Trophoblast invasion during implantation mimics the process of tumor metastasis in some aspects, however with a significant difference in that the former is strictly controlled. Several members of matrix metalloproteinases (MMPs) family have been shown to play crucial roles in regulating trophoblast cell invasion. MMP-26 is a newly identified MMP family member, and its functions in human trophoblast cells have yet to be elucidated. A well-accepted human trophoblast cell model———choriocarcinoma cell line (JEG-3) was used as in vitro model to investigate the involvement of MMP-26 in trophoblast cell invasion. Expressing pCR3.1 plasmid containing MMP-26 cDNA was transfected into JEG-3 cells, and a stable cell line with over-expressed MMP-26 was named as JEG-3/MMP-26. Transwell insert invasion assay showed that cell invasiveness in JEG-3/MMP-26 cells was significantly promoted. Data of RT-PCR and gelatin zymography demonstrated that the mRNA expression and zymogen secretion of MMP-9 were increased in these cells. Double immunofluorescent staining revealed co-localization of MMP-26 and MMP-9 proteins in JEG-3/MMP-26 cells. The data indicated that MMP-26 was involved in invasion-promoting regulation of human trophoblast cells, and it may function, at least in part, through coordination with other MMPs such as MMP-9.