The effect of docosahexaenoic acid combined with cyclooxygenase-2 selective inhibitor NS-398 on the apoptosis of bile duct cancer cells QBC939

Abstract

Objective To investigate the effect of docosahexaenoic acid (DHA) combined with cyclooxygenase-2 (COX-2) selective inhibitor NS-398 on the apoptosis of cholangiocarcinoma QBC939 cells and the mechanism. Methods In vitro, cholangiocarcinoma QBC939 cells were treated with 0, 15, 30, 45, 60 and 75 μg/ml DHA with 0, 25, 50, 100, 150 and 200 μmol/L NS-398, respectively. The absorbances of the QBC939 cells were measured by CCK8 and its growth inhibition ratios were analyzed. Flow cytometry was applied to detect cell apoptosis. The level of β-catenin and COX-2 mRNA and protein were measured by real-time PCR, immunocytochemistry and enzyme-linked immunoadsordent assay, respectively. Results DHA combined with NS-398 could significantly suppress the growth of QBC939 cells (P<0.05). When the concentration of DHA went up to 45 μg/ml and NS-398 was 100 μmol/L, the relative growth inhibition rate of QBC939 cells was 90.0%. If the concentrations were increased, the result showed no significant differences. Furthermore, flow cytometry analysis indicated that DHA combined with NS-398 could induce QBC939 cells apoptosis at the early stage, and the apoptosis rate was significantly different between the experimental and control groups (P<0.01). Real-time PCR showed low β-catenin and COX-2 expression in QBC939 cells disposed by DHA combined with NS-398, and their expression were significantly different between the experimental and control groups (P<0.01). Immunocytochemistry and ELISA demonstrated that DHA combined with NS-398 could decrease β-catenin and COX-2 protein expression in QBC939 cells. Conclusion DHA combined with NS-398 induced apoptosis and inhibited proliferation of cholangiocarcinoma cells QBC939 in vitro through targeting β-catenin and COX-2. Key words: Docosahexaenoic acid (DHA); NS-398; Cholangiocarcinoma; Apoptosis; β-catenin; Cyclooxygenase-2 (COX-2)