Previous attempts at identifying the constitutive source(s) of granulocyte colony-stimulating factor (G-CSF) in human bone marrow have been unsuccessful despite the fact that normal bone marrow supports abundant myelopoiesis in vivo. We hypothesized that the intimate physical association between bone and hematopoietic cells facilitates interactions between osteoblasts and hematopoietic stem cells. Here we provide the first direct evidence that human osteoblasts participate in hematopoiesis by constitutively producing G-CSF and present the protein in a membrane-associated fashion to human hematopoietic progenitors. These results suggest a direct and central role for osteoblasts in normal myelopoiesis.