Transcriptome changes in hMSCs during expansion

Human mesenchymal stem cells or multipotent stromal cells (MSCs) are of interest for clinical therapy, in part because of their capacity for proliferation and differentiation. However, results from clinical trials and in vitro models have been variable, possibly due to MSC heterogeneity and a lack of standardization between MSC in vitro expansion protocols. Here we defined changes in MSCs during expansion in vitro. In low density cultures, MSCs expand through distinct lag, exponential growth and stationary phases. We assayed cultures of passage 2 human MSCs from three donors at low density (50 cells/cm2) at about 5% confluence on Day 2 and after the cultures had expanded to about 70% confluence on Day 7. On Day 2 genes involved in cell division were up-regulated. On Day 7 genes for cell development were up-regulated. The variations between three donors were less than the variation within the expansion of MSCs from a single donor. The microarray data for selected genes were confirmed by real-time PCR, ELISA and FACScan. About 50% of cells at Day 2 were in S-phase compared to 10% at Day 7. The results demonstrated major differences in early and late stage cultures of MSCs that should be considered in using the cells in experiments and clinical applications. Keywords: Time course We assayed cultures of passage 2 human MSCs from three donors at low density (50 cells/cm2) at about 5% confluence on Day 2 and after the cultures had expanded to about 70% confluence on Day 7.