A Concise Review on the Use of Mesenchymal Stem Cells in Cell Sheet-Based Tissue Engineering with Special Emphasis on Bone Tissue Regeneration

Figure 2

Analyses of the MSC sheets on the basis of Alizarin Red S staining and reverse transcription polymerase chain reaction (RT-PCR). Mineralization of MSC sheets was analyzed on the basis of Alizarin Red S staining in all groups. Osteogenic gene expression profiles in all groups of MSC sheets were analyzed on the basis of RT-PCR. MSC sheets maintained in osteogenesis-stimulating medium supplemented with the triple cocktail for 21 days stained positive with Alizarin Red S (b) and they exhibited increased levels of collagen type I (Col1a2), osteocalcin (OC), osteopontin (OP), and alkaline phosphatase (ALP) mRNA (d) in comparison to MSC sheets maintained in basal medium supplemented with only ascorbic acid (a, d). MSC sheets maintained in basal medium supplemented with only ascorbic acid showed slight osteogenic differentiation (a, d) when compared to the MSC sheets maintained in basal medium without any supplementation (c, d); however, this differentiation was not as strong as seen in the triple cocktail supplemented group (b, d). Sequences and sources of the primers used for the gene expression analyses were given in Table 1. (d) Lane 1: MSC sheets maintained in basal medium supplemented with only ascorbic acid; lane 2: MSC sheets maintained in osteogenesis-stimulating medium; lane 3: MSC sheets maintained in basal medium without any supplementation; GAPDH: glyceraldehyde 3-phosphate dehydrogenase (housekeeping gene used as loading control in this experiment). Scale bar = 200 μm for (a, b, c).

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