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 （2...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 p-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, HIVF-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 resulte^l 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.展开更多
文摘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 p-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, HIVF-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 resulte^l 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.
