N-butanol fraction of Entada phaseoloides ethanol extract inhibits hepatocellular carcinoma HepG2 cell proliferation by inducing apoptosis.

Purpose: To screen for substances with inhibitory effects on the proliferation of hepatocellular carcinoma (HCC) HepG2 cell line from extracts of traditional Chinese medicinal plants including Heliciopsis lobata (Merr.) Sleum, Bidens pilosa, Entada phaseoloides, Plantago major, and Smilax, and unveil their mechanism of action. Methods: 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was used to assess the inhibition of HepG2 cell proliferation by plant extracts. Cell apoptosis was evaluated by Hoechst 33342 staining and mitochondrial transmembrane potential (Delta Psi m) dissipation was measured using JC-1 probe by fluorescence microscopy. Results: Heliciopsis lobata, Bidens, Plantago, and Smilax extracts showed reduced inhibitory effects on HepG2 cell proliferation compared with Entada phaseoloides (all p<0.05). The n-butanol fraction of Entada phaseoloides ethanol extract exhibited the highest inhibition rate. Treatment of HepG2 cells with 500, 250, and 100 mu g/ml n-butanol extract resulted in 89.92 +/- 0.58%, IC50 81.66 +/- 0.42%, 68.85 +/- 0.57% decrease in cell viability, respectively, indicating an IC50 of 9.27 mu g/ml. In the presence of 100 mu g/ml entada phaseoloides n-butanol extract for 24h, apoptotic nuclei and hyperchromatic, dense fluorescent massive granules were observed in the cytoplasm, effects that increased with extract concentrations in HepG2 cells. Finally, we showed that Entada phaseoloides n-butanol extract induced depolarization of mitochondrial membrane potential. Conclusions: Entada phaseoloides n-butanol extract inhibits HepG2 cell proliferation by inducing cell apoptosis likely through mitochondrial apoptotic pathway. This extract is therefore a potential natural source towards the discovery for a new drug-candidate for the treatment of HCC.