Abstract
In a previous study, it was demonstrated that Rhein lysinate (RHL) inhibited HeLa cell proliferation via a specific mechanism. The aim of the present study was to clarify the mechanism of RHL by investigating its effect on mitochondrial damage and cell apoptosis. The results indicated that RHL inhibited cell growth and proliferation in HeLa cells. HeLa cells treated with RHL developed extensive vacuolization in a dose- and time-dependent manner. Ultrastructure analysis using transmission electron microscopy revealed that the vacuoles observed were damaged mitochondria and endoplasmic reticulum. The effects of RHL on mitochondria were further confirmed by a decrease in mitochondrial membrane potential and increased generation of reactive oxygen species. The mitochondrial proteome was analyzed, and the results demonstrated that the expression of the cytoskeletal protein keratin and dermal papilla derived protein 12 (associated with the oxidation-reduction process), which are associated with mitochondrial structure and function, were decreased compared with the untreated control group. Hoechst staining, flow cytometry and western blotting also revealed that apoptosis was induced at 24 h following RHL treatment. These results confirm that RHL toxicity in HeLa cells is a dynamic process. Vacuolar degeneration appeared in HeLa cells treated with 160 µmol/l RHL during the first 6 h and with the extension of RHL treatment, cell apoptosis was presented at ~24 h in HeLa cells.