To establish HP14.5 cell line stably expressing hepatitis B virus X (HBx) and detect the effect of HBx on cell proliferation, cell cycle and the wnt/β-catenin signal pathway of hepatic progenitor cells.The plasmid of pSEB-Flag-HBx and the packaging plasmid pAmpho were co-transfected into HEK 293T cells to construct the recombinant retrovirus carrying HBx gene. The recombinant retrovirus wan then transfected into mouse hepatic progenitor cells HP14.5. The blasticidin-resistant clones of HBx cells (HP14.5/HBx) were selected out and cultured as the experimental group, paralleled with the vector control HP14.5/Rv (HP14.5/pSEEB-Flag) and HP14.5 cell as negative control. The cell proliferation was tested by MTS assay, the cell cycle was measured by flow cytometry, the expression levels of cyclin D1 and c-myc mRNA were tested by real time PCR, and the expression leveIs of HBx, cyclin D1, c-myc, GSK3β, p-GSK3β (ser-9), β-catenin proteins were examined by Western blotting.The expression of HBx at both mRNA and protein levels was positive in HP14.5/HBx cell line as confirmed by RT-PCR and Western blotting. Compared with the control groups, the proliferation of HP14.5/HBx cells increased significantly (P0.05).The hepatic stem cell line stably expressing HBx has been constructed successfully. HBx can promote the proliferation and malignant transformation of hepatic stem cells via the activation of Wnt/β-catenin signal pathway.