Effects On Cytotoxicity And EGFP DNA Methylation Of Six Kinds Of Heavy Metal Compounds In Human BEL-7402 Liver Cancer Cells

Objective: Human liver cancer Bel-7402 cells were used to estimate the cell toxicity and demethylation toxicity of 6 compounds including Pb Cl2, Pb(CH3COO)2, Pb(NO3)2, Cu Cl2, Cu(CH3COO)2 and Cu(NO3)2.Method: Cell viability of different compounds was estimated firstly by MTS assay and their demethylation toxicity was quantified with a method established and reported already- quantification method of total demethylation potential of chemicals.(DNA methylase was used methylation modification for p EGFP-C3 plasmid of green fluorescent protein gene promoter region. Then p EGFP-C3 plasmid was transfected into Bel-7402 cells and the transfected cell were exposed with compound samples. Last, demethylation epigenetic toxicity of compound samples was assessed quantitatively based on green fluorescence intensity and the proportion of fluorescence positive cells QDMP evaluation method).Result: The results showed 6 kinds of heavy metal compounds all significantly inhibited the Bel-7402 cell proliferation. By paired t test, between the same heavy metal of different compounds, In addition to Pb(NO3)2 and Pb(CH3COO)2 effect on the cell survival rate are different, there were no significant difference of cell viability between other compounds. But the differences between different kinds of heavy metals had statistical significance, its inhibition for copper above lead, and there was significant difference of demethylation equivalency between lead compounds and copper compounds, the result were lead compounds higher than copper compounds. However significant demethylation epigenetic toxicity was observed for Pb Cl2, Pb(CH3COO)2, Pb(NO3)2, Cu Cl2, Cu(CH3COO)2, Cu(NO3)2 and their demethylation toxicity equivalency was 2.82E-03, 1.50E-03, 5.09E-04, 3.31E-05, 6.87E-05 and 3.45E-05 folds of 5-AZA-Cd R, respectively. No significant difference was observed between the same heavy metal of different compounds.Conclusion: The study revealed that lead and copper compounds can inhibit Bel-7402 cell proliferation and induce obvious demethylation effect on DNA methylation levels at promoter region of EGFP gene. Between the same heavy metal of different compounds, their demethylation equivalency was no significant. There was significant difference of demethylation equivalency between lead compounds and copper compounds, the result were lead compounds higher than copper compounds. There was significant difference of cell viability y between lead compounds and copper compounds, the result were lead compounds higher than copper compounds.