Zanthoxylum avicennae extract enhances GSK-3β to attenuate β-catenin via phosphatase 2A to block metastatic effects of HA22T cells and hepatocellular carcinoma xenografted nude mice.

Hepatocellular carcinoma (HCC) metastasis is often associated with the activation of Wnt/β-catenin signaling pathway. Zanthoxylum avicennae (Ying Bu Bo, YBB), a traditional herb with hepatoprotective effect, has been proven to inhibit human HCC in in vivo models however, the in vitro and in vivo effect of YBB on tumor metastasis is not clear yet. To determine whether YBB could inhibit HA22T human HCC cell by acting on β-catenin metastatic signaling in vitro and in vivo, HA22T cells were treated with different concentrations of YBB extracts (YBBE) and analyzed by Immunofluorescence staining assay, western blot analysis, siRNA mediated gene knock-down assays and co-immunoprecipitation assay. Additionally, the HA22T-implanted xenograft nude mice were used to confirm the assessed cellular effects. Mice treated with YBBEs showed a strong increasing trend in PP2Acα, GSK-3β, APC, and β-TrCP/HOS levels, however the expression of β-catenin, p-GSK-3β, TBX 3, and IL8 proteins showed a decreasing trend. YBBE significantly downregulated the nuclear and cytosolic β-catenin levels by facilitating the proteosomal degradation of β-catenin. Moreover, as observed by co-immunoprecipitation assay, YBBE directly promoted the protein interactions between GSK-3β, β-TrCP, APC, PP2A, and β-catenin. In conclusion, both in vitro and in vivo models clearly demonstrated that YBBE inhibits β-catenin involved metastatic signaling in highly metastatic HA22T cells through PP2A activation.