Humans are exposed to perfluorooctane sulfonate (PFOS) from sources of both continuing discharges and environmental legacies, and the health effects of low dose exposure remain unknown. We assessed the effects of PFOS at 0.2-200 nmol/L on osteogenic and adipogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSCs). PFOS affected hBMSCs differentiation in a nonmonotonic dose-response relationship, where the effects peaked at 100 nmol/L. PFOS repressed calcium deposition on day 14 of osteogenic differentiation. The molecular osteogenic biomarkers, osteopontin, osteonectin and osteocalcin, and the transcription factor β-catenin, were also decreased on differentiation day 7. In hBMSCs co-treated by PFOS and WNT signaling agonist, LiCl, the osteogenic marker levels were higher than those treated by PFOS alone. Moreover, the osteogenic effect of PFOS was repressed by DKK1, the WNT signaling antagonist. PFOS additionally supported the receptor activator of nuclear factor κB ligand/osteoprotegerin ratio. In contrast, PFOS enhanced adipogenesis in regard to lipid droplet formation and marker gene expression of PPARγ, CCAAT/enhancer-binding protein-α (C/EBPα), lipoprotein lipase and leptin. Microarray analysis identified 597 differentially expressed genes at p < .05 and log2 (fold change) > 0.3. These results demonstrated that PFOS repressed osteogenesis and enhanced adipogenesis, which could well explain the homeostatic imbalance seen in human bone associated with exposure to PFOS.