This study was to investigate the effect of vascular endothelial cell conditioned medium on the proliferation and migration of gastric cancer cells, and further to explore the potential mechanism of gastric cancer metastasis. Methods Control group and co-culture group were set up. Gastric cancer cells HGC27 were stimulated with human umbilical vein endothelial cells (HUVEC ) conditioned mediumr or normal medium, respectively, and the proliferation and migration of HGC27 were detected by MTT assay and scratch test, respectively. Then control group, 6h group, 12h group and 24h group were built the following tests. The expression of epithelial-mesenchymal transition (EMT) and tight junction were detected by Western blot after HGC27 treated with HUVEC conditioned medium for 0h, 6h, 12h and 24h durations.Furthermore, the distribution changes of cytoskeleton and tight junctions were observed with laser confocal microscopy.Results MTT and scratch test showed that HUVEC medium promoted the proliferation and migration of HGC27, respectively.Compared with the Control group, the morphology of HGC27 turned to mesenchymal after co-culture, and the number of filamentous pseudopod-like protrusions (FLPs) increased significantly.WB results showed that the expression level of E-cadherin decreased gradually, while N-cadherin and MMP-9 increased gradually in a time-depend manner.The expression of ZO-1 and Occludin also displayed a decreasing trend, and the distribution were both decreased obviously.Conclusion Vascular endothelial cells promote gastric cancer cell migration by up-regulating MMP-9 and destroying tight junctions in the indirect co-culture condition