Clinical Orthopaedics and Related Research®

, Volume 467, Issue 12, pp 3206–3212 | Cite as

Successful Spinal Fusion by E. coli-derived BMP-2-adsorbed Porous β-TCP Granules: A Pilot Study

  • Sho Dohzono
  • Yuuki Imai
  • Hiroaki Nakamura
  • Shigeyuki Wakitani
  • Kunio Takaoka
Symposium: Tribute to Dr. Marshall Urist: Musculoskeletal Growth Factors


Bone morphogenetic proteins (BMPs) were originally identified as osteoinductive proteins. With cloning of BMP genes, studies of BMPs and their clinical application have advanced. However, with increasing clinical applications, drug delivery systems and production costs have become more important issues. To address these issues, we asked whether E. coli-derived rhBMP-2 (E-BMP-2)-adsorbed porous β-TCP granules could achieve posterolateral lumbar fusion in a rabbit model similar to autogenous bone grafts. Lumbar spinal fusion masses were evaluated by 3-D computed tomography, mechanical testing, and histological analyses 8 weeks after surgery. By these measures E-BMP-2-adsorbed β-TCP granules achieved lumbar spinal fusion in dose-dependent fashion in a rabbit model as well as autogenous bone graft. Our preliminary findings suggest E-BMP-2-adsorbed porous β-TCP could be a novel, effective alternative to autogenous bone grafting for generating new bone and promoting regenerative repair of bone, and potentially utilizable in the clinical setting for treating spinal disorders.


Spinal Fusion Transverse Process Autogenous Bone Autogenous Bone Graft Lumbar Spinal Fusion 
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.



We thank Drs. A. Suzuki, K. Yano, T. Matsumoto, H. Yasuda, K. Sugama, H. Irie, and A. Yamada, W. Fukushima, M Fukui, and also Ms. A. Inagaki, Ms. K. Hata, Ms. K. Kamei, and Ms. Y. Hanamoto for technical and statistical assistance. E-BMP-2 was produced and kindly provided by Dr. W. Sebald (Würzburg, Germany) and donated to us through Osteopharma Inc. (Osaka, Japan). The β-TCP granules were donated by HOYA Corp. (Tokyo, Japan).


