar
  22. 22.
    Nellaiappan K, Risitano A, Liu G, Nicklas G, Kagan H (2000) Fully processed lysyl oxidase catalyst translocates from the extracellular space into nuclei of aortic smooth-muscle cells. J Biol Chem 79:576–582Google Scholar
  23. 23.
    Li W, Nellaiappan K, Strassmaier T, Graham L, Thomas K, Kagan H (1997) Localization and activity of lysyl oxidase within nuclei of fibrogenic cells. Proc Natl Acad Sci USA 94:12817–12822PubMedCrossRefGoogle Scholar
  24. 24.
    Palamakumbura A, Jeay S, Guo Y, Pischon N, Sommer P, Sonenshein G et al (2004) The propeptide domain of lysyl oxidase induces phenotypic reversion of ras-transformed cells. J Biol Chem 279:40593–40600PubMedCrossRefGoogle Scholar
  25. 25.
    Jeay S, Pianetti S, Kagan H, Ge S (2003) Lysyl oxidase inhibits ras-mediated transformation by preventing activation of NF-kappa B. Mol Cell Biol 23:2251–2263PubMedCrossRefGoogle Scholar
  26. 26.
    Giampuzzi M, Oleggini R, Di Donato A (2003) Demonstration of in vitro interaction between tumor suppressor lysyl oxidase and histones H1 and H2: definition of the regions involved. Biochim Biophys Acta 1647:245–251PubMedGoogle Scholar
  27. 27.
    Min C, Kirsch KH, Zhao Y, Jeay S, Palamakumbura AH, Trackman PC et al (2007) The tumor suppressor activity of the lysyl oxidase propeptide reverses the invasive phenotype of Her-2/neu-driven breast cancer. Cancer Res 67:1105–1112PubMedCrossRefGoogle Scholar
  28. 28.
    Wu M, Min C, Wang X, Yu Z, Kirsch K, Trackman PC et al (2007) Repression of BCL2 by the tumor suppressor activity of the lysyl oxidase propeptide inhibits transformed phenotype of lung and pancreatic cancer cells. Cancer Res 67:6278–6285PubMedCrossRefGoogle Scholar
  29. 29.
    Hong HH, Pischon N, Santana RB, Palamakumbura AH, Chase HB, Gantz D et al (2004) A role for lysyl oxidase regulation in the control of normal collagen deposition in differentiating osteoblast cultures. J Cell Physiol 200:53–62PubMedCrossRefGoogle Scholar
  30. 30.
    Mäki JM, Räsänen J, Tikkanen H, Sormunen R, Mäkikallio K, Kivirikko KI et al (2002) Inactivation of the lysyl oxidase gene Lox leads to aortic aneurysms, cardiovascular dysfunction, and perinatal death in mice. Circulation 106:2503–2509PubMedCrossRefGoogle Scholar
  31. 31.
    Hornstra I, Birge S, Starcher B, Bailey A, Mecham R, Shapiro S (2003) Lysyl oxidase is required for vascular and diaphragmatic development in mice. J Biol Chem 278:14387–14393PubMedCrossRefGoogle Scholar
  32. 32.
    Mäki J, Sormunen R, Lippo S, Kaarteenaho-Wiik R, Soininen R, Myllyharju J (2005) Lysyl oxidase is essential for normal development and function of the respiratory system and for the integrity of elastic and collagen fibers in various tissues. Am J Pathol 167:927–936PubMedGoogle Scholar
  33. 33.
    Radlanski RJ, Renz H, Klarkowski MC (2003) Prenatal development of the human mandible. 3D reconstructions, morphometry and bone remodelling pattern, sizes 12–117 mm CRL. Anat Embryol 207:221–232PubMedCrossRefGoogle Scholar
  34. 34.
    Bellows CG, Aubin JE, Heersche JNM, Antosz ME (1986) Mineralized bone nodules formed in vitro from enzymatically released rat calvaria cell populations. Calcif Tissue Int 38:143–154PubMedCrossRefGoogle Scholar
  35. 35.
    Ecarot-Charrier B, Glorieux FH, Van der Rest M, Pereira G (1983) Osteoblasts isolated from mouse calvaria initiate matrix mineralization in culture. J Cell Biol 96:639–643PubMedCrossRefGoogle Scholar
  36. 36.
