Low-intensity pulsed ultrasound (LIPUS) is an established therapy for fracture healing where bone marrow stromal cells (BMSCs) migration is crucial to bone regeneration. This work focused on different performances of C-X-C-receptor 4 (CXCR4), integrin-1β and chemokine-chemokine receptor2 (CCR-2) in BMSCs migration by LIPUS stimulation. Single 20-min LIPUS treatment was applied to BMSCs during wound healing assay with or without the inhibitor AMD3100. The migration rate of BMSCs with LIPUS stimulation exhibited a higher closure rate than that of BMSCs without LIPUS stimulation, which was 1.89 μm/h and 1.38 μm/h, respectively. After LIPUS stimulation, significant elevation of the expression of CXCR4, integrin-1β and CCR-2 was observed. When AMD3100 was added, the migration rate of the BMSCs was obviously declined with or without LIPUS treatment. Furthermore, the expression of CXCR4 was significantly down-regulated by AMD3100, while integrin-1β and CCR-2 were less affected. It suggested that the enhancement of the migration of the BMSCs by LIPUS was inhibited by AMD3100. The results confirmed that LIPUS stimulation was able to activate and improve migration of BMSCs. Nevertheless, CXCR4 and both integrin-1β and CCR-2 had different roles in BMSCs migration after LIPUS treatment.