  1. 1.
    Akamaru T, Suh D, Boden SD, Kim HS, Minamide A, Louis-Ugbo J. Simple carrier matrix modifications can enhance delivery of recombinant human bone morphogenetic protein-2 for posterolateral spine fusion. Spine. 2003;28:429–434.CrossRefPubMedGoogle Scholar
  2. 2.
    Allen RT, Lee YP, Stimson E, Garfin SR. Bone morphogenetic protein-2 (BMP-2) in the treatment of pyogenic vertebral osteomyelitis. Spine. 2007;32:2996–3006.CrossRefPubMedGoogle Scholar
  3. 3.
    Arosarena OA, Collins WL. Bone regeneration in the rat mandible with bone morphogenetic protein-2: a comparison of two carriers. Otolaryngol Head Neck Surg. 2005;132:592–597.CrossRefPubMedGoogle Scholar
  4. 4.
    Arrington ED, Smith WJ, Chambers HG, Bucknell AL, Davino NA. Complications of iliac crest bone graft harvesting. Clin Orthop Relat Res. 1996;329:300–309.CrossRefPubMedGoogle Scholar
  5. 5.
    Boden SD, Kang J, Sandhu H, Heller JG. Use of recombinant human bone morphogenetic protein-2 to achieve posterolateral lumbar spine fusion in humans: a prospective, randomized clinical pilot trial: 2002 Volvo Award in clinical studies. Spine. 2002;27:2662–2673.CrossRefPubMedGoogle Scholar
  6. 6.
    Boden SD, Schimandle JH, Hutton WC. An experimental lumbar intertransverse process spinal fusion model. Radiographic, histologic, and biomechanical healing characteristics. Spine. 1995;20:412–420.PubMedCrossRefGoogle Scholar
  7. 7.
    David SM, Gruber HE, Meyer RA, Jr., Murakami T, Tabor OB, Howard BA, Wozney JM, Hanley EN, Jr. Lumbar spinal fusion using recombinant human bone morphogenetic protein in the canine. A comparison of three dosages and two carriers. Spine. 1999;24:1973–1979.CrossRefPubMedGoogle Scholar
  8. 8.
    Dimar JR, Glassman SD, Burkus KJ, Carreon LY. Clinical outcomes and fusion success at 2 years of single-level instrumented posterolateral fusions with recombinant human bone morphogenetic protein-2/compression resistant matrix versus iliac crest bone graft. Spine. 2006;31:2534–2539; discussion 2540.CrossRefPubMedGoogle Scholar
  9. 9.
    Glassman SD, Dimar JR, 3rd, Burkus K, Hardacker JW, Pryor PW, Boden SD, Carreon LY. The efficacy of rhBMP-2 for posterolateral lumbar fusion in smokers. Spine. 2007;32:1693–1698.CrossRefPubMedGoogle Scholar
  10. 10.
    Govender S, Csimma C, Genant HK, Valentin-Opran A, Amit Y, Arbel R, Aro H, Atar D, Bishay M, Borner MG, Chiron P, Choong P, Cinats J, Courtenay B, Feibel R, Geulette B, Gravel C, Haas N, Raschke M, Hammacher E, van der Velde D, Hardy P, Holt M, Josten C, Ketterl RL, Lindeque B, Lob G, Mathevon H, McCoy G, Marsh D, Miller R, Munting E, Oevre S, Nordsletten L, Patel A, Pohl A, Rennie W, Reynders P, Rommens PM, Rondia J, Rossouw WC, Daneel PJ, Ruff S, Ruter A, Santavirta S, Schildhauer TA, Gekle C, Schnettler R, Segal D, Seiler H, Snowdowne RB, Stapert J, Taglang G, Verdonk R, Vogels L, Weckbach A, Wentzensen A, Wisniewski T. Recombinant human bone morphogenetic protein-2 for treatment of open tibial fractures: a prospective, controlled, randomized study of four hundred and fifty patients. J Bone Joint Surg Am. 2002;84:2123–2134.CrossRefPubMedGoogle Scholar
  11. 11.
    Grauer JN, Patel TC, Erulkar JS, Troiano NW, Panjabi MM, Friedlaender GE. 2000 Young Investigator Research Award winner. Evaluation of OP-1 as a graft substitute for intertransverse process lumbar fusion. Spine. 2001;26:127–133.CrossRefPubMedGoogle Scholar
  12. 12.
    Hoshino M, Namikawa T, Kato M, Terai H, Taguchi S, Takaoka K. Repair of bone defects in revision hip arthroplasty by implantation of a new bone-inducing material comprised of recombinant human BMP-2, Beta-TCP powder, and a biodegradable polymer: an experimental study in dogs. J Orthop Res. 2007;25:1042–1051.CrossRefPubMedGoogle Scholar
  13. 13.
    Hsu WK, Wang JC. The use of bone morphogenetic protein in spine fusion. Spine J. 2008;8:419–425.CrossRefPubMedGoogle Scholar
  14. 14.
