Study on the Change of Iron Transporter Expression in k562 Cells Apoptosis Induced by Artesunate

Background and objective The antitumor effect of artemisinin was widely reported in vivo and ex vivo, but the possible mechanisms have yet been poorly understood. As is the anti-malaria effect of artemisinin, the tumour cells have the richer iron than the normal cells, which maybe result to be sensitive to artemisinins. Moreover, it is a very interesting question whether artesininin and its derivates can affect the iron metabolism of tumor cells and finally result to the antitumor effect. So in this study, we investigate the expression change of iron transporter including TfR1 (transferrin receptor-1, CD71), DMT1(Divalent metal transporter 1) and FPN1(Ferroportin 1) in the wild-type K562 cell(K562-W) and imatinib-resistant K562 cell(K562-R) treated with artesunates.

Methods The K562-W cells and K562-R cells were treated with 4×10⁻⁵ mol/L, 2×10⁻⁴ mol/L and 1×10⁻³ mol/L artesunate for 24, 48 and 72 hours respectively. The control groups include the untreated group and the group treated with desferrioxamine(20 umol/L). The different expressions of TfR1, DMT1 and FPN1 between the groups treated with artesunates and the control groups were analyzed by Semi-quantitative RT-PCR, Western-Blotting or flow cytometry. Apoptosis was measured by Annexin V-PI and Hochest 33258 staining.

Results The artesunate can induce K562-W and K562-R cells apoptosis in time- and concentration-dependent manner. When K562-W cells treated with 4×10⁻⁵ mol/L, 2×10⁻⁴ mol/L and 1×10⁻³ mol/L of artesunate for 48 hours, the mRNA expression of TfR1 is lower in artesunate groups than in the untreated groups. DMT1 mRNA expression in low concentration of artesunate and desferrioxamine group is lower than untreated group, and there is no expression of DMT1 mRNA in middle and high concentration artesunate group. The expression of FPN1 mRNA in artesunate group and desferrioxamine group is lower than the untreared group. The positive rate of CD71 is lower in artesunate groups than untreated group by flow cytometry analysis. The expression of DMT1 protein in desferrioxamine group is lower than untreated group and no expression of DMT1 protein in artesunate groups by Western blot. The TfR1 mRNA expression of K562-R cells treated with 4×10⁻⁵mol/L, 2×10⁻⁴mol/L and 1×10⁻³mol/L artesunate for 48 hours is lower in artesunate groups than untreated group by Semi-quantity RT-PCR. DMT1 mRNA expression in low concentration artesunate group is lower than untreated group, and there is no expression of +IRE-DMT1 mRNA in middle and high concentration artesunate group. The expression of FPN1 mRNA in artesunate group is lower than that in the untreated group. The positive rate of CD71 is lower in artesunate groups than untreated group by flow cytometry analysis. There is no expression of DMT1 protein in middle concentration artesunate groups by Western blot.

Conclusions Artesunate may reduce TfR1 and DMT1 expression in mRNA and/or protein level in K562-W and K562-R and lead to reduce the iron intake, iron transportation from the endosome to the cytoplasm and iron utilization in the mitochondria. Furthermore, Artesunate also reduce the expression of FPN1 mRNA to inhibit the outflow of iron. In addition to the generation of free radicals inducing to cell death, Artesunate can inhibit cell proliferation and induce cell apoptosis by the regulation of iron transporter.