    Hippenstiel S, Schmeck B, N’Guessan PD, Seybold J, Krull M, Preissner K et al (2002) Rho protein inactivation induced apoptosis of cultured human endothelial cells. Am J Physiol Lung Cell Mol Physiol 283:L830–L838PubMedGoogle Scholar
  37. 37.
    N’Guessan PD, Vigelahn M, Bachmann S, Zabel S, Opitz B, Schmeck B et al (2007) The UspA1 protein of Moraxella catarrhalis induces CEACAM-1-dependent apoptosis in alveolar epithelial cells. J Infect Dis 195:1651–1660PubMedCrossRefGoogle Scholar
  38. 38.
    N’Guessan PD, Schmeck B, Ayim A, Hocke AC, Brell B, Hammerschmidt S et al (2005) Streptococcus pneumoniae R6x induced p38 MAPK and JNK-mediated caspase-dependent apoptosis in human endothelial cells. Thromb Haemost 94:295–303PubMedGoogle Scholar
  39. 39.
    Livak KJ, Schnmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-∆∆C(T) method. Methods Enzymol 25:402–408CrossRefGoogle Scholar
  40. 40.
    Stein G, Lian J, Stein J, Van Wijnen J, Montecino M (1996) Transcriptional control of osteoblast growth and differentiation. Physiol Rev 76:593–629PubMedGoogle Scholar
  41. 41.
    Atsawasuwan P, Mochida Y, Parisuthiman D, Yamauchi M (2005) Expression of lysyl oxidase isoforms in MC3T3–E1 osteoblastic cells. Biochem Biophys Res Commun 327:1042–1046PubMedCrossRefGoogle Scholar
  42. 42.
    Pischon N, Darbois LM, Palamakumbura AH, Kessler E, Trackman PC (2004) Regulation of collagen deposition and lysyl oxidase by tumor necrosis factor-alpha in osteoblasts. J Biol Chem 279:30060–30065PubMedCrossRefGoogle Scholar
  43. 43.
    Turecek C, Fratzl-Zelman N, Rumpler M, Buchinger B, Spitzer S, Zoehrer R et al (2008) Collagen cross-linking influences osteoblastic differentiation. Calcif Tissue Int 82:392–400PubMedCrossRefGoogle Scholar
  44. 44.
    Jordan MA (2002) Mechanism of action of antitumor drugs that interacts with microtubules and tubulin. Curr Med Chem Anti-Cancer Agents 2:1–17CrossRefGoogle Scholar
  45. 45.
    Hock JM, Krishnan V, Onyia JE, Bidwell JP, Milas J, Stanislaus D (2001) Osteoblast apoptosis and bone turnover. J Bone Miner Res 16:975–984PubMedCrossRefGoogle Scholar
  46. 46.
    Ducy P, Zhang R, Geoffroy V, Ridall AL, Karsenty G (1997) Osf2/Cbfa1: a transcriptional activator of osteoblast differentiation. Cell 89:747–754PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • N. Pischon
    • 1
  • J. M. Mäki
    • 2
  • P. Weisshaupt
    • 1
  • N. Heng
    • 1
  • A. H. Palamakumbura
    • 3
  • P. N’Guessan
    • 4
  • A. Ding
    • 5
  • R. Radlanski
    • 5
  • H. Renz
    • 5
  • T. A. L. J. J. Bronckers
    • 6
  • J. Myllyharju
    • 2
  • A. M. Kielbassa
    • 1
  • B. M. Kleber
    • 1
  • J.-P. Bernimoulin
    • 1
  • P. C. Trackman
    • 3
  1. 1.Department of Operative Dentistry and Periodontology, CharitéCentrum 3, University School of Dental MedicineCharité – Universitätsmedizin BerlinBerlinGermany
  2. 2.Oulu Center for Cell-Matrix Research, Biocenter Oulu, Department of Medical Biochemistry and Molecular BiologyUniversity of OuluOuluFinland
  3. 3.Department of Oral Biology and Periodontology, Goldman School of Dental MedicineBoston UniversityBostonUSA
  4. 4.Department of Infectious Diseases and PulmonologyCharité – UniversitätsmedizinBerlinGermany
  5. 5.Department of Experimental Dental MedicineCharité – UniversitätsmedizinBerlinGermany
  6. 6.Department of Oral Cell Biology, ACTAVrije UniversiteitAmsterdamThe Netherlands

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