Isolation and Characterization of Multipotent Postnatal Stem Cells from the Alveolar Bone and Gingival Cervical Margin

Periodontitis is a destructive inflammatory disorder of the periodontium branded by the destruction of periodontal tissues namely the PDL, cementum, alveolar bone, and gingiva. Once these tissues are lost, the foremost goal of periodontal therapy is to regenerate the diseased tissues if possible to their original form, architecture, and function. This a demanding task, that requires the harmonization of many actions at both cellular and molecular levels. Successful periodontal regeneration encompasses the formation of new gingival connective tissue, the restoration of bone, and most decisively, the insertion of new connective tissue fibres into newly formed cementum on the formerly diseased root surfaces. A variety of procedures to regenerate the lost tissue have been suggested, including the utilization of the principles of guided tissue regeneration and the application of an assortment of growth factors as well as bone and enamel matrix proteins on the root surfaces. However, these strategies showed only limited regeneration of periodontal tissue and were associated with incoherent and unpredictable clinical outcomes. Stem cells are progenitor cells characterized by their ability to self-renew and differentiate to produce specialized cells. There are two main categories of stem cells, the pluripotent embryonic stem cells and the multipotent adult ones. Recently, dental tissues such as the PDL, the dental pulp and the tooth follicle have been recognized as readily available sources of adult stem cells. Employing the criteria defined by Bartold et al. (2006) as well as the minimal standards accepted by the International Society for Cellular Therapy (Dominici et al. 2006) to define human MSCs and bearing in mind that the PDL shares the same developmental path with the cervical gingival margin and alveolar bone, the aim of this study was to present two protocols for the isolation and characterization of populations of mesenchymal multipotent stem cells-periodontal stem cells (PDSCs) derived from the alveolar bone and the cervical gingival margin of the periodontium. In this study the sample consisted of eleven partially impacted wisdom teeth, which were surgically extracted together with a piece of alveolar bone and cervical gingival margin attached to them. Cells isolated from the alveolar bone and the cervical gingival margin of the periodontium showed adherence to tissue culture plastics under standard in vitro culture conditions. They both exhibited the ability to form colony-forming units and were highly positive for the surface markers CD73, CD90, CD105 (> 95%), moderately positive for the markers CD146 and STRO-1, and negative for CD14, CD34, and CD45. On the mRNA level unstimulated cells from both sources expressed the osteogenic markers alkaline phosphatase, type I and III collagen, osteonectin, osteopontin, and osteocalcin. In addition, they both expressed type V collagen. Finally, cells from both sources demonstrated in vitro multilineage differentiation potential as evidenced by histological staining and PCR assays. It is concluded from this study that adult postnatal multipotent PDSCs can be readily isolated from the alveolar bone as well as from the cervical gingival margin.


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