title: Cobalt chloride supplementation induces stem-cell marker expression and inhibits osteoblastic differentiation in human periodontal ligament cells.
abstract: OBJECTIVE
  Low oxygen tension is one of the crucial factors of the stem-cell niche. However, the long-term hypoxic culture of stem cells is difficult and requires special equipment. In this study, we investigated whether mimicking hypoxia using cobalt chloride (CoCl2) could maintain human periodontal ligament (HPDL) cell stemness.
  
  
  METHODS
  HPDL cells were treated with either 50 or 100 μM CoCl2. Cell proliferation was determined by an MTT assay. The mRNA expression of stem-cell marker and osteogenic associated genes were analyzed by RT-PCR and Real-time PCR. Osteogenic differentiation was determined by assaying alkaline phosphatase activity and in vitro mineralization.
  
  
  RESULTS
  The results showed that the CoCl2 supplementation had no effect on cell proliferation. CoCl2 treatment increased the mRNA expression of the embryonic stem-cell markers REX1 and OCT4. Culturing HDPL cells in osteogenic medium containing CoCl2 resulted in a decrease in alkaline phosphatase activity, down-regulation of osteogenic associated gene expression, and suppression of mineralization. The use of Apigenin, an HIF-1α inhibitor, indicated that CoCl2 might inhibit osteogenic differentiation through an HIF-1α- dependent mechanism.
  
  
  CONCLUSIONS
  This study shows that CoCl2 treatment can induce stem-cell marker expression and inhibit the osteoblastic differentiation of HPDL cells. These findings suggest the potential application of CoCl2 for maintaining the stem-cell state in the laboratory.
authors.affiliation: Department of Anatomy, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Developing Research Unit in Genetic and Craniofacial Analyses of Craniofacial Structures, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Thanaphum
authors.initials: T
authors.lastname: Osathanon
authors.affiliation: Department of Anatomy, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Developing Research Unit in Genetic and Craniofacial Analyses of Craniofacial Structures, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Philaiporn
authors.initials: P
authors.lastname: Vivatbutsiri
authors.affiliation: Department of Pediatric Dentistry, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Developing Research Unit in Tissue Engineering, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Waleerat
authors.initials: W
authors.lastname: Sukarawan
authors.affiliation: Department of Pediatric Dentistry, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Developing Research Unit in Cell Signaling and Protein Function, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Wannakorn
authors.initials: W
authors.lastname: Sriarj
authors.affiliation: Department of Anatomy, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Prasit
authors.initials: P
authors.lastname: Pavasant
authors.affiliation: Developing Research Unit in Cell Signaling and Protein Function, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand; Department of Pharmacology, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand. Electronic address: pl_sireerat@yahoo.com.
authors.forename: Sireerat
authors.initials: S
authors.lastname: Sooampon
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003035:Cobalt / Q000494:pharmacology*; D005434:Flow Cytometry; D006801:Humans; D010012:Osteogenesis / Q000187:drug effects*; D010513:Periodontal Ligament / Q000166:cytology*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000378:metabolism*
chemical_list: D012333:RNA, Messenger; D003035:Cobalt; D000469:Alkaline Phosphatase; C018021:cobaltous chloride
keywords: Cobalt chloride; Human periodontal ligament cells; Osteoblastic differentiation; Stem cells