The mechanism research of non-Smad dependent TAK1 signaling pathway in the treatment of bone defects by recombination BMP-2-loaded hollow hydroxyapatite microspheres/chitosan composite

  • Jingtang Li
  • Shilang Xiong
  • Linghua Ding
  • Jianhua Zeng
  • Peng Qiu
  • Jianguo Zhou
  • Xingen Liao
  • Long XiongEmail author
Delivery Systems Original Research
Part of the following topical collections:
  1. Delivery Systems



The present study aimed to evaluate whether the non-Smad dependent TAK1 signaling pathway (BMP-2-TAK1-p38-Osx signaling pathway) played an important role in bone repair mediated by hollow hydroxyapatite (HA) microspheres/chitosan (CS) composite.


Firstly, the biological activity of rhBMP-2 released from the complex was investigated. Then, differentiation test of osteoblasts including ALP activity and calcium deposition, X-ray scoring and three-point bending test were performed. Finally, the mRNAs expression of TAK1, p38, Osx and osteogenic markers was tested by reverse transcription-polymerase chain reaction (RT-PCR).


RhBMP-2 could be loaded and released from the complex in bioactive form. Additionally, the complex provided a prolonged period of time compared with HA/CS scaffolds. Serum ALP activity was significantly decreased in the TAK1 inhibitor group and p38 inhibitor group. In the X-ray radiography, bone callus was observed in rhBMP-2-loaded hollow HA microspheres/CS composite group. In the three-point bending test, load values in p38 inhibitor group decreased. In the animal model, the mRNA expression of BSP on day 90 was significantly decreased in the p38 inhibitor group and TAK1 inhibitor group. In MC3T3-E1 cells, the mRNA expression of OSX was remarkably up-regulated in both rhBMP-2 group or rhBMP-2-loaded hollow HA microspheres/CS composite group; while the mRNA expression of OSX was significantly down-regulated in TAK1 inhibitor group and p38 inhibitor group.


The BMP-2-TAK1-p38-OSX signaling pathway may play an important role in bone formation and repair mediated by rhBMP-2-loaded hollow HA microspheres/CS composite.



This study was funded by National Natural Science Foundation of China of “The mechanism research of non-Smad dependent TAK1 signaling pathway in the treatment of bone defects by recombination BMP-2-loaded hollow hydroxyapatite microspheres/chitosan composite” (81560355) and “Mechanism study of bone repair mediated by rhBMP-2/rhCXCL13-loaded hollow hydroxyapatite microspheres/chitosan composite” (81560377).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Research involving Animals

All animal studies were approved by the Jiangxi Provincial People’s Hospital Affiliated to Nanchang University and were in accordance with the Guidelines for Care and Use of Experimental Animals.


