title: Isolation and characterization of human mesenchymal stem cells from gingival connective tissue.
abstract: OBJECTIVE
  The main purpose of this study was to isolate and characterize gingival connective tissue-derived mesenchymal stem cells (GMSCs). The secondary purpose was to present a modified isolation method for the GMSCs.
  
  
  METHODS
  Collected healthy gingival tissue samples were de-epithelialized and minced into small fragments. The tissues were digested by dispase and collagenase IV for 30 min. The first digested cell suspension was discarded, and then additional digestion was performed to the remaining cells in the same solution for 90 min. The isolated cells from gingiva was incubated in 37°C humidified condition and observed by inverted microscope. Cytoskeletal morphology was evaluated by phalloidin immunofluorescence. Potency of the cells was tested by colony-forming unit fibroblast assay. GMSCs were characterized by osteogenic, adipogenic and chondrogenic differentiation, and flow cytometric, immunofluorescence analysis.
  
  
  RESULTS
  GMSCs showed spindle-shaped, fibroblast-like morphology, colony-forming abilities, adherence to plastic and multilineage differentiation (osteogenic, adipogenic, chondrogenic) potency. GMSCs expressed CD44, CD73, CD90 and CD105, but did not express CD14, CD45, CD34 and CD19 in flow cytometry. Expression of stem cell markers (SSEA-4, STRO-1, CD146, CD166 and CD271) and a mesenchymal marker (vimentin) were observed by immunofluorescence.
  
  
  CONCLUSIONS
  In conclusion, we isolated and characterized stem cells from human gingival connective tissue with modified protocol. GMSCs showed multipotency with high proliferation and characteristics of mesenchymal stem cells. GMSCs are promising sources for tissue engineering and may be obtained during routine procedures under local anesthesia. Further research is needed to evaluate the potential of GSMCs' proliferation and cryopreservation.
authors.affiliation: Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: S H
authors.initials: SH
authors.lastname: Jin
authors.affiliation: Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: J E
authors.initials: JE
authors.lastname: Lee
authors.affiliation: Division of Periodontology, Department of Dentistry, School of Medicine, Inha University, Incheon, Korea.
authors.forename: J-H
authors.initials: JH
authors.lastname: Yun
authors.affiliation: Division of Oral and Maxillofacial Surgery, Department of Dentistry, Uijeongbu St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Uijeongbu, Korea.
authors.forename: I
authors.initials: I
authors.lastname: Kim
authors.affiliation: Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: Y
authors.identifier: http://orcid.org/0000-0002-6564-9156
authors.initials: Y
authors.lastname: Ko
authors.affiliation: Department of Periodontics, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: J B
authors.initials: JB
authors.lastname: Park
mesh_terms: D015720:5'-Nucleotidase / Q000032:analysis; D050156:Adipogenesis / Q000502:physiology; D015703:Antigens, CD / Q000032:analysis; D000954:Antigens, Surface / Q000032:analysis; D051929:CD146 Antigen / Q000032:analysis; D002448:Cell Adhesion / Q000502:physiology; D015816:Cell Adhesion Molecules, Neuronal / Q000032:analysis; D002449:Cell Aggregation / Q000502:physiology; D002454:Cell Differentiation / Q000502:physiology; D002469:Cell Separation / Q000379:methods*; D048430:Cell Shape; D020219:Chondrogenesis / Q000502:physiology; D017364:Collagenases / Q000008:administration & dosage; D003239:Connective Tissue Cells / Q000166:cytology; D003599:Cytoskeleton / Q000648:ultrastructure; D000071063:Endoglin / Q000032:analysis; D010450:Endopeptidases / Q000008:administration & dosage; D005326:Fetal Proteins / Q000032:analysis; D005347:Fibroblasts / Q000166:cytology; D058851:GPI-Linked Proteins / Q000032:analysis; D005881:Gingiva / Q000166:cytology*; D006801:Humans; D018960:Hyaluronan Receptors / Q000032:analysis; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D039902:Multipotent Stem Cells / Q000166:cytology; D009419:Nerve Tissue Proteins / Q000032:analysis; D010012:Osteogenesis / Q000502:physiology; D017475:Receptors, Nerve Growth Factor / Q000032:analysis; D055746:Stage-Specific Embryonic Antigens / Q000032:analysis; D018800:Thy-1 Antigens / Q000032:analysis; D013997:Time Factors; D014746:Vimentin / Q000032:analysis; D020410:Activated-Leukocyte Cell Adhesion Molecule
chemical_list: D015720:5'-Nucleotidase; D015703:Antigens, CD; D000954:Antigens, Surface; D051929:CD146 Antigen; D015816:Cell Adhesion Molecules, Neuronal; D017364:Collagenases; D000071063:Endoglin; D010450:Endopeptidases; D005326:Fetal Proteins; D058851:GPI-Linked Proteins; D018960:Hyaluronan Receptors; D009419:Nerve Tissue Proteins; D017475:Receptors, Nerve Growth Factor; D055746:Stage-Specific Embryonic Antigens; D018800:Thy-1 Antigens; D014746:Vimentin; C105613:ALCAM protein, human; D020410:Activated-Leukocyte Cell Adhesion Molecule; C497483:CD44 protein, human; C064206:ENG protein, human; C497902:MCAM protein, human; C484065:NGFR protein, human; C525591:STRO-1 antigen, human; C066584:stage-specific embryonic antigen-4; C548241:NT5E protein, human; C023139:dispase
keywords: gingiva; mesenchymal stromal cells; multipotent stem cells; stem cells