Apoptosis of HepG2 Cell Induced by Ouabain and Cinobufogenin through ERK Signaling Pathway / 中国药房

ABSTRACT

OBJECTIVE:To investigate the effect of ouabain and cinobufogenin on the apoptosis of HepG2 cell in order to study its apoptosis mechanism through ERK signaling pathway. METHODS:The activity of Na+/K+-ATPase was detected with HepG2 cell as target. The morphological changes of HepG2 cell and DNA injury were determined by Hoechst 33342 fluorescent staining and single cell gel electrophoresis (SCGE) respectively. The changes of the concentration of calcium ion were investigated by Fura-3 AM, and the expressions of Caspase-3 and ERK were detected by Western blotting assay. RESULTS:Ouabain and cinobufogenin inhibited the activity of Na+/K+-ATPase in HepG2 cell. Hoechst 33342 fluorescent staining showed that the clear apoptotic characteristics of HepG2 cell. The results of SCGE demonstrated that DNA double-strand of HepG2 cell was injured by ouabain and cinobufogenin. The concentration of calcium ion also increased after the treatment of ouabain and cinobufogenin according to the result of Fura-3 AM assay. The results of Western blot proved that ouabain and cinobufogenin promoted the production of p-ERK and the activition of caspase-3. CONCLUSION:Ouabain and cinobufogenin can induce the apoptosis of HepG2 cell and DNA damage, the increase of free calcium concentration and the activity of Caspase-3. ERK signaling pathway plays an important role in the apoptosis of HepG2 induced by ouabain and cinobufogenin.