ABSTRACT
Objective: To investigate the effect of combination treatments of cisplatin and KK4 and ICG15042 peanut testa extracts against cholangiocarcinoma cells in vitro.Methods: The growth inhibition, cell cycle arrest and apoptosis of cholangiocarcinoma cells were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and flow cytometry analysis, respectively. The levels of proteins involved in apoptosis were assessed using Western blotting assays. The caspase activity was assessed using a colorimetric caspase activity assay. Results: Cisplatin and peanut (KK4 and ICG15042) testa extracts inhibited the growth of cholangiocarcinoma cell lines (KKU-M214 and KKU-100 cells) in a dose- and time-dependent manner. The combination treatments reduced cell viability and induced apoptosis of cholangiocarcinoma cells more efficiently than single-drug treatments. Cancer cell death synergistically mediated by cisplatin and peanut testa extracts was observed in KKU-M214 cells (combination index < 1.0) but not in KKU-100 cells (combination index > 1.0). The combination treatments also increased the sub-G1 population and caused KKU-M214 cell cycle arrest at S and G2/M phases, which were the combined effects of cisplatin (S phase arrest) and peanut testa extracts (G2/M phase arrest). In addition, pERK1/2, Ac-H3, Bcl-2 and proteins related to apoptosis, including Bax and caspases 3, 8, 9, exhibited enhanced expression in KKU-M214 cells. The combination treatments caused down-regulation of p53, whereas the expression of p21 was fairly constant when compared with cisplatin single drug treatment. Conclusions: Peanut testa extracts in combination with cisplatin synergistically reduce cell viability and induce apoptosis through stimulation of caspases 3, 8 and 9 in KKU-M214 cells.