Surface functionalization of TiO2 nanotubes with bone morphogenetic protein 2 and its synergistic effect on the differentiation of mesenchymal stem cells.

To investigate the influence of surface-functionalized substrates with nanostructures on the behaviors of mesenchymal stem cells, we conjugated bone morphogenetic protein 2 (BMP2) onto TiO(2) nanotubes with different diameter sizes of 30, 60, and 100 nm for in vitro study. Polydopamine was employed as the intermediate layer for the conjugation of BMP2. The successful conjugation of BMP2 onto TiO(2) nanotubes was revealed by field-emission scanning electron microscopy (FE-SEM), X-ray photoelectron spectroscopy (XPS), and contact angle measurements. Immunofluorescence staining of vinculin, osteocalcin (OCN), and osteopontin (OPN) revealed that BMP2-functionalized TiO(2) nanotubes was favorable for cell growth. More importantly, MSCs cultured onto BMP2-functionalized TiO(2) nanotubes displayed significantly higher (p < 0.05 or p < 0.01) differentiation levels of ALP and mineralization after 7 and 14 day cultures, respectively. The results suggested that surface functionalization of TiO(2) nanotubes with BMP2 was beneficial for cell proliferation and differentiation. The approach presented here has potential application for the development of titanium-based implants for enhanced bone osseointegration.

[1]  K. Miyazono,et al.  Interaction and Functional Cooperation of PEBP2/CBF with Smads , 1999, The Journal of Biological Chemistry.

[2]  Joachim P Spatz,et al.  Impact of order and disorder in RGD nanopatterns on cell adhesion. , 2009, Nano letters.

[3]  Li Yang,et al.  Surface mediated in situ differentiation of mesenchymal stem cells on gene-functionalized titanium films fabricated by layer-by-layer technique. , 2009, Biomaterials.

[4]  G S Stein,et al.  Molecular mechanisms mediating proliferation/differentiation interrelationships during progressive development of the osteoblast phenotype. , 1993, Endocrine reviews.

[5]  Patrik Schmuki,et al.  TiO2 nanotube surfaces: 15 nm--an optimal length scale of surface topography for cell adhesion and differentiation. , 2009, Small.

[6]  C. Azevedo.  Failure analysis of a commercially pure titanium plate for osteosynthesis , 2003 .

[7]  Christoph Ballestrem,et al.  Vinculin controls focal adhesion formation by direct interactions with talin and actin , 2007, The Journal of cell biology.

[8]  M. Yoshinari,et al.  An immunoelectron microscopic localization of noncollagenous bone proteins (osteocalcin and osteopontin) at the bone-titanium interface of rat tibiae. , 1998, Journal of biomedical materials research.

[9]  P. Giannoudis,et al.  Mesenchymal stem cell tissue engineering: techniques for isolation, expansion and application. , 2007, Injury.

[10]  K. Anseth,et al.  Multifunctional Hydrogels that Promote Osteogenic Human Mesenchymal Stem Cell Differentiation Through Stimulation and Sequestering of Bone Morphogenic Protein 2 , 2007 .

[11]  Haeshin Lee,et al.  Mussel-Inspired Surface Chemistry for Multifunctional Coatings , 2007, Science.

[12]  J. Macák,et al.  Mechanistic aspects and growth of large diameter self-organized TiO2 nanotubes , 2008 .

[13]  Vladimir Volloch,et al.  Bone morphogenetic protein-2 decorated silk fibroin films induce osteogenic differentiation of human bone marrow stromal cells. , 2004, Journal of biomedical materials research. Part A.

[14]  K. Miyazono.  Signal transduction by bone morphogenetic protein receptors: functional roles of Smad proteins. , 1999, Bone.

[15]  A. Bloch-Zupan,et al.  Delayed Osteoprogenitor Differentiation in Cleft-Palate Models , 2010, Cells Tissues Organs.

[16]  Eugeniu Balaur,et al.  Tailoring the wettability of TiO2 nanotube layers , 2005 .

[17]  Julian H. George,et al.  Exploring and Engineering the Cell Surface Interface , 2005, Science.

[18]  P. Rubenstein,et al.  Vinculin Nucleates Actin Polymerization and Modifies Actin Filament Structure* , 2009, The Journal of Biological Chemistry.

[19]  Y. Iwakura,et al.  Periprosthetic osteolysis: Characterizing the innate immune response to titanium wear‐particles , 2010, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[20]  Haeshin Lee,et al.  Facile Conjugation of Biomolecules onto Surfaces via Mussel Adhesive Protein Inspired Coatings , 2009, Advanced materials.

[21]  B. Nebe,et al.  Cell architecture-cell function dependencies on titanium arrays with regular geometry. , 2010, Biomaterials.

