Isolation and cellular properties of mesenchymal stem cells from human periosteum

Background: Cell-based therapy has achieved good functional recovery for tissue repair. Mesenchymal stem cells (MSCs) exhibit multilineage potential, long-term viability, and capacity for self-renewal. Periosteum-derived mesenchymal stem cells (PD cells) may be an attractive cell source for tissue engineering because of their easy accessibility and reduced ethical concerns.

Objectives: To isolate and investigate the phenotypic and functional characteristics of mesenchymal stem cells derived from human periosteum. We also examined the differentiation of PD cells with a trilineage differentiation assay to determine whether they were MSCs.

Materials and Methods: Periosteum-derived cells were cultured in osteogenic, chondrogenic or adipogenic media to evaluate their multilineage differentiation potential. Adherent fibroblast-like cells were analyzed by flow cytometry for MSC cell surface markers. Differentiation of PD cells into osteogenic, chondrogenic, and adipogenic lineages was also evaluated by von Kossa, Alizarin red, Alcian blue, and oil red O stains, respectively. Expression of mesenchymal stem cell markers were assessed using reverse transcriptase-polymerase chain reaction (RT-PCR) analysis.

Results: We successfully isolated and expanded MSCs from human periosteum. Flow cytometry revealed that PD cells were positive for mesenchymal adhesion cell markers (CD29, CD44, CD90, and CD105) and negative for hematopoietic markers (CD34 and CD45). In osteogenic differentiation, calcium accumulation (positive von Kossa and Alizarin red) and RUNX2, alkaline phosphatase, collagen type I, osteopontin genes were detected. In adipogenic differentiation, the cells displayed oil red O positive and expressed lipoprotein lipase and peroxisome proliferator-activated receptor-gamma (PPAR-γ) associated with adipogenesis. The cells grown in chondrogenic conditions were positively stained for Alcian blue and expressed SOX-9.

Conclusion: PD cells presented osteogenic, chondrogenic, and adipogenic differentiation abilities in vitro and could provide an alternative cellular source for tissue repair in clinical applications.