title: Optimizing the osteogenic differentiation of human mesenchymal stromal cells by the synergistic action of growth factors.
abstract: A variety of different growth factors, most notably bone morphogenetic proteins (BMPs), have been shown to stimulate the osteogenic differentiation of mesenchymal stromal cells (MSCs) in vitro. Yet, due to the lack of comparative studies it remains unclear which protocol is the most effective in the induction of osteogenesis in MSC cultures. The aim of this study was to compare the most potent growth factors in regard to their osteoinductive potential. Human MSCs were cultured for 10 days in the presence of BMP-2, BMP-6, BMP-9 + IGF-2 and BMP-2, -6, -9 (day 1 + 2: 50 ng/ml; days 3-6: 100 ng/ml; days 7-10: 200 ng/ml). The formation of the osteoblast phenotype was assessed by quantification of osteoblast-related marker genes using reverse transcription polymerase chain reaction (RT-PCR) and alkaline phosphatase (ALP) staining. Matrix mineralization was assessed by alizarin red S and von Kossa staining. Statistical analysis was carried out using the one-way analysis of variance (ANOVA) followed by Scheffe's post hoc procedure. Among the tested growth factors the combination of BMP-2 + BMP-6 + BMP-9 most effectively induced the upregulation of collagen type I, collagen type V, osteocalcin, alkaline phosphatase, RUNX2, BMP-2, osteonectin and DLX5 (p < 0.01) and resulted in a consistent matrix mineralization. The findings suggest the combined addition of BMP-2, BMP-6 and BMP-9 to the osteoinductive culture medium containing dexamethasone, β-glycerophosphate and ascorbate-2-phosphate produces more potent osteoblast differentiation of human MSCs in vitro.
authors.affiliation: Department of Oral & Maxillofacial Surgery, Christian-Albrechts-University, Kiel, Germany.
authors.forename: Yahya
authors.initials: Y
authors.lastname: Açil
authors.affiliation: Department of Oral & Maxillofacial Surgery, Christian-Albrechts-University, Kiel, Germany.
authors.forename: Amir-Alexander
authors.initials: AA
authors.lastname: Ghoniem
authors.affiliation: Department of Oral & Maxillofacial Surgery, Christian-Albrechts-University, Kiel, Germany.
authors.forename: Jörg
authors.initials: J
authors.lastname: Wiltfang
authors.affiliation: Department of Plastic Surgery, University of Lübeck, Lübeck, Germany. Electronic address: matthias_gierloff@icloud.com.
authors.forename: Matthias
authors.initials: M
authors.lastname: Gierloff
mesh_terms: D000469:Alkaline Phosphatase / Q000032:analysis; D001205:Ascorbic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D055396:Bone Morphogenetic Protein 2 / Q000494:pharmacology; D055418:Bone Morphogenetic Protein 6 / Q000494:pharmacology; D002113:Calcification, Physiologic / Q000187:drug effects; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D024042:Collagen Type I / Q000032:analysis; D024062:Collagen Type V / Q000032:analysis; D050659:Core Binding Factor Alpha 1 Subunit / Q000032:analysis; D003470:Culture Media; D003907:Dexamethasone / Q000494:pharmacology; D005994:Glycerophosphates / Q000494:pharmacology; D055427:Growth Differentiation Factor 2; D055412:Growth Differentiation Factors / Q000494:pharmacology; D018398:Homeodomain Proteins / Q000032:analysis; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology*; D059630:Mesenchymal Stem Cells / Q000187:drug effects*; D010006:Osteoblasts / Q000187:drug effects*; D015675:Osteocalcin / Q000032:analysis; D010012:Osteogenesis / Q000187:drug effects*; D015676:Osteonectin / Q000032:analysis; D010641:Phenotype; D014157:Transcription Factors / Q000032:analysis
chemical_list: C525718:BMP2 protein, human; C525816:BMP6 protein, human; D055396:Bone Morphogenetic Protein 2; D055418:Bone Morphogenetic Protein 6; D024042:Collagen Type I; D024062:Collagen Type V; D050659:Core Binding Factor Alpha 1 Subunit; D003470:Culture Media; C100596:DLX5 protein, human; C525887:GDF2 protein, human; D005994:Glycerophosphates; D055427:Growth Differentiation Factor 2; D055412:Growth Differentiation Factors; D018398:Homeodomain Proteins; C035170:IGF2 protein, human; D036341:Intercellular Signaling Peptides and Proteins; D015676:Osteonectin; C493709:RUNX2 protein, human; D014157:Transcription Factors; D015675:Osteocalcin; C011669:ascorbate-2-phosphate; D007335:Insulin-Like Growth Factor II; D003907:Dexamethasone; D000469:Alkaline Phosphatase; D001205:Ascorbic Acid; C031463:beta-glycerophosphoric acid
keywords: BMP; Bone tissue engineering; Growth factors; MSC; Osteogenic differentiation