Genistein reduces the activation of AKT and EGFR, and the production of IL6 in cholangiocarcinoma cells involving estrogen and estrogen receptors

Cholangiocarcinoma (CCA) is a malignant tumor of the biliary epithelium associated with Opisthorchis viverrini, primary sclerosing cholangitis and hepatitis viral infection. In the global population, men have higher incidence rates for CCA than women; thus, a gender disparity in the progression of chronic inflammation of the biliary duct leading to malignancy may involve the effects of estrogen (E2). Genistein (GE), a prominent phytoestrogen found in soy products, is an estrogen receptor (ER) agonist and a tyrosine kinase inhibitor. The present study investigated the effects of GE on the growth of CCA cells by cell viability assay. The effects on signaling proteins were detected by western blot analysis and ELISA. Gene expression was examined by RT-qPCR. Two human intrahepatic CCA cell lines, HuCCA-1 and RMCCA-1, were utilized. GE (50-200 mu M) reduced the viability of the two cell lines, and also inhibited the activation of epidermal growth factor receptor (EGFR) and AKT, as evidenced by decreasing protein levels of phosphorylated (p)-EGFR (Tyr1173) and p-AKT (Ser473), respectively. GE altered the mitogen-activated protein kinase signaling cascade by mediating decreased protein levels of p-p38 and increased protein levels of p-ERK1/2. GE significantly decreased the levels of interleukin 6 (IL6) and induced the expression of inducible nitric oxide synthase (iNOS). GE also down-regulated the expression of p-ER (Ser118) protein and ER mRNA levels. Finally, GE induced the downregulation of the protein levels of ER. Of note, E2 deprivation potentiated the GE-induced reduction of p-EGFR (Tyr1173) and total AKT proteins and production of IL6, and mediated the downregulation of GE-induced iNOS protein. In conclusion, GE inhibited the growth of human CCA cell lines by reducing the activation of EGFR and AKT, and by attenuating the production of IL6. E2 and ER were also involved in the growth-inhibitory effect of GE in CCA cells.