The Toxic Effects Of Organotins On HepG2 Cells And Their Mechanisms

Organotins are among the most widely used organometallic compounds. Their accumulation in environment is due to their wide use in agricultural and industrial applications. The toxicity of tribuyltin (TBT) and trimethyltin (TMT) have been confirmed in many cells. But the molecular mechanism of their toxicity has not elucidated clearly. In the present study, we treated HepG2 cells with TBT (0.5, 1, 2 and 4μM) and TMT (4,8, 16,32 and 64μM) , investigated their toxic effects on HepG2 cells and analyzed the DNA damage induced by them and the apoptosis-related proteins and enzymes, the results are as follows:1. The toxicity of TBT and TMT on HepG2 cellsIn MTT experiment, we found that the viability of HepG2 cells treated with TBT or TMT was inhibited in a concentration-dependent manner. The cell morphology was changed, including cell adhesion rate decreased significantly and the intracellular connections were destruct. The results of Hoechst 33342 stainining and flow cytometry experiments indicated that TBT and TMT could induce apoptosis and the apoptotic rate was concentration-dependent manner. But overall, the cytotoxicity of TBT is higher than that of TMT. It is reflected primarily that the vibility of cells treated with TBT 0.5μM for 3 h can be significantly inhibited (p < 0.5) , while we obtain the similar result with TMT 64μM for 24 h.2. The effect of TBT and TMT on DNA damageIn the comet assay experiment, we found that the tail length, tail DNA% and tail moment of HepG2 cells treated with TBT and TMT were all increased with the elevation of concentrations, there was an obvious concentration-dependent. From the results of the intracellular ROS levels detection, we observed the ROS levels in treated groups were increased with the elevation of the concentration of TBT and TMT and there was an obvious difference between 4μM TBT treated group and the control group (p<0.05) . It suggested that the DNA damage induced by TBT and TMT has association with the change of intracellular ROS levels.3. The effect of TBT and TMT on intracellular apoptosis-related proteinsThe results of the Western blot of intracellular apoptosis-related proteins treated by TBT and TMT suggested that the levels of Cytochrome C and the ratio of Bax and Bcl-2 were increased as the concentration of TBT and TMT increased, but the p53 was not concerned. The results of caspase-3, 8 and 9 activities indicated that the caspase-3 activity was increased with the concentration of elevation of TBT and TMT. In TBT treated groups the caspase-8 and 9 activities were increased with the contentration of elevation of TBT, but in the TMT treated groups, the caspase-8 activity was first increased then decreased and the caspase-9 activity was not evident. All these results indicated that both mitochondrial pathway and death receptor pathway take part in the TBT-induced apoptosis, while TMT induced apoptosis mainly through the death receptor pathway.