[22]  K. Popat,et al.  Template synthesized poly(epsilon-caprolactone) nanowire surfaces for neural tissue engineering. , 2010, Biomaterials.

[23]  Michael Müller,et al.  Surface structure and composition of flat titanium thin films as a function of film thickness and evaporation rate , 2005 .

[24]  T. Tsukazaki,et al.  BMP‐2 promotes differentiation of osteoblasts and chondroblasts in Runx2‐deficient cell lines , 2007, Journal of cellular physiology.

[25]  A L Boskey,et al.  Mineral-matrix interactions in bone and cartilage. , 1992, Clinical orthopaedics and related research.

[26]  K. Yao,et al.  Poly(D,L-lactic acid) surfaces modified by silk fibroin: effects on the culture of osteoblast in vitro. , 2002, Biomaterials.

[27]  Sang Hoon Lee,et al.  Bone regeneration using hyaluronic acid-based hydrogel with bone morphogenic protein-2 and human mesenchymal stem cells. , 2007, Biomaterials.

[28]  R. Tuan,et al.  Mechanism of BMP‐2 stimulated adhesion of osteoblastic cells to titanium alloy , 1999, Biology of the cell.

[29]  Jeffrey L. Wrana,et al.  Mechanism of activation of the TGF-β receptor , 1994, Nature.

[30]  G. Im,et al.  Electroporation-mediated transfer of Runx2 and Osterix genes to enhance osteogenesis of adipose stem cells. , 2011, Biomaterials.

[31]  Sungho Jin,et al.  Stem cell fate dictated solely by altered nanotube dimension , 2009, Proceedings of the National Academy of Sciences.

[32]  C. Wilkinson,et al.  The control of human mesenchymal cell differentiation using nanoscale symmetry and disorder. , 2007, Nature materials.

[33]  P. Dijke.  Bone morphogenetic protein signal transduction in bone , 2006 .

[34]  M. Carstens,et al.  In Situ Osteogenesis: Regeneration of 10-cm Mandibular Defect in Porcine Model Using Recombinant Human Bone Morphogenetic Protein-2 (rhBMP-2) and Helistat Absorbable Collagen Sponge , 2005, The Journal of craniofacial surgery.

[35]  P. Ducheyne,et al.  Apoptosis and Survival of Osteoblast-like Cells Are Regulated by Surface Attachment* , 2005, Journal of Biological Chemistry.

[36]  D W Hutmacher,et al.  The effect of rhBMP-2 on canine osteoblasts seeded onto 3D bioactive polycaprolactone scaffolds. , 2004, Biomaterials.

[37]  C. Colnot,et al.  Bone morphogenetic protein 2 stimulates endochondral ossification by regulating periosteal cell fate during bone repair. , 2010, Bone.

[38]  Kozo Akino,et al.  Bone morphogenetic protein‐2 regulates proliferation of human mesenchymal stem cells , 2003, Wound repair and regeneration : official publication of the Wound Healing Society [and] the European Tissue Repair Society.

[39]  M. Textor,et al.  Chemically patterned, metal oxide based surfaces produced by photolithographic techniques for studying protein- and cell-surface interactions I: Microfabrication and surface characterization. , 2003, Biomaterials.

[40]  K. Neoh,et al.  The effect of VEGF functionalization of titanium on endothelial cells in vitro. , 2010, Biomaterials.

[41]  C. Whang,et al.  Titanium oxide films on Si(100) deposited by electron-beam evaporation at 250 °C , 2000 .

[42]  T. Goto,et al.  Effects of Static Magnetic Fields on Bone Formation in Rat Osteoblast Cultures , 2003, Journal of dental research.

[43]  Kam W Leong,et al.  Nanopattern-induced changes in morphology and motility of smooth muscle cells. , 2005, Biomaterials.

[44]  Jingyuan Fu,et al.  Signal Transduction of Bone Morphogenetic Proteins in Osteoblast Differentiation , 2003, The Journal of bone and joint surgery. American volume.

[45]  Juan Luo,et al.  Surface modification of poly (D,L-lactic acid) with chitosan and its effects on the culture of osteoblasts in vitro. , 2002, Journal of biomedical materials research.

[46]  Patrik Schmuki,et al.  Nanosize and vitality: TiO2 nanotube diameter directs cell fate. , 2007, Nano letters.

[47]  Marc Aucouturier,et al.  Anodic oxidation of titanium and TA6V alloy in chromic media. An electrochemical approach , 1999 .

[48]  C. Dunning,et al.  Bone stresses before and after insertion of two commercially available distal ulnar implants using finite element analysis , 2011, Journal of orthopaedic research : official publication of the Orthopaedic Research Society.

[49]  J. Waite.  Mussel power. , 2008, Nature materials.