title: Human pseudoarthrosis tissue contains cells with osteogenic potential.
abstract: BACKGROUND
  Nonunion is a challenging problem that may occur after certain bone fractures. The treatment of nonunion is closely related to its type. To develop an effective treatment strategy for each type of nonunion, biological analysis of nonunion tissue is essential. Pseudoarthrosis is a distinct pathologic entity of nonunion. To understand the pathology of pseudoarthrosis, we investigated the cellular properties of pseudoarthrosis tissue-derived cells (PCs) in vitro.
  
  
  METHODS
  PCs were isolated from four patients with pseudoarthrosis and cultured. Cells were evaluated for cell-surface protein expression by using flow cytometry. Osteogenic differentiation capacity was assessed by using Alizarin Red S staining, alkaline phosphatase (ALP) activity assay, and reverse transcription polymerase chain reaction (RT-PCR) after osteogenic induction. Chondrogenic differentiation capacity was assessed via Safranin O staining and RT-PCR after chondrogenic induction.
  
  
  RESULTS
  PCs were consistently positive for the mesenchymal stem cell-related markers CD29, CD44, CD105, and CD166, but were negative for the haematopoietic-lineage markers CD31, CD34, CD45, and CD133. Alizarin Red S staining revealed that PCs formed a mineralised matrix that was rich in calcium deposits after osteogenic induction. ALP activity under osteogenic conditions was significantly higher than that under control conditions. Gene expression of ALP, Runx2, osterix, osteocalcin, and bone sialoprotein was observed in PCs cultured under osteogenic conditions. Induced pellets were negatively stained by Safranin O staining. Gene expression of aggrecan, collagen II, collagen X, SOX5, and SOX9 was not observed.
  
  
  CONCLUSIONS
  We have shown for the first time the properties of cells in patients with pseudoarthrosis. Our results indicated that osteogenic cells existed in the pseudoarthrosis tissue. This study might provide insights into understanding the pathology of pseudoarthrosis and improving the treatment for pseudoarthrosis.
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Takahara
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Japan. Electronic address: tniikura@med.kobe-u.ac.jp.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Niikura
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Japan.
authors.forename: Sang Yang
authors.initials: SY
authors.lastname: Lee
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Iwakura
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Japan.
authors.forename: Etsuko
authors.initials: E
authors.lastname: Okumachi
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Japan.
authors.forename: Ryosuke
authors.initials: R
authors.lastname: Kuroda
authors.affiliation: Department of Orthopaedic Surgery, Kobe University Graduate School of Medicine, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Kurosaka
mesh_terms: D000368:Aged; D001842:Bone and Bones / Q000166:cytology*; D002113:Calcification, Physiologic / Q000502:physiology*; D049109:Cell Proliferation; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000502:physiology*; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D008297:Male; D008875:Middle Aged; D015675:Osteocalcin / Q000378:metabolism*; D010012:Osteogenesis / Q000502:physiology*; D011542:Pseudarthrosis / Q000378:metabolism / Q000473:pathology*; D023822:Tissue Engineering
chemical_list: D015675:Osteocalcin
keywords: Fracture healing; Nonunion; Osteogenic potential; Pseudoarthrosis