乙酰紫草素对黑色素瘤A375细胞增殖和凋亡作用研究

Effect of acetyl shikonin on the proliferation and apoptosis of malignant melanoma A375 cell

目的研究乙酰紫草素对黑色素瘤A375细胞增殖和凋亡的影响。方法采用CCK-8法观察不同浓度乙酰紫草素对A375细胞的增殖抑制情况;在倒置显微镜下观察乙酰紫草素处理48 h后,A375细胞形态的变化;采用Hoechst 33342荧光染色法在荧光显微镜下观察乙酰紫草素干预后A375细胞的凋亡现象;流式细胞术检测乙酰紫草素对A375细胞凋亡水平的影响;采用Western blot法检测乙酰紫草素处理A375细胞后,凋亡相关蛋白Bax、Bcl-2、Cleaved caspase-9的表达。结果乙酰紫草素能明显抑制A375细胞的增殖,且呈时间依赖性与剂量依赖性;形态学观察与流式细胞术以及Hoechst 33342荧光染色法结果显示,与对照组比较,乙酰紫草素低、中、高剂量组A375细胞凋亡率升高,差异有统计学意义(P<0.05);Western blot结果显示,与对照组相比,乙酰紫草素低、中、高剂量A375细胞中Bcl-2表达降低,Bax、Cleaved caspase-9表达升高,Bax/Bcl-2比值增加,差异有统计学意义(P<0.05)。结论乙酰紫草素能够呈剂量依赖性抑制A375细胞的增殖,上调促凋亡蛋白Bax、Cleaved caspase-9的表达,抑制抑凋亡蛋白Bcl-2蛋白的表达,促进A375细胞的凋亡。

Objective To study the effect of acetyl shikonin on the proliferation and apoptosis of Melanoma A375 cells.Methods The human melanoma A375 cells were treated with different concentrations of acetyl shikonin,and the proliferation inhibition of A375 cells was observed by CCK-8 assay.The morphological changes of A375 cells were observed under an inverted microscope.Hoechst 33342 fluorescence staining method was used to observe the apoptosis of cells.Flow cytometry was used to test the apoptosis.Western blotting was used to detect the expression of apoptosis related proteins Bax,Bcl-2 and Cleaved caspase-9 expression level.Results Acetyl shikonin significantly inhibited the proliferation of A375 cells dose-and time-dependently.Compared with the control group,the apoptosis rate of A375 cells in the low,medium and high dose of acetyl shikonin groups was increased,with statistical significance(P<0.05).Compared with the control group,the expression of Bcl-2 was decreased,Bax and Cleaved caspase-9 was increased in A375 cells with low,medium and high doses of acetyl shikonin,and Bax/Bcl-2 ratio was increased,all with statistical significance(P<0.05).Conclusion Acetyl shikonin can inhibit the proliferation of A375 cells,up-regulate the expression of Bax and Cleaved caspase-9,and inhibit Bcl-2,thereby promote the apoptosis of A375 cells.