The aim of the present study was to investigate the effect of co-culture with amniotic epithelial cells (AECs) on the biological characteristics of amniotic mesenchymal stem cells (AMSCs), to compare the expression of C-X-C motif chemokine receptor 4 (CXCR4) in co-cultured AMSCs and to investigate the roles of the stromal cell-derived factor-1 (SDF-1);CXCR4 axis in the homing and migration of AMSCs. AMSCs were isolated from human amniotic membranes, purified and then differentiated into osteoblasts and adipocytes in vitro, which was verified by von Kossa Staining and Oil Red 0 staining. Cell viability was measured by Cell Counting kit-8 and trypan blue assays at 24, 48 and 72 h, the expression of CXCR4 was analyzed by immunofluorescence-based flow cytometry and reverse transcription-quantitative polymerase chain reaction, and the migration ability of AMSCs in vitro was observed by a migration assay. The results demonstrated that cell viability (at 48 and 72 h) and survival (at 24, 48 and 72 h) in the co-culture and serum groups were higher compared with the serum-free group. Furthermore, CXCR4 mRNA and protein expression, and migration along the SDF-1 gradient, in the co-culture and serum-free groups were higher compared with the serum group. Overall, the results indicated that AMSCs co-cultured with AECs exhibited enhanced proliferation activity and survival rate. In conclusion, the present study demonstrated that co-culture of AMSCs with AECs upregulated CXCR4 on the surface of AMSCs and enhanced the migration ability of AMSCs in vitro. This result may improve the directional migration and homing ability of AMSCs, as well as provide a theoretical basis for the application of AMSCs in clinical practice as a novel strategy to increase the success of hematopoietic stem cell transplantation.