title: Culturing bone marrow cells with dexamethasone and ascorbic acid improves osteogenic cell sheet structure.
abstract: OBJECTIVE
  To assess the structure and extracellular matrix molecule expression of osteogenic cell sheets created via culture in medium with both dexamethasone (Dex) and ascorbic acid phosphate (AscP) compared either Dex or AscP alone.
  
  
  METHODS
  Osteogenic cell sheets were prepared by culturing rat bone marrow stromal cells in a minimal essential medium (MEM), MEM with AscP, MEM with Dex, and MEM with Dex and AscP (Dex/AscP). The cell number and messenger (m)RNA expression were assessed in vitro, and the appearance of the cell sheets was observed after mechanical retrieval using a scraper. β-tricalcium phosphate (β-TCP) was then wrapped with the cell sheets from the four different groups and subcutaneously implanted into rats.
  
  
  RESULTS
  After mechanical retrieval, the osteogenic cell sheets from the MEM, MEM with AscP, and MEM with Dex groups appeared to be fragmented or incomplete structures. The cell sheets cultured with Dex/AscP remained intact after mechanical retrieval, without any identifiable tears. Culture with Dex/AscP increased the mRNA and protein expression of extracellular matrix proteins and cell number compared with those of the other three groups. More bridging bone formation was observed after transplantation of the β-TCP scaffold wrapped with cell sheets cultured with Dex/AscP, than in the other groups.
  
  
  CONCLUSIONS
  These results suggest that culture with Dex/AscP improves the mechanical integrity of the osteogenic cell sheets, allowing retrieval of the confluent cells in a single cell sheet structure. This method may be beneficial when applied in cases of difficult tissue reconstruction, such as nonunion, bone defects, and osteonecrosis.Cite this article: M. Akahane, T. Shimizu, T. Kira, T. Onishi, Y. Uchihara, T. Imamura, Y. Tanaka. Culturing bone marrow cells with dexamethasone and ascorbic acid improves osteogenic cell sheet structure. Bone Joint Res 2016;5:569-576. DOI: 10.1302/2046-3758.511.BJR-2016-0013.R1.
authors.affiliation: Department of Public Health, Health Management and Policy, Nara Medical University Faculty of Medicine, Kashihara, Nara, Japan makahane@naramed-u.ac.jp.
authors.forename: M
authors.initials: M
authors.lastname: Akahane
authors.affiliation: Department of Orthopedic Surgery, Nara Medical University Faculty of Medicine, Kashihara, Nara, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Shimizu
authors.affiliation: Department of Orthopedic Surgery, Nara Medical University Faculty of Medicine, Kashihara, Nara, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Kira
authors.affiliation: Department of Orthopedic Surgery, Nara Medical University Faculty of Medicine, Kashihara, Nara, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Onishi
authors.affiliation: Department of Orthopedic Surgery, Nara Medical University Faculty of Medicine, Kashihara, Nara, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Uchihara
authors.affiliation: Department of Public Health, Health Management and Policy, Nara Medical University Faculty of Medicine, Kashihara, Nara, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Imamura
authors.affiliation: Department of Orthopedic Surgery, Nara Medical University Faculty of Medicine, Kashihara, Nara, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Tanaka
keywords: Cell sheet; Mechanical retrieval; Osteogenesis