In human cancers, multiple members of microRNAs (miRNA) are often repressed, such as the let-7 family miRNA. The let-7 family regulates self-renewal and tumorigenicity of cancer cells, and plays critical roles in embryonic development. The RNA binding proteins Lin-28 and Lin-28B are recently demonstrated to negatively regulate let-7 biogenesis by inducing terminal uridylation and degradation of the let-7 precursors. In this study, we showed that in hepatocellular carcinoma (HCC), only Lin-28B but not Lin-28 was highly expressed. Lin-28B expression in HCC was more frequently found in high grade tumors with high α-fetoprotein levels. Among the HCC cell lines, Hep3B and HCC36 expressed higher levels of Lin-28B than the other cell lines, and were chosen for gene knockdown of Lin-28B. HA22T cell line which had very low level or undetectable Lin-28B, was chosen for ectopic expression of Lin-28B. Knockdown of Lin-28B by RNA interference in the Hep3B and HCC36 led to suppressed proliferation in vitro and reduced in vivo tumor growth in NOD/SCID mice. In contrast, overexpression of Lin-28B in HA22T enhanced proliferation and tumorigenecity. Moreover, overexpression of Lin-28B induced epithelial-mesenchymal transition in HA22T cells, accompanied by increased invasion and migration capacity. To further delineate the role of Lin-28B in the expression of miRNA family, we did a large-scale real-time PCR array analysis. The results revealed that only let-7/mir-98 family members were regulated by Lin-28B. Lin-28B overexpression enhanced the expression of the known let-7 targets c-myc and HMGA2. In this study, we also found that Lin-28B enhanced the expression of type 1 insulin-like growth factor receptor (IGF1R) in a let-7-dependent manner. These findings highlight the important role for Lin-28B in tumor formation and invasion in HCC, through coordinated repression of the let-7/mir-98 family and induction of multiple oncogenic pathways.