The maintenance of bone marrow stromal stem cells (BMSSCs) is tightly controlled by the local microenvironment and by autocrine regulatory factors secreted by BMSSCs. To identify such factors, a cDNA subtraction library was generated from purified BMSSCs, based on their high expression of the STRO-1 antigen. Stromal-derived factor-1 (SDF-1) was one differentially expressed gene highly expressed by purified BMSSCs prior to culture. In vitro, immature preosteogenic cells expressed greater levels of SDF-1 when compared with mature cell types representative of osteoblasts and osteocytes/bone lining cells. Furthermore, SDF-1 expression was rapidly down-regulated when BMSSCs were cultured under osteoinductive conditions. BMSSCs were also shown to express functional cell surface SDF-1 receptors (CXCR4). Transduced BMSSC lines, secreting high SDF-1 levels, displayed an enhanced ability to form ectopic bone in vivo, in comparison with control BMSSC lines. Moreover, high SDF-1-expressing BMSSCs displayed an increased capacity for cellular growth and protection against interleukin-4-induced apoptosis. Similarly, fibroblast colony-forming units (CFU-Fs) also displayed increased growth and resistance to alpha-interferon-2a-induced apoptosis, in synergy with platelet-derived growth factor BB (PDGF-BB) and SDF-1 in vitro. These studies indicate that the chemokine, SDF-1, may play a role in the maintenance, survival, and osteogenic capacity of immature BMSSC populations.