Effect of puerarin on the apoptosis of human cholangiocarcinoma RBE cells

Abstract

Objective To study the effects of puerarin on the proliferation and apoptosis of human cholangiocarcinoma (RBE) cells and the expression of related genes. Methods RBE cells cultured were treated with puerarin at different concentrations (0, 100, 250, 500, 1 000, 1 500 μg/ml). And then methyl thiazol tetrazolium (MTT) assay was used to measure cell viability. Flow cytometry (FCM) was used to detect the apoptosis of RBE cells treated with puerarin at (500, 1 000 μg/ml) concentrations. The expression levels of B cell lymphoma/leukemia-2 (bcl-2) and bcl-2 associated X protein (bax) genes were detected by real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR). Western blotting was used to detect the expression of apoptosis-related proteins. Results MTT assay revealed that puerarin (500, 1 000, 1 500 μg/ml) inhibited the proliferation of RBE cells in a time- and dose-dependent manner, with inhibition rate at 12, 24 and 48 h being (8.14±1.85)%, (12.57±3.10)% and (18.26±2.74)% (P=0.000, 0.005, 0.003), (22.38±2.77)%, (29.14±3.85)% and (37.43±2.11)% (P=0.005, 0.007, 0.002), and (35.47±3.75)%, (40.81±2.52)% and (52.29±3.98)% (P=0.007, 0.000, 0.002) respectively. After RBE cells were treated with puerarin at (500, 1 000 μg/ml) for 24 h, the apoptosis rate [(14.21±2.53)% and (22.87±2.74)%] was increased greatly as compared with the control group [(3.246±2.25)%] (P=0.013, 0.008). The expression of bax was up-regulated (2.37±0.17 and 3.78±0.41) times as compared with the control group (0.98±0.04) (P=0.005, 0.003), and the expression of bcl-2 was down-regulated (1.57±0.14 and 2.86±0.55) times as compared with the control group (0.99±0.03) (P=0.002 and 0.008). The expression levels of Cleaved-Caspase-8, Cleaved-Caspase-9 and bax protein were increased (P=0.015, 0.003 and 0.008) and those of bcl-2 protein were decreased (P=0.021). Conclusion Puerarin can inhibit the proliferation of RBE cells and promote the apoptosis. Key words: Puerarin; Cholangiocarcinoma; Proliferation; Apoptosis