青蒿琥酯抑制FOXP3基因对K562/ADR细胞增殖、凋亡及多药耐药的影响

目的 探讨青蒿琥酯（Artesunate）抑制FOXP3基因表达对慢性髓系白血病（CML）耐阿霉素（ADR）细胞株 K562/ADR增殖、凋亡及多药耐药的影响，并探讨其作用机制。方法 实时荧光定量PCR（Real-time PCR）技术检测 FOXP3 mRNA在K562和K562/ADR细胞中的表达；蛋白印迹法（Western blot）检测FOXP3蛋白的表达；青蒿琥酯不同 质量浓度（2.5、5.0、7.5、10.0、12.5 mg/L）处理K562/ADR细胞24 h，利用CCK-8法检测不同浓度青蒿琥酯对K562/ADR 细胞的细胞毒性，筛选无细胞毒性浓度的青蒿琥酯完成后续实验；RT-PCR法、Western blot法检测无细胞毒性浓度青 蒿琥酯处理下FOXP3 mRNA、FOXP3蛋白表达变化；CCK-8法检测ADR对细胞毒性的变化；流式细胞术（FCM）检测 ADR平均荧光强度，即蓄积浓度变化。结果 FOXP3基因在CML耐药细胞株K562/ADR中表达升高；无毒剂量浓度 青蒿琥酯（2.5、5.0、7.5 mg/L）处理下K562/ADR细胞FOXP3 mRNA、蛋白表达受抑制（P＜0.05）；K562/ADR细胞胞内 ADR毒性增强，浓度升高（P＜0.05）。结论 FOXP3基因在CML K562/ADR耐药细胞株中高表达；青蒿琥酯可以通过 抑制FOXP3基因表达，增加K562/ADR胞内ADR浓度，逆转多药耐药，且呈一定的剂量依赖性。

Effects of artesunate inhibiting the expression of FOXP3 on proliferation,apoptosis andmultidrug resistance of K562/ADR cells

Objective To investigate the effect of artesunate inhibiting the expression of FOXP3 on proliferation, apoptosis and multidrug resistance of adriamycin (ADR) -resistant K562/ADR cells in chronic myeloid leukemia (CML), and to explore its mechanism. Methods The expressions of FOXP3 mRNA in K562 and K562/ADR cells were detected by real-time PCR. The expressions of FOXP3 proteins in K562 and K562/ADR cells were detected by Western blot assay. The K562/ADR cells were treated with different concentrations of artesunate (2.5, 5.0, 7.5, 10.0 and 12.5 mg/L) for 24 h. The toxicities of different concentrations of artesunate to K562/ADR cells were detected by CCK-8 assay, and the non-cytotoxic concentrations were screened. The expressions of FOXP3 mRNA and proteins in K562/ADR cells treated by non-cytotoxic concentration of artesunate were detected by RT-PCR and Western blot assay. The changes of toxicities of ADR in K562/ ADR cells were detected by CCK-8 assay. The average fluorescence intensities of ADR were detected by FCM assay. Results The expressions of FOXP3 were higher in K562/ADR cells than those in K562 cells. The mRNA and proteins expressions of FOXP3 were significantly lower in 2.5 mg/L group, 5 mg/L group and 7.5 mg/L group than those in the control group. The toxicities and concentrations of ADR were increased in K562/ADR cells treated by artesunate (both P＜0.05). Conclusion FOXP3 gene is highly expressed in adriamycin-resistant K562/ADR cells in CML. Artesunate can increase the concentrations of ADR and reverse multidrug resistance in K562/ADR cells by inhibiting the expression of FOXP3 in a dose-dependent manner.