OL-023 Activation of canonical Wnt signaling pathway promotes proliferation and self-renewal of rat hepatic oval cell line WB-F344 in vitro

Abstract

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 ce l l s were t reated wi th 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, subcel lu lar 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-6 ) 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