AIM:To investigate the effect of activation of canonical Wnt signaling pathway on the proliferation and differentiation of hepatic oval cells in vitro.METHODS:WB-F344 cells were treated with recombinant Wnt3a (20,40,80,160,200 ng/mL) in serum-free medium for 24 h.Cell proliferation was measured by Brdu incorporation analysis; untreated WB-F344 cells were taken as controls.After treatment with Wnt3a (160 ng/mL) for 24 h,subcellular localization and protein expression of β-catenin in WB-F344 cells treated and untreated with Wnt3a were examined by immunofluorescence staining and Western blot analysis.CyclinD1 mRNA expression was determined by semi-quantitative reverse-transcript polymerase chain reaction (RT-PCR).The mRNA levels of some phenotypic markers (AFP,CK-19,ALB) and two hepatic nuclear factors (HNF-4,HNF-d) were measured by RT-PCR.Expressions of CK-19 and AFP protein were detected by Western blot analysis.RESULTS:Wnt3a promoted proliferation of WB-F344 cells.Stimulation of WB-F344 cells with recombinant Wnt3a resulted in accumulation of the transcriptional activator β-catenin,together with its translocation into the nuclei,and up-regulated typical Wnt target gene CyclinD1.After 3 d of Wnt3a treatment in the absence of serum,WB-F344 cells retained their bipotential to express several specific phenotypic markers of hepatocytes and cholangiocytes,such as AFP and CK-19,following activation of the canonical Wnt signaling pathway.CONCLUSION:The canonical Wnt signaling pathway promotes proliferation and self-renewal of rat hepatic oval cells.