Growing evidences show that mesenchymal stem cells (MSCs) home to tumorgenesis and inhibit tumor cells, however, the molecular mechanisms underlying are unclear. The human mesenchymal stem cells (hMSCs) derived from human fetal bone marrow in 4 months were established without immortalization, designated BMMS-03 cells. In order to clarify the molecular mechanism underlying, the effect of conditioned media from hMSCs on human breast cancer MCF-7 cells was examined. The results showed that conditioned media from BMMS-03 cells were able to decrease the colony-forming units and proliferation of MCF-7 cells by colony-forming assay and MTT assay. Western blot analysis revealed that β-catenin and its target gene c-Myc, Bcl-2, PCNA and survivin were downregulated in MCF-7 cells treated with conditioned media from BMMS-03 cells. In addition, the treatment significantly reduced β-catenin nuclear assembly in MCF-7 cells by immunoflorescence staining. The finding demonstrated that the expression level of Dkk-1 in hMSCs was much higher than that in MCF-7 cells. Moreover, treatment with the antibody of rabbit against Dkk-1 abolished the inhibitory effects of conditioned media from BMMS-03 on the tumor cells. Conditioned media from BMMS-03 cells transiently transfected with pcDNA3.1(-)-Dkk-1 produced stronger downregulation of β-catenin and c-Myc expression in MCF-7 cells. The transfection with pcDNA3.1(-)-Dkk-1 in MCF-7 cells resulted in the downregulation of β-catenin and c-Myc as well．The data suggest that hMSCs are able to decrease the malignant phenotype of tumor cells in vitro. Taken together, hMSCs may suppress tumor growth via Wnt/β-catenin pathway, in which Dkk-1 released from the hMSCs is responsible for the depression.