The specific effects of interferon alpha (IFNβ£), on the differentiation pathways of human osteogenic cells are not known. The aim of this study was to investigate possible effects of IFNβ£ on osteogenic development by investigating cell differentiation, colony formation (colony forming unit-fibroblastic, CFU-F), cell proliferation, and gene expression, in particular bone morphogenetic protein (BMP) expression, of human bone marrow osteoprogenitor cells. Human bone marrow fibroblasts were cultured with or without the addition of IFNβ£ (5-1,000 IU/ml) in the presence and absence of dexamethasone (10 nM) and ascorbate (100 Β΅M), which are agents known to affect osteogenic differentiation. IFNβ£ produced a significant dose-dependent inhibition of cell proliferation and alkaline phosphatase specific activity at concentrations as low as 50 IU/ml. IFNβ£ (50-1,000 IU/ml) inhibited the stimulation of alkaline phosphatase specific activity induced by ascorbate and dexamethasone. Examination of CFU-F showed dose-and time-dependent inhibitions of colony formation and reductions in both colony size and alkaline phosphatase-positive CFU-F colonies particularly at earlier times. Reactivity with an antibody specific for osteoprogenitors (HOP-26), was reduced in IFNβ£-treated cultures. Northern blot analysis showed a significant dose-dependent up-regulation of BMP-2 mRNA, estrogen receptor alpha mRNA and osteocalcin mRNA expression in ascorbate/dexamethasone cultures. In contrast, IFNβ£ significantly inhibited BMP-2 mRNA expression in the absence of ascorbate and dexamethasone. In conclusion, IFNβ£ inhibits human osteoprogenitor cell proliferation, CFU-F formation, HOP-26 expression, and alkaline phosphatase specific activity and modulates BMP-2 gene expression. These results suggest a role for IFNβ£ in local bone turnover through the specific and direct modulation of osteoprogenitor proliferation and differentiation.