盐酸小檗碱对人黑色素瘤A375-S2细胞增殖、凋亡的影响及其机制研究

Effects of Berberine Hydrochloride on the Proliferation and Apoptosis of Human Melanoma A375-S2 Cells and Underlying Mechanism

目的:探讨盐酸小檗碱对人黑色素瘤A375-S2细胞增殖、凋亡以及核因子κB(NF-κB)通路相关蛋白表达的影响。方法:体外培养A375-S2细胞,实验分为对照组(不添加任何药物处理)、盐酸小檗碱高、中、低剂量组(25,50,100μmol·L^(-1))、阳性对照组(顺铂,0.01 mmol·L^(-1))。采用CCK-8法检测细胞增殖情况;流式细胞仪检测细胞凋亡以及周期分布情况;免疫印迹法(WB)检测细胞中NF-κB的抑制蛋白α(IκB-α)、p-NF-κB、NF-κB、B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、剪切的半胱氨酸天冬氨酸蛋白酶3(cleaved-caspase3)蛋白表达情况。结果:与对照组相比,盐酸小檗碱不同剂量组A375-S2细胞增殖抑制率、凋亡率、G1期DNA量、IκB-α、Bax、cleaved-caspase3蛋白表达均显著升高,p-NF-κB、Bcl-2蛋白表达显著降低(P<0.05),且盐酸小檗碱高剂量组、阳性对照组与盐酸小檗碱中、低剂量组比较,差异有统计学意义(P<0.05),高剂量组与阳性对照组间比较差异无统计学意义(P>0.05)。结论:盐酸小檗碱可明显抑制人黑色素瘤A375-S2细胞增殖并诱导细胞凋亡,其可能与引起细胞周期G1期阻滞及抑制NF-κB信号通路激活有关。

Objective: To investigate the effects of berberine hydrochloride on the proliferation,apoptosis and expression of NFkappa B pathway-related proteins in human melanoma A375-S2 cells. Methods: A375-S2 cells were cultured in vitro and divided into control group( without any drug treatment),berberine hydrochloride high,medium and low dose groups( 25,50 and 100 μmol·L^(-1))and positive control group( cisplatin,0.01 mmol·L^(-1)). The cell proliferation was detected by CCK-8 assay;the cell apoptosis and cell cycle distribution were detected by flow cytometry;the expressions of IκB-α,p-NF-κB,NF-κB and B cell lymphoma-2( Bcl-2),Bcl-2 related X protein( Bax) and cleaved-caspase 3 protein were detected by Western blot( WB). Results: Compared with those in the control group,the inhibition rate,apoptosis rate,G1 DNA,protein expressions of IκB-α,Bax and cleaved-caspase3 of A375-S2 cells in different dose groups of berberine hydrochloride were significantly increased,and the protein expressions of p-NF-κB and Bcl-2 were significantly decreased( P<0.05). Compared with berberine hydrochloride high dose group and the positive control group,the differences were statistically significant in berberine hydrochloride medium and low dose groups( P< 0.05). There were no significant differences between berberine hydrochloride high dose group and the positive control group( P> 0.05). Conclusion: Berberine hydrochloride can significantly inhibit the proliferation and induce apoptosis of human melanoma A375-S2 cells,which may be related to arrest of cell cycle G1 phase and inhibition of activation of NF-κB signaling pathway.