    Itoh H, Ebara S, Kamimura M, Tateiwa Y, Kinoshita T, Yuzawa Y, Takaoka K. Experimental spinal fusion with use of recombinant human bone morphogenetic protein 2. Spine. 1999;24:1402–1405.CrossRefPubMedGoogle Scholar
  15. 15.
    Johnsson R, Stromqvist B, Aspenberg P. Randomized radiostereometric study comparing osteogenic protein-1 (BMP-7) and autograft bone in human noninstrumented posterolateral lumbar fusion: 2002 Volvo Award in clinical studies. Spine. 2002;27:2654–2661.CrossRefPubMedGoogle Scholar
  16. 16.
    Joseph V, Rampersaud YR. Heterotopic bone formation with the use of rhBMP2 in posterior minimal access interbody fusion: a CT analysis. Spine. 2007;32:2885–2890.CrossRefPubMedGoogle Scholar
  17. 17.
    Konishi S, Nakamura H, Seki M, Nagayama R, Yamano Y. Hydroxyapatite granule graft combined with recombinant human bone morphogenic protein-2 for solid lumbar fusion. J Spinal Disord Tech. 2002;15:237–244.PubMedGoogle Scholar
  18. 18.
    Kubler NR, Reuther JF, Faller G, Kirchner T, Ruppert R, Sebald W. Inductive properties of recombinant human BMP-2 produced in a bacterial expression system. Int J Oral Maxillofac Surg. 1998;27:305–309.CrossRefPubMedGoogle Scholar
  19. 19.
    Kurz LT, Garfin SR, Booth RE, Jr. Harvesting autogenous iliac bone grafts. A review of complications and techniques. Spine. 1989;14:1324–1331.CrossRefPubMedGoogle Scholar
  20. 20.
    Lewandrowski KU, Nanson C, Calderon R. Vertebral osteolysis after posterior interbody lumbar fusion with recombinant human bone morphogenetic protein 2: a report of five cases. Spine J. 2007;7:609–614.CrossRefPubMedGoogle Scholar
  21. 21.
    Martin GJ, Jr., Boden SD, Marone MA, Marone MA, Moskovitz PA. Posterolateral intertransverse process spinal arthrodesis with rhBMP-2 in a nonhuman primate: important lessons learned regarding dose, carrier, and safety. J Spinal Disord. 1999;12:179–186.PubMedGoogle Scholar
  22. 22.
    Minamide A, Kawakami M, Hashizume H, Sakata R, Tamaki T. Evaluation of carriers of bone morphogenetic protein for spinal fusion. Spine. 2001;26:933–939.CrossRefPubMedGoogle Scholar
  23. 23.
    Minamide A, Kawakami M, Hashizume H, Sakata R, Yoshida M, Tamaki T. Experimental study of carriers of bone morphogenetic protein used for spinal fusion. J Orthop Sci. 2004;9:142–151.CrossRefPubMedGoogle Scholar
  24. 24.
    Nakagawa K, Imai Y, Ohta Y, Takaoka K. Prostaglandin E2 EP4 agonist (ONO-4819) accelerates BMP-induced osteoblastic differentiation. Bone. 2007;41:543–548.CrossRefPubMedGoogle Scholar
  25. 25.
    Namikawa T, Terai H, Hoshino M, Kato M, Toyoda H, Yano K, Nakamura H, Takaoka K. Enhancing effects of a prostaglandin EP4 receptor agonist on recombinant human bone morphogenetic protein-2 mediated spine fusion in a rabbit model. Spine. 2007;32:2294–2299.CrossRefPubMedGoogle Scholar
  26. 26.
    Namikawa T, Terai H, Suzuki E, Hoshino M, Toyoda H, Nakamura H, Miyamoto S, Takahashi N, Ninomiya T, Takaoka K. Experimental spinal fusion with recombinant human bone morphogenetic protein-2 delivered by a synthetic polymer and beta-tricalcium phosphate in a rabbit model. Spine. 2005;30:1717–1722.CrossRefPubMedGoogle Scholar
  27. 27.
    Ohta Y, Nakagawa K, Imai Y, Katagiri T, Koike T, Takaoka K. Cyclic AMP enhances Smad-mediated BMP signaling through PKA-CREB pathway. J Bone Miner Metab. 2008;26:478–484.PubMedGoogle Scholar
  28. 28.
    Raisz LG. Pathogenesis of osteoporosis: concepts, conflicts, and prospects. J Clin Invest. 2005;115:3318–3325.CrossRefPubMedGoogle Scholar
  29. 29.
    Ruppert R, Hoffmann E, Sebald W. Human bone morphogenetic protein 2 contains a heparin-binding site which modifies its biological activity. Eur J Biochem. 1996;237:295–302.CrossRefPubMedGoogle Scholar
  30. 30.
    Saito N, Okada T, Horiuchi H, Murakami N, Takahashi J, Nawata M, Ota H, Nozaki K, Takaoka K. A biodegradable polymer as a cytokine delivery system for inducing bone formation. Nat Biotechnol. 2001;19:332–335.CrossRefPubMedGoogle Scholar
  31. 31.
