Phenotype and gene expression of human mesenchymal stem cells in alginate scaffolds

Human mesenchymal stem cells (MSC) are popular candidates for tissue engineering. MSC are defined by their properties in two-dimensional (2D) culture systems. Cells in 2D are known to differ from their in vivo counterparts in cell shape, proliferation, and gene expression. Little is so far known about the phenotype and gene expression of cells in three-dimensional (3D) culture systems. To begin to unravel the impact of 3D versus 2D culture conditions on MSC, we have established MSC from adipose tissue and bone marrow in 3D cultures in alginate beads covalently modified with the tripeptide arginine-glycine-aspartic acid (RGD), the integrin-binding motif found in several molecules within the extracellular matrix. The MSC changed from their fibroblastoid shape (2D) to a small, compact shape when embedded in RGD alginate (3D). High viability was maintained throughout the experiment. The MSC retained expression of integrins known to bind RGD, and practically ceased to proliferate. Microarray analysis revealed that the gene expression in cells in RGD alginate was different both from the cells cultured in 2D and from prospectively isolated, uncultured MSC, but more similar to 2D cells. As alginate may be entirely dissolved, leaving the cells as single cell suspensions for various analyses, this represents a useful model for the study of cells in 3D cultures.