ERK5 silencing inhibits invasion of human osteosarcoma cell via modulating the Slug/MMP-9 pathway.

OBJECTIVE ERK5 is over expressed in a many of human cancers and this overexpression has been associated with metastasis and invasion. Furthermore, ERK5 silencing inhibits aggressive phenotypes of cancer cells. However, mechanisms by which ERK5 regulates tumour progression or metastasis have not been elucidated. In this study, using human osteosarcoma cell lines U2OS as a model, we explored the involvement of ERK5 silencing on invasiveness of U2OS cells. MATERIALS AND METHODS ERK5 siRNA targeting ERK5 was stably transfected into the human osteosarcoma cell lines U2OS. ERK5 knocked-down U2OS cells was then transfected with Slug cDNA or MMP-9 cDNA plasmid to re-express Slug or MMP-9. Cell proliferation was detected by MTT assay. Cell invasion and metastasis was detected by Matrigel invasion and wound healing assay. An orthotopic nude mouse model of U2OS was applied for in vivo lung metastasis experiments. ERK5, Slug, MMP-9 and E-cadherin were analyzed by real-time PCR, and Western blotting. RESULTS ERK5 silencing by siRNA in U2OS cells decreased Slug and MMP-9 expression. Compared with the vector-transfected cells, ERK5 knocked-down cells showed reduced migration and invasion in vitro, as well as decreased metastatic potential in experimental metastasis. Re-expression of Slug or MMP-9 in ERK5 knocked-down cells restored the invasive phenotypes. We also discovered that Re-expression of Slug in ERK5 knocked-down cells restored the MMP-9 expression, and re-expression of MMP-9 in ERK5 knocked-down cells did not affect Slug and ERK5 expression. CONCLUSIONS Our data suggest that ERK5 knockdown inhibits aggressive behaviour of human U2OS cells through modulating Slug signaling and MMP-9 expression.