There has been considerable interest in developing new therapies with adult multipotent progenitor stromal cells or mesenchymal stem cells (MSCs) in organ replacement and repair. To be effectively seeded into scaffolds for therapy, large numbers of cells are needed, but concerns remain regarding their chromatin stability in long-term culture. We therefore expanded four donors of human MSCs (hMSCs) from bone marrow aspirates with a protocol that maintains the cells at low density. MSCs initially proliferated at average doubling times of 24 h and then gradually reached senescence after 8-15 passages (33-55 population doublings) without evidence of immortalization. Comparative genomic hybridization assays of two preparations revealed no abnormalities through 33 population doublings. One preparation had a small amplification of unknown significance in chromosome 7 (7q21:11) after 55 population doublings. Microarray assays demonstrated progressive changes in the transcriptome of the cells. However, the transcriptomes clustered more closely over time within a single passage, rather than with passage number, indicating a partial reversibility of the patterns of gene expression. One of the largest changes was a decrease in mRNA for Sox11, a transcription factor previously identified in neural progenitor cells. Knockdown of Sox11 with siRNA decreased the proliferation and osteogenic differentiation potential of hMSCs. The results suggested that assays for Sox11 may provide a biomarker for early progenitor hMSCs.