To study the gene expression changes in mesenchymal stem cells from bone marrow stroma (MSCs) during in vitro expansion (from low density), passage 1 MSC were grown in culture for 15 days with medium change every 2-3 days. Samples for microarrays were taken at day 5 (early log-phase), 10 (late log-phase) and 15 (stationary phase). The data was queried for expression changes in Wnt signaling molecules and cell surface markers. Several components of the canonical Wnt signaling pathway were expressed, including Dkk-1; Wnt-5a; alpha-catenin; beta-catenin; frizzled 1, 4, 6, and 7; disheveled; glycogen synthetase kinase 3 beta; and glycogen synthetase kinase 3 alpha. In addition, the expression of over 10 cell surface transcripts decreased and an almost equal number increased during expansion. The two of the transcripts with the largest decreases coded for proteins previously shown to be linked to cell motility and tumor progression: PODXL, and alpha6-integrin (CD49f). As the cultures expanded, the largest increase was for mRNA for the cell adhesion protein VCAM-1. To study the gene expression changes in more detail, real-time RT-PCR, RT-PCR, ELISAs, FACS, and western blotting were performed for additional MSC donors. The results demonstrated dramatic changes in the transcriptome of MSCs during in vitro expansion.