title: [A novel tissue-engineered bone constructed by using human adipose-derived stem cells and biomimetic calcium phosphate scaffold coprecipitated with bone morphogenetic protein-2].
abstract: To construct a novel biomimetic calcium phosphate (BioCaP) scaffold loaded with bone morphogenetic protein-2 (BMP-2), and to investigate its role in the osteogenesis of human adipose-derived stem cells (hASCs) in vitro and in vivo.
  
  
  
  The BioCaP scaffold coprecipitated with BMP-2 (BMP-2-BioCaP) was constructed in this study. Field emission scanning electron microscopy (SEM) was used to analyze the morphology of the surfaces. The release kinetics was measured to evaluate the slow-release characteristics in vitro. BMP-2-BioCaP was immersed in proliferation medium (PM) or osteogenic medium (OM), respectively. The supernatants were collected and used to culture hASCs in vitro. Cell numbers were determined using the cell-counting kit-8 (CCK-8) to assess the cell proliferation. After 7 and 14 days, alkaline phosphatase (ALP) staining and quantification were performed to test the activity of ALP. After 14 and 21 days, the calcification deposition was determined by alizarin red S (ARS) staining and quantification. The expressions of the osteoblast-related genes were tested on day 4 and day 14. In the in vivo study, 6 nude mice were used and implanted subcutaneously into the back of the nude mice for 4 groups: (1) BioCaP scaffold only, (2) BioCaP scaffold+hASCs, (3) BMP-2-BioCaP scaffold, (4) BMP-2-BioCaP scaffold+hASCs (test group). After 4 weeks of implantation, hematoxylin-eosin (HE) staining was performed to evaluate the in vivo osteogenesis of hASCs.
  
  
  
  SEM observations showed that BioCaP and BMP-2-BioCaP scaffold were entirely composed of straight, plate-like and sharp-edged crystal units, and the length of the crystal units varied between 5 and 10 μm. Release kinetics analysis demonstrated that BMP-2 incorporated with BioCaP could be released at certain concentration and last for more than 21 days, and the accumulative protein release could reach 20%. CCK-8 assays showed that cell proliferation was not significantly affected by BMP-2-BioCaP. ALP activity was higher by the induction of OM+BMP-2-BioCaP than of the other groups (P<0.01). More mineralization deposition and more expressions of osteoblast-related genes such as Runt-related transcription factor 2 (RUNX2), ALP, osteopontin (OPN) and osteocalcin (OC) were determined in the OM+BMP-2-BioCaP group at different time points (P<0.01). HE staining showed that, in the test group and BMP-2-BioCaP scaffold group, the extracellular matrix (ECM) with eosinophilic staining were observed around hASCs, and newly-formed bone-like tissues could be found in ECM around the scaffold materials. Moreover, compared with the BMP-2-BioCaP scaffold group, more bone-like tissues could be observed in ECM with typical structure of bone tissue in the test groups. No obvious positive results were found in the other groups.
  
  
  
  BMP-2-BioCaP scaffold could achieve slow-release of BMP-2 and promote the osteogenic differentiation of hASCs in vitro and in vivo. The novel tissue-engineered bone composed of hASCs and BMP-2-BioCaPis promising for the repair of bone defect.
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, Beijing 100081, China.
authors.forename: W R
authors.initials: WR
authors.lastname: Jiang
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, Beijing 100081, China.
authors.forename: X
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, Beijing 100081, China.
authors.forename: Y S
authors.initials: YS
authors.lastname: Liu
authors.affiliation: Department of Oral Implantology and Prosthetic Dentistry, Academic Centre for Dentistry Amsterdam (ACTA), Research Institute MOVE, VU University and University of Amsterdam, Amsterdam, the Netherlands 1081 LA.
authors.forename: G
authors.initials: G
authors.lastname: Wu
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, Beijing 100081, China.
authors.forename: Y J
authors.initials: YJ
authors.lastname: Ge
authors.affiliation: Department of Prosthodontics, Peking University School and Hospital of Stomatology & National Engineering Laboratory for Digital and Material Technology of Stomatology & Beijing Key Laboratory of Digital Stomatology, Beijing 100081, China.
authors.forename: Y S
authors.initials: YS
authors.lastname: Zhou
mesh_terms: D000273:Adipose Tissue; D000469:Alkaline Phosphatase / Q000187:drug effects / Q000378:metabolism; D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry / Q000494:pharmacology; D040761:Biomimetic Materials / Q000737:chemistry / Q000494:pharmacology; D055396:Bone Morphogenetic Protein 2 / Q000008:administration & dosage / Q000493:pharmacokinetics* / Q000494:pharmacology* / Q000627:therapeutic use; D001842:Bone and Bones; D002113:Calcification, Physiologic / Q000187:drug effects*; D002130:Calcium Phosphates / Q000008:administration & dosage / Q000493:pharmacokinetics* / Q000494:pharmacology* / Q000627:therapeutic use; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured / Q000166:cytology / Q000187:drug effects; D050659:Core Binding Factor Alpha 1 Subunit / Q000187:drug effects / Q000378:metabolism; D065546:Drug Liberation / Q000187:drug effects*; D005109:Extracellular Matrix / Q000187:drug effects / Q000378:metabolism; D064593:Heterografts / Q000737:chemistry / Q000502:physiology / Q000637:transplantation; D006801:Humans; D045164:Mesenchymal Stem Cell Transplantation / Q000379:methods*; D059630:Mesenchymal Stem Cells / Q000187:drug effects; D051379:Mice; D008819:Mice, Nude; D008855:Microscopy, Electron, Scanning; D015675:Osteocalcin / Q000187:drug effects / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects*; D053495:Osteopontin / Q000187:drug effects / Q000378:metabolism; D013499:Surface Properties; D023822:Tissue Engineering / Q000379:methods*; D054457:Tissue Scaffolds / Q000737:chemistry
chemical_list: C525718:BMP2 protein, human; D001672:Biocompatible Materials; D055396:Bone Morphogenetic Protein 2; D002130:Calcium Phosphates; D050659:Core Binding Factor Alpha 1 Subunit; D015675:Osteocalcin; D053495:Osteopontin; D000469:Alkaline Phosphatase