We investigated the role of galectin-3 in metastasis of human breast carcinoma BT549 cells using the experimental liver metastasis model. Underlying mechanisms were then elucidated using the liver/tumor coculture and cell culture systems. After intrasplenic injection, galectin-3 cDNA transfected BT549 cells (BT549gal-3 wt) formed metastatic colonies in the liver, while galectin-3 null BT549 cells (BT549par) did not, demonstrating that galectin-3 enhances metastatic potential. More than 90% of BT549gal-3 wt cells survived after 24 hours-co-culture with the liver fragments isolated following ischemia treatment. In contrast, more than half of BT549par cells showed metabolic death following co-culture with the liver fragments. When the liver from inducible nitric oxide synthase (iNOS) knockout mice was used, no cytotoxicity to BT549par cells was observed. Thus, iNOS exerts cytotoxicity on BT549par cells and galectin-3 can protect against iNO-Sinduced cytotoxicity. BT549gal-3 wt also exhibited enhanced survival against peroxynitrite (up to 400 μmol/L) in vitro. A single mutation in the NWGR motif of galectin-3 obliterated both metastatic capability and cell survival, indicating that the antiapoptotic function of galectin-3 is involved in enhanced metastasis. In conclusion, galectin-3 enhances the metastatic potential of BT549 cells through resistance to the products of iNOS, possibly through its bcl-2-like antiapoptotic function.