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目的探讨穿心莲内酯(AD)对人皮肤基底细胞癌A431细胞生长、凋亡及增殖细胞核抗原(PCNA)蛋白表达的影响。方法应用酸性磷酸酶(APA)法检测细胞增殖抑制率,荧光显微镜观察细胞形态,Annexin V-FITC/PI双染法、流式细胞术(FCM)检测细胞凋亡率,Rh123染色FCM检测细胞线粒体膜电位,免疫细胞化学法检测细胞PCNA蛋白表达。结果 AD呈时间、剂量依赖性抑制A431细胞生长增殖;且同一时间点50mg/L、100mg/LAD组与溶媒对照组相比差异显著(P<0.05)。AD组作用A431细胞24h时,部分细胞核染色质出现典型凋亡形态学改变。不同浓度AD作用A431细胞24h,早期凋亡率、晚期凋亡及坏死率均随AD浓度升高而逐渐增加,且50mg/L、100mg/L AD组与溶媒对照组相比差异显著(P<0.05)。AD作用A431 24h细胞内PCNA蛋白随AD浓度升高表达逐渐减少。AD作用A431细胞24h后,线粒体膜电位下降明显,与溶媒对照组相比较,50mg/L、100mg/LAD组差异有统计学意义(P<0.05)。结论 AD可明显抑制A431细胞生长增殖,其抑制细胞增殖可能与下调PCNA蛋白表达有关。AD能诱导A431细胞凋亡,其凋亡机制可能与细胞线粒体膜电位下降有关。
Objective To investigate the effects of andrographolide (AD) on the growth, apoptosis and the expression of proliferating cell nuclear antigen (PCNA) protein in human dermal basal cell carcinoma A431 cells. Methods The inhibitory rate of cell proliferation was detected by acid phosphatase (APA) method. The cell morphology was observed by fluorescence microscopy. The apoptosis rate was detected by Annexin V-FITC / PI double staining and flow cytometry (FCM) Membrane potential and immunocytochemistry were used to detect the expression of PCNA protein. Results AD inhibited the growth and proliferation of A431 cells in a dose-and time-dependent manner. At the same time point, there was a significant difference (P <0.05) between 50 mg / L and 100 mg / L AD groups. A4 group of AD cells treated 24h, part of the typical nuclear chromatin apoptosis morphological changes. The effect of different concentrations of AD on A431 cells 24h, early apoptosis rate, late apoptosis rate and necrosis rate increased with the increase of AD concentration gradually, and 50mg / L, 100mg / L AD group compared with the vehicle control group significant difference (P < 0.05). The effect of AD on A431 cells 24h PCNA protein decreased with the increase of AD concentration. After 24 hours, the mitochondrial membrane potential of A431 cells decreased significantly. Compared with the vehicle control group, the difference was statistically significant (P <0.05) between 50mg / L and 100mg / LAD groups. Conclusion AD can significantly inhibit the growth and proliferation of A431 cells, and its inhibition of cell proliferation may be related to the down-regulation of PCNA protein expression. AD can induce apoptosis in A431 cells, which may be related to the decrease of mitochondrial membrane potential.