title: Osteogenic potential of rhBMP9 combined with a bovine-derived natural bone mineral scaffold compared to rhBMP2.
abstract: OBJECTIVE
  Combination therapies of growth factors and scaffolds for bone tissue engineering are becoming routine for clinical use. BMP9 has previously been characterized as one of the most osteogenic inducers among the BMP superfamily; however, up until recently, BMP9 has only been available through adenovirus transfection experiments (gene therapy). While recombinant human (rh)BMP2 is regarded as the gold standard for bone regeneration with recombinant growth factors, recently the successful development of rhBMP9 brings intriguing new possibilities for future clinical use. The purpose of this pioneering study was to investigate the effects of rhBMP9 in comparison with rhBMP2 on an in vitro cell behavior of bone-forming osteoblasts when combined with a bone grafting material.
  
  
  METHODS
  Undifferentiated mouse ST2 stromal bone marrow cells were seeded onto bovine-derived natural bone mineral (NBM) particles treated with (i) control, (ii) rhBMP2 (10 ng/ml), (iii) rhBMP2 (100 ng/ml), (iv) rhBMP9 (10 ng/ml) and (v) rhBMP9 (100 ng/ml). The effects of rhBMPs were compared for cell adhesion at 8 h, cell proliferation at 1, 3 and 5 days and osteoblast differentiation as assessed by real-time PCR at 3 and 14 days for genes encoding Runx2, collagen1alpha2 (COL1a2), alkaline phosphatase (ALP) and osteocalcin (OCN). Furthermore, ALP staining and alizarin red staining were used to investigate localization of osteoblast differentiation marker and mineralization on NBM.
  
  
  RESULTS
  Although neither rhBMP2 nor rhBMP9 influenced cell attachment to NBM particles, both were able to stimulate cell proliferation at 3 days. Furthermore, all concentrations of rhBMPs were able to significantly induce mRNA levels of Runx2, COL1a2 and OCN at 3 days. Interestingly, only rhBMP9 was able to significantly upregulate mRNA levels of ALP up to eightfold, and ALP staining up to 25-fold, when compared to rhBMP2. In addition, only rhBMP9 (100 ng/ml) significantly increased alizarin red staining when compared to control and rhBMP2 (10 ng/ml) samples.
  
  
  CONCLUSIONS
  These results demonstrate that both rhBMP2 and rhBMP9 have osteopromotive properties on osteoblast differentiation. It was found that rhBMP9 additionally stimulated the osteopromotive potential of osteoblasts when compared to rhBMP2 by demonstrating higher levels of ALP expression and alizarin red staining. Further animal studies comparing both recombinant proteins are necessary to further characterize the osteoinductive potential of BMP9.
authors.affiliation: Department of Cranio-Maxillofacial Surgery, Bern University Hospital, Inselspital, Bern, Switzerland.|Masako Fujioka-Kobayashi, Department of Oral Surgery, Clinical Dentistry, Institute of Biomedical Sciences, Tokushima University Graduate School, Tokushima, Japan.
authors.forename: Masako
authors.initials: M
authors.lastname: Fujioka-Kobayashi
authors.affiliation: Department of Cranio-Maxillofacial Surgery, Bern University Hospital, Inselspital, Bern, Switzerland.|Kosaku Sawada, Advanced Research Center, School of Life Dentistry at Niigata, The Nippon Dental University, Niigata, Japan.
authors.forename: Kosaku
authors.initials: K
authors.lastname: Sawada
authors.affiliation: Department of Cranio-Maxillofacial Surgery, Bern University Hospital, Inselspital, Bern, Switzerland.
authors.forename: Eizaburo
authors.initials: E
authors.lastname: Kobayashi
authors.affiliation: Department of Cranio-Maxillofacial Surgery, Bern University Hospital, Inselspital, Bern, Switzerland.
authors.forename: Benoit
authors.initials: B
authors.lastname: Schaller
authors.affiliation: Department of Oral Implantology, University of Wuhan, Wuhan, China.
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Preventive, Restorative and Pediatric Dentistry, University of Bern, Bern, Switzerland.|Department of Periodontology, Nova Southeastern University, Fort Lauderdale, FL, USA.
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Miron
mesh_terms: D000818:Animals; D040761:Biomimetic Materials; D055396:Bone Morphogenetic Protein 2 / Q000494:pharmacology*; D002113:Calcification, Physiologic / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D055427:Growth Differentiation Factor 2; D055412:Growth Differentiation Factors / Q000494:pharmacology*; D049673:History, 20th Century; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D010006:Osteoblasts / Q000187:drug effects; D010012:Osteogenesis / Q000187:drug effects*; D011994:Recombinant Proteins / Q000494:pharmacology*; D023822:Tissue Engineering
chemical_list: C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; C525887:GDF2 protein, human; D055427:Growth Differentiation Factor 2; D055412:Growth Differentiation Factors; D011994:Recombinant Proteins
keywords: bone formation; bone induction; bone morphogenetic protein; growth factor; guided bone regeneration; osteoinduction; osteoinductive