  1. 1.
    Giannoudis PV, Dinopoulos H, Tsiridis E. Bone substitutes: an update. Injury. 2005;36(Suppl 3):S20. Google Scholar
  2. 2.
    Bauer TW, Muschler GF. Bone graft materials. An overview of the basic science. Clin Orthop Relat Res. 2000;10:27.Google Scholar
  3. 3.
    Fleming JE,Jr, Cornell CN, Muschler GF. Bone cells and matrices in orthopedic tissue engineering. Orthop Clin North Am. 2000;31:357.Google Scholar
  4. 4.
    Laurencin C, Khan Y, El-Amin SF. Bone graft substitutes. Expert Rev Med Devices. 2006;3:49. Google Scholar
  5. 5.
    Perry CR. Bone repair techniques, bone graft, and bone graft substitutes. Clin Orthop Relat Res. 1999;71:86.Google Scholar
  6. 6.
    Zimmermann G, Moghaddam A. Allograft bone matrix versus synthetic bone graft substitutes. Injury. 2011;42(Suppl 2):S16. Google Scholar
  7. 7.
    Crowley C, Wong JM, Fisher DM, Khan WS. A systematic review on preclinical and clinical studies on the use of scaffolds for bone repair in skeletal defects. Curr Stem Cell Res Ther. 2013;8:243.Google Scholar
  8. 8.
    Moore WR, Graves SE, Bain GI. Synthetic bone graft substitutes. ANZ J Surg. 2001;71:354.Google Scholar
  9. 9.
    Finkemeier CG. Bone-grafting and bone-graft substitutes. J Bone Jt Surg Am. 2002;84:454.Google Scholar
  10. 10.
    Kirker-Head CA. Potential applications and delivery strategies for bone morphogenetic proteins. Adv Drug Deliv Rev. 2000;43:65.Google Scholar
  11. 11.
    Fu H, Rahaman MN, Day DE, Brown RF. Hollow hydroxyapatite microspheres as a device for controlled delivery of proteins. J Mater Sci Mater Med. 2011;22:579. Scholar
  12. 12.
    Shiels SM, Solomon KD, Pilia M, Appleford MR, Ong JL. BMP-2 tethered hydroxyapatite for bone tissue regeneration: coating chemistry and osteoblast attachment. J Biomed Mater Res Part A. 2012;100:3117. Google Scholar
  13. 13.
    Babensee JE, McIntire LV, Mikos AG. Growth factor delivery for tissue engineering. Pharm Res. 2000;17:497.Google Scholar
  14. 14.
    Hong MH, Son JS, Kim KM, Han M, Oh DS, Lee YK. Drug-loaded porous spherical hydroxyapatite granules for bone regeneration. J Mater Sci Mater Med. 2011;22:349. Google Scholar
  15. 15.
    Kean T, Thanou M. Biodegradation, biodistribution and toxicity of chitosan. Adv Drug Deliv Rev. 2010;62:3. Google Scholar
  16. 16.
    Onishi H, Machida Y. Biodegradation and distribution of water-soluble chitosan in mice. Biomaterials. 1999;20:175.Google Scholar
  17. 17.
    Dimitriou R, Tsiridis E, Giannoudis PV. Current concepts of molecular aspects of bone healing. Injury. 2005;36:1392. Google Scholar
  18. 18.
    Abarrategi A, Gutierrez MC, Moreno-Vicente C, Hortigüela MJ, Ramos V, López-Lacomba JL, et al. Multiwall carbon nanotube scaffolds for tissue engineering purposes. Biomaterials. 2008;29:94. Google Scholar
  19. 19.
    Abarrategi A, Moreno-Vicente C, Ramos V, Aranaz I, Sanz Casado JV, Lopez-Lacomba JL. Improvement of porous beta-TCP scaffolds with rhBMP-2 chitosan carrier film for bone tissue application. Tissue Eng Part A. 2008;14:1305. Google Scholar
  20. 20.
    Lopez-Lacomba JL, Garcia-Cantalejo JM, Sanz Casado JV, Abarrategi A, Correas Magana V, Ramos V. Use of rhBMP-2 Activated Chitosan Films To Improve Osseointegration. Biomacromolecules. 2006;7:792. Google Scholar
  21. 21.
    Stephan SJ, Tholpady SS, Gross B, Petrie-Aronin CE, Botchway EA, Nair LS, et al. Injectable tissue-engineered bone repair of a rat calvarial defect. Laryngoscope. 2010;120:895.
  22. 22.
    Yao AH, Li XD, Xiong L, Zeng JH, Xu J, Wang DP. Hollow hydroxyapatite microspheres/chitosan composite as a sustained delivery vehicle for rhBMP-2 in the treatment of bone defects. J Mater Sci Mater Med. 2015;26:5336.
  23. 23.
    Xiong L, Zeng J, Yao A, Tu Q, Li J, Yan L, et al. BMP2-loaded hollow hydroxyapatite microspheres exhibit enhanced osteoinduction and osteogenicity in large bone defects. Int J Nanomed. 2015;10:517.
  24. 24.
    Nakashima K, Zhou X, Kunkel G, Zhang Z, Deng JM, Behringer RR, et al. The novel zinc finger-containing transcription factor osterix is required for osteoblast differentiation and bone formation. Cell. 2002;108:17.Google Scholar
  25. 25.
    Zhou X, Zhang Z, Feng JQ, Dusevich VM, Sinha K, Zhang H, et al. Multiple functions of Osterix are required for bone growth and homeostasis in postnatal mice. Proc Natl Acad Sci USA. 2010;107:12919. Google Scholar
  26. 26.
    Chen G, Deng C, Li YP. TGF-β and BMP signaling in osteoblast differentiation and bone formation. Int J Biol Sci. 2012;8:272. Google Scholar
  27. 27.
    Yang F, Wang J, Hou J, Guo H, Liu C. Bone regeneration using cell-mediated responsive degradable PEG-based scaffolds incorporating with rhBMP-2. Biomaterials. 2013;34:1514. Google Scholar
  28. 28.
    Athanasiou VT, Papachristou DJ, Panagopoulos A, Saridis A, Scopa CD, Megas P. Histological comparison of autograft, allograft-DBM, xenograft, and synthetic grafts in a trabecular bone defect: an experimental study in rabbits. Med Sci Monit. 2010;16:Br24.Google Scholar
  29. 29.
    Marom R, Shur I, Solomon R, Benayahu D. Characterization of adhesion and differentiation markers of osteogenic marrow stromal cells. J Cell Physiol. 2005;202:41. Google Scholar
  30. 30.
    Stucki U, Schmid J, Hammerle CF, Lang NP. Temporal and local appearance of alkaline phosphatase activity in early stages of guided bone regeneration. A descriptive histochemical study in humans. Clin Oral Implants Res. 2001;12:121.Google Scholar
  31. 31.
    Bigham AS, Dehghani SN, Shafiei Z, Torabi Nezhad S. Xenogenic demineralized bone matrix and fresh autogenous cortical bone effects on experimental bone healing: radiological, histopathological and biomechanical evaluation. J Orthop Traumatol. 2008;9:73. Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of OrthopaedicsJiangxi Provincial People’s Hospital Affiliated to Nanchang UniversityNanchangChina
  2. 2.Department of Clinincal MedicineHe UniversityShenyangChina
  3. 3.Department of Joint SurgeryThe Affiliated Ganzhou Hospital of Nanchang UniversityGanzhouChina

Personalised recommendations