Strontium attenuates rhBMP-2-induced osteogenic differentiation via formation of Sr-rhBMP-2 complex and suppression of Smad-dependent signaling pathway

Wenjing Zhang; Yu Tian; Hongyan He; Rui Chen; Yifan Ma; Han Guo; Yuan Yuan; Changsheng Liu

doi:10.1016/j.actbio.2016.01.042

Strontium attenuates rhBMP-2-induced osteogenic differentiation via formation of Sr-rhBMP-2 complex and suppression of Smad-dependent signaling pathway

Acta Biomater. 2016 Mar:33:290-300. doi: 10.1016/j.actbio.2016.01.042. Epub 2016 Jan 29.

Authors

Wenjing Zhang¹, Yu Tian¹, Hongyan He², Rui Chen³, Yifan Ma⁴, Han Guo⁵, Yuan Yuan⁶, Changsheng Liu⁷

Affiliations

¹ The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, PR China; Key Laboratory for Ultrafine Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237, PR China.
² Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237, PR China.
³ The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, PR China; Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237, PR China.
⁴ The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, PR China.
⁵ Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, PR China.
⁶ The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, PR China; Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237, PR China. Electronic address: yyuan@ecust.edu.cn.
⁷ The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, PR China; Key Laboratory for Ultrafine Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237, PR China; Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237, PR China. Electronic address: liucs@ecust.edu.cn.

PMID: 26828127
DOI: 10.1016/j.actbio.2016.01.042

Abstract

Strontium (Sr(2+)) has pronounced effects on stimulating bone formation and inhibiting bone resorption in bone regeneration. In this current study, the effect and the underlying mechanism involved of Sr(2+) on the biological activity of bone morphogenetic protein-2 (BMP-2) were studied in detail with pluripotent skeletal muscle myogenic progenitor C2C12 model cell line. The results indicated that Sr(2+) could bind recombinant human BMP-2 (rhBMP-2) rapidly, even in the presence of Ca(2+) and Mg(2+), and inhibited rhBMP-2-induced osteogenic differentiation in vitro and osteogenetic efficiency in vivo. Further studies demonstrated that Sr(2+) treatment undermined the binding capacity of rhBMP-2 with its receptor BMPRIA and thus attenuated Smad 1/5/8 phosphorylation without affecting their dephosphorylation in C2C12 cells. Furthermore, circular dichroism spectroscopy, fluorescence spectroscopy and X-ray photoelectron spectroscopy all revealed that the inhibitory effect of Sr(2+) on the rhBMP-2 osteogenic activity was associated with the formation of Sr-rhBMP-2 complex and ensuing enhancement of β-sheet structure. Our work suggests the activity of rhBMP-2 to induce osteogenic differentiation was decreased by directly interaction with free Sr ions in solution, which should provide guide and assist for development of BMP-2-based materials for bone regeneration.

Statement of significance: Due to easy denaturation and ensuing the reduced activity of rhBMP-2, preserving/enhancing the capacity of rhBMP-2 to induce osteogenic differentiation is of critical importance in developing the protein-based therapy. Cations as effective elements influence the conformation and thereby the bioactivity of protein. Strontium (Sr(2+)), stimulating bone formation and inhibiting bone resorption, has been incorporated into biomaterials/scaffold to improve the bioactivity for bone-regeneration applications. However, Sr(2+)-induced changes in the conformation and bioactivity of BMP-2 have never been investigated. In this study, the formation of Sr-rhBMP-2 complex inhibited the osteogenic differentiation in vitro and osteogenetic efficiency in vivo through the inhibition of BMP/Smad signaling pathway, providing guidance for development of Sr-containing BMP-2-based bone scaffold/matrice and other Sr-dopped protein therapy.

Keywords: Bone formation; Sr ions; Sr-rhBMP-2 complex; rhBMP-2.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Blotting, Western
Bone Morphogenetic Protein 2 / chemistry
Bone Morphogenetic Protein 2 / pharmacology*
Bone Morphogenetic Protein Receptors / metabolism
Cell Differentiation / drug effects*
Cell Line
Cell Membrane / drug effects
Cell Membrane / metabolism
Choristoma / pathology
Humans
Ions
Mice
Models, Biological
Osteogenesis / drug effects*
Protein Structure, Secondary
Recombinant Proteins / chemistry
Recombinant Proteins / pharmacology
Signal Transduction / drug effects*
Smad Proteins / metabolism*
Strontium / pharmacology*
Thermodynamics
Transforming Growth Factor beta / chemistry
Transforming Growth Factor beta / pharmacology*

Substances

Bone Morphogenetic Protein 2
Ions
Recombinant Proteins
Smad Proteins
Transforming Growth Factor beta
recombinant human bone morphogenetic protein-2
Bone Morphogenetic Protein Receptors
Strontium