Our objective was to delineate the role of nitric oxide (NO) in osteopontin (OPN)-associated metastatic properties in HepG2 cells. OPN is the major phosphoprotein secreted by malignant cells in patients with advanced metastatic cancer, is frequently overexpressed in human tumors, and has been implicated as a key mediator of tumor cell metastasis. OPN is significantly overexpressed in hepatocellular cancer (HCC) and correlates with capsular infiltration and behavior. In addition, significantly increased inducible nitric oxide synthase (iNOS) and NO expression are found in HCC. In archived human samples of normal, cirrhotic, and HCC livers, we demonstrate that iNOS and OPN protein are strongly coexpressed in hepatoma cells. In the setting of cirrhosis, hepatocytes express iNOS, but not OPN. Further in vitro studies performed with HepG2 hepatocellular cancer cells demonstrate that exogenous NO transcriptionally upregulates OPN expression. Enhanced expression of OPN in this setting is associated with increased in vitro cell adhesion and invasion. These data suggest that NO enhances HCC expression of OPN and, as a result, conveys a metastatic phenotype.