    Sandhu HS, Kanim LE, Kabo JM, Toth JM, Zeegen EN, Liu D, Delamarter RB, Dawson EG. Effective doses of recombinant human bone morphogenetic protein-2 in experimental spinal fusion. Spine. 1996;21:2115–2122.CrossRefPubMedGoogle Scholar
  32. 32.
    Santoni BG, Pluhar GE, Motta T, Wheeler DL. Hollow calcium phosphate microcarriers for bone regeneration: in vitro osteoproduction and ex vivo mechanical assessment. Biomed Mater Eng. 2007;17:277–289.PubMedGoogle Scholar
  33. 33.
    Sasaoka R, Terai H, Toyoda H, Imai Y, Sugama R, Takaoka K. A prostanoid receptor EP4 agonist enhances ectopic bone formation induced by recombinant human bone morphogenetic protein-2. Biochem Biophys Res Commun. 2004;318:704–709.CrossRefPubMedGoogle Scholar
  34. 34.
    Schimandle JH, Boden SD, Hutton WC. Experimental spinal fusion with recombinant human bone morphogenetic protein-2. Spine. 1995;20:1326–1337.PubMedGoogle Scholar
  35. 35.
    Seeherman HJ, Bouxsein M, Kim H, Li R, Li XJ, Aiolova M, Wozney JM. Recombinant human bone morphogenetic protein-2 delivered in an injectable calcium phosphate paste accelerates osteotomy-site healing in a nonhuman primate model. J Bone Joint Surg Am. 2004;86:1961–1972.PubMedGoogle Scholar
  36. 36.
    Taguchi S, Namikawa T, Ieguchi M, Takaoka K. Reconstruction of bone defects using rhBMP-2-coated devitalized bone. Clin Orthop Relat Res. 2007;461:162–169.PubMedGoogle Scholar
  37. 37.
    Takayanagi H. Osteoimmunology: shared mechanisms and crosstalk between the immune and bone systems. Nat Rev Immunol. 2007;7:292–304.CrossRefPubMedGoogle Scholar
  38. 38.
    Toyoda H, Terai H, Sasaoka R, Oda K, Takaoka K. Augmentation of bone morphogenetic protein-induced bone mass by local delivery of a prostaglandin E EP4 receptor agonist. Bone. 2005;37:555–562.CrossRefPubMedGoogle Scholar
  39. 39.
    Urist MR. Bone: formation by autoinduction. Science. 1965;150:893–899.CrossRefPubMedGoogle Scholar
  40. 40.
    Weigel U, Meyer M, Sebald W. Mutant proteins of human interleukin 2. Renaturation yield, proliferative activity and receptor binding. Eur J Biochem. 1989;180:295–300.CrossRefPubMedGoogle Scholar
  41. 41.
    Wong DA, Kumar A, Jatana S, Ghiselli G, Wong K. Neurologic impairment from ectopic bone in the lumbar canal: a potential complication of off-label PLIF/TLIF use of bone morphogenetic protein-2 (BMP-2). Spine J. 2007;8:1011–1018.CrossRefPubMedGoogle Scholar
  42. 42.
    Wozney JM, Rosen V, Celeste AJ, Mitsock LM, Whitters MJ, Kriz RW, Hewick RM, Wang EA. Novel regulators of bone formation: molecular clones and activities. Science. 1988;242:1528–1534.CrossRefPubMedGoogle Scholar
  43. 43.
    Wu CH, Hara K, Ozawa H. Enhanced osteoinduction by intramuscular grafting of BMP-beta-TCP compound pellets into murine models. Arch Histol Cytol. 1992;55:97–112.CrossRefPubMedGoogle Scholar
  44. 44.
    Yamamoto Y, Udagawa N, Matsuura S, Nakamichi Y, Horiuchi H, Hosoya A, Nakamura M, Ozawa H, Takaoka K, Penninger JM, Noguchi T, Takahashi N. Osteoblasts provide a suitable microenvironment for the action of receptor activator of nuclear factor-kappaB ligand. Endocrinology. 2006;147:3366–3374.CrossRefPubMedGoogle Scholar
  45. 45.
    Yano K, Hoshino M, Ohta Y, Manaka T, Naka Y, Imai Y, Sebald W, Takaoka K. Osteoinductive capacity and heat stability of recombinant human bone morphogenetic protein-2 produced by Escherichia coli and dimerized by biochemical processing. J Bone Miner Metab. 2009;27:355–363.CrossRefPubMedGoogle Scholar
  46. 46.
    Zaidi M. Skeletal remodeling in health and disease. Nat Med. 2007;13:791–801.CrossRefPubMedGoogle Scholar

Copyright information

© The Association of Bone and Joint Surgeons® 2009

Authors and Affiliations

  • Sho Dohzono
    • 1
  • Yuuki Imai
    • 1
  • Hiroaki Nakamura
    • 1
  • Shigeyuki Wakitani
    • 1
  • Kunio Takaoka
    • 1
  1. 1.Department of Orthopaedic SurgeryOsaka City University Graduate School of MedicineOsakaJapan

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