Hepatitis B virus X-interacting protein (HBXIP), a cofactor of survivin, previously had been found to promote cell proliferation. In order to demonstrate the molecular mechanismsthe effect of HBXIP on transcriptional activity of NF-κB was investigated. The eukaryotic expression vector of HBXIP (pcDNA3-hbxip), or the RNA interference vector of HBXIP (pSilencer-hbxip) and NF-κB luciferase reporter (pNF-κB-Luc) were co-transfected into H7402 hepatoma cells. The pRL-TK vector was used as an internal control for normalization of luciferase activity, and the luciferase activity was determined by using the dual luciferase reporter assay system. The luciferase assay indicated that the over-expression of HBXIP could stimulated NF-κB transcriptional activity in H7402 hepatoma cells. RNA interference (RNAi) targeting HBXIP mRNA resulted in the opposite effects. Statistically no significant difference was observed between the control cells and pcDNA3 empty vector-transfected cells or pSilencer-control transfected cells. The phosphorylation level of IκBα, an inhibitor of NF-κB, was increased by Western blot analysis when over-expression of HBXIP in H7402 cells. The p65/NF-κB level in nucleus extraction from H7402 cells was examined by Western blot analysis after transfection. The data indicated that over-expression of HBXIP in H7402 cells was able to increase the levels of p65/NF-κB in the nucleus. However, down-regulation of HBXIP mRNA in the cells by RNAi resulted in the opposite results. In conclusion, HBXIP is involved in cell proliferation by regulating NF-κB signal pathway.