Objective:To study the effect of transfecting the neuronal guidance factor Netrin-1 and enhanced green fluorescent protein gene(EGFP) to rat bone mesenchymal stem cells(BMSCs) by adenovirus on the cytokine secretion, differentiation and tube formation. Methods:BMSCs were separated and purified in vitro. Surface receptors of adenovirus infected BMSCs and cytokines were detected by real-time PCR or RT-PCR.Tube formation of HUVEC was observed by coculture with BMSCs. BMSCs migration was observed by Transwell experiment. Results:Adenovirus infected BMSCs expressed some kinds of Netrin-1 receptors and secreted vascular endothelial growth factor(VEGF),basic fibroblast growth factor(bFGF),brain derived neurophic factor(BDNF) and nerve growth factor(NGF). BMSCs cultured with Netrin-1 and VEGF markedly increased the tube formation of HUVEC and BMSCs migration,compared with Netrin-1 or VEGF alone(P < 0.05). Conclusion:BMSCs successfully infected with Ad5-Netrin-1-CMV-EGFP can express EGFP and Netrin-1.Netrin-1 has the ability to accelerate tube formation and BMSCs migration and VEGF can enhance the effect when used together.