Activated hepatic stellate cells (HSCs) are the major subtype of stromal cells in the liver tumor microenvironment which can promote the growth and migration of hepatocellular carcinoma (HCC) cells. However, the underlying mechanisms by which activated HSCs exert their oncogenic effects are not fully understood to date. In the present study, we investigated the number of activated HSCs and its clinicopathological significance in HCC and uncovered its correlation with focal adhesion kinase (FAK)-MMP9 signaling. A higher number of activated HSCs was associated with tumor invasion of the portal vein, advanced TNM stage and poorer tumor differentiation. The number of activated HSCs was positively correlated with the expression levels of p-FAK and MMP9 in HCC. Furthermore, we studied the effects of activated HSCs on the migration and invasion of HCC cells in vitro. Conditioned medium (CM) from activated HSCs or co-culture with activated HSCs significantly induced the migration and invasion of HCC cells. In addition, activation of FAK-MMP9 signaling in HCC was demonstrated in the presence of activated HSC-CM and of co-culture. Inhibition of FAK-MMP9 signaling in HCC cells with FAK short hairpin RNA (shRNA) abrogated the effects of activated HSCs on HCC cells. Taken together, our data suggest that activated HSCs in the tumor microenvironment promote HCC cell migration and invasion via activation of FAK-MMP9 signaling.