In vitro and in silico approaches to unveil the mechanisms underlying the cytotoxic effect of juncunol on human hepatocarcinoma cells

Abstract

Background: Juncunol is a phenanthrene isolated from the halophyte species Juncus acutus, with selective cytotoxic activity towards human hepatocarcinoma (HepG2) cells. However, its mechanism of action is unknown.

Methods: The in vitro cytotoxic mechanism of juncunol was evaluated on HepG2 cells through several methods to elucidate its potential to induce apoptotic features, decrease mitochondrial membrane potential, promote internal ROS production and influence cell cycle. We also report its haemolytic activity on human erythrocytes and in silico DNA-binding studies.

Results: Juncunol induced an increase in the number of apoptotic cells in a concentration-dependent manner, accompanied by a decrease in the mitochondrial membrane potential. No significant differences were observed in production of reactive oxygen species (ROS). Moreover, juncunol application at the IC₅₀ value significantly induced cell cycle arrest in the G0/G1 phase comparatively to the control group. No significant haemolysis was detected. In silico studies indicate that juncunol seems to bind between GC base pairs.

Conclusion: Juncunol reduced HepG2 cells proliferation through the induction of apoptotic cellular death, in a concentration-dependent manner. Apoptosis induction seems to be related with a decrease of the mitochondrial membrane potential but not with ROS production. Juncunol had no haemolytic activity and may act as a DNA intercalator. Our data suggests juncunol as a suitable candidate for more detailed studies, including in vivo experiments, in order to completely characterize its mode of action.

Keywords: Apoptosis; Cell cycle; DNA binding; Flow cytometry; Haemolysis; Mitochondrial membrane potential; Reactive oxygen species.