Enhanced cytotoxic effects of arsenite in combination with anthocyanidin compound, delphinidin, against a human leukemia cell line, HL-60

Among five major anthocyanin compounds, delphinidin exhibited the most potent and selective cytocidal effect against HL-60, a trivalent arsenic (As(III))-resistant cell line. Co-treatment with delphinidin and As(III) resulted in the reduction of IC50 value for As(III) from 11.2 to 1.5 mu M, which was considered as clinically achieved concentrations of As(III). The combination treatment strongly preferred to selectively enhance the cytotoxicity of As(III) against HL-60 cells rather than human peripheral blood mononuclear cells. The induction of apoptosis as evidenced by the increase of sub-G1 cells, DNA fragmentation, annexin V-positive cells and the activation of caspase-8, -9 and -3 was observed in HL-60 cells co-treated with As(III) and delphinidin. Similar to the activation pattern of caspases, a substantial decrease in the expression level of Bid along with the loss of mitochondrial membrane potential was also observed. These results suggested that the combination treatment triggered a convergence of the intrinsic and extrinsic pathways of apoptosis via the activation of caspase-8 and cleaved Bid. Delphinidin itself significantly decreased the intracellular GSH ([i] GSH) and nuclear factor-kappa B (NF-kappa B) binding activity, and further returned As(III)-triggered increment of [i] GSH and enhancement of NF-kappa B binding activity to control level. Additionally, buthionine sulfoximine, a GSH depletor; JSH-23, a NF-kappa B inhibitor, also mimicked the capacity of delphinidin to significantly induce the reduction of [i] GSH along with the potentiation of As(III) cytotoxicity in HL-60 cells. These observations suggested that delphinidin-induced sensitization of HL-60 cells to As(III) was caused by the reduction of [i] GSH, which was probably associated with the inhibitory effect of delphinidin on NF-kappa B binding activity. These findings further suggest that delphinidin-induced sensitization of HL-60 cells to As(III) may lead to dose reduction of As(III) in clinical application, and ultimately contribute to minimizing its side effects.