槲皮素对人黑素瘤A375细胞增生和凋亡的影响

目的探讨槲皮素对体外培养人黑素瘤A375细胞增生和凋亡的影响。方法 CCK8法检测0、20、40、80、160μmol/L浓度槲皮素作用于A375细胞24、48和72 h后细胞增生水平。用0、20、40、80、160μmol/L浓度槲皮素作用24 h后,以Annexin-V/PI法观察A375细胞凋亡,分别测定Caspase 3、Caspase 8和Caspase 9的活性,并以蛋白印迹法(Western blot)检测p53,Bax及Bcl-2的基因蛋白水平。结果与空白对照组比较,20、40、80、160μmol/L槲皮素作用24、48和72 h后均对A375细胞的增生有抑制作用,且有时间、浓度依赖性。0、20、40、80、160μmol/L槲皮素在作用24 h时,A375细胞早期凋亡率由(3.02±0.49)%分别上升至(11.20±1.68)%、(12.44±1.68)%、(24.55±0.58)%和(47.68±0.79)%,各组与对照组相比差异均有统计学意义(P0.05或0.01),80、160μmol/L作用组的Caspase 3,Caspase8,Caspase 9活性升高,且与对照组比较,差异均有统计学意义(P0.05或0.01)。p53及Bax的蛋白水平随药物浓度的增加而升高,Bcl-2的蛋白水平随药物浓度增加而降低。结论槲皮素对黑素瘤A375细胞具有抑制增生和诱导凋亡的作用,其作用机制可能与调节p53及Bcl-2/Bax基因有关。

Effects of quercetin on the proliferation and apoptosis of A375 human melanoma cells

ObjectiveTo study the effect of quercetin on the proliferation and apoptosis of A375 human melanoma cells. Methods Cell counting kit-8 (CCK8) was used to evaluate the proliferative activity of A375 cells which treated with quercetin of 0, 20, 40, 80, 160μmol/L for 24 h, 48 h and 72 h respectively. Subsequently, A375 cells that treated with the same method underwent double staining with annexin V and propidium iodide for the detection of cell apoptosis, and lfuorometric assay for the estimation of Caspase 3, Caspase 8 and Caspase 9 activity, and Western blot for the determination of the levels of p53, Bax and Bcl-2 proteins.ResultsWithin the investigated concentration and time ranges, quercetin signiifcantly inhibited the proliferative activity of A375 cells in a concentration-dependent and time-dependent manner. Compared with blank groups, a signiifcant increase was observed in the early apoptosis rate in A375 cells treated with quercetin of 0, 20, 40, 80, 160 μmol/L for 24 h (11.20±1.68)%, (12.44±1.68)%, (24.55±0.58)% and (47.68±0.79)% vs (3.02±0.49)%, (P<0.05 or 0.01), as well as in the activity of Caspase 3, 8 and 9 in A375 cells treated with quercetin of 80, 160μmol/L for 24 h (P<0.05 or 0.01). After 24 h treatment, the protein levels of p53 and Bax were elevated, but Bcl-2 was decreased in A375 cells with the increase of quercetin concentration.ConclusionQuercetin can inhibit the proliferation and induce the apoptosis of A375 cells, and the apoptosis-inducing effect may occur through modulating the express of p53 and Bcl-2/Bax genes.