© 2015 American Society for Blood and Marrow Transplantation.Granulocyte colony stimulating factor (G-CSF) is sometimes administered to donors before bone marrow (BM) harvest. G-CSF-primed (G-BM) and unprimed BM (U-BM)-derived mesenchymal stem cells (MSC) were obtained from 16 healthy donors and were expanded in vitro. Their proliferative characteristics, morphology, and differentiation capacity were examined. Supernatants of the second passage of MSCs were evaluated for transforming growth factor β1, hepatocyte growth factor, and prostaglandin E2 (PGE2) levels and compared with controls. The analyses of cytokines in the G-BM- and U-BM-derived MSCs supernatants revealed that PGE2 levels were significantly lower in the G-CSF-primed samples. These cytokines were also measured in BM plasma. The level of hepatocyte growth factor in G-BM plasma was significantly increased. The current study is the first to show the effects of G-CSF on the BM microenvironment of healthy human donors. The preliminary data suggest that G-BM- and U-BM-derived MSCs have similar morphologic/phenotypic properties and differentiation capacity but differ in their secretory capacity. Significant changes in cytokine levels of BM plasma in G-CSF-primed donors were also demonstrated. These findings suggest that BM MSCs and changes in the BM microenvironment may contribute to the effects of G-CSF on inflammation and immunomodulation.