蛇床子素联合顺铂对人肺癌细胞的杀伤效应及机制

Anticancer activity and mechanism of osthole combined with cisplatin in human lung cancer cells

目的探讨蛇床子素联合顺铂对人肺癌细胞NCI-H460的杀伤效应及机制。方法采用噻唑蓝(MTT)法检测蛇床子素(50、100、200μmol/L)、顺铂(1.5、3.0、6.0μmol/L)及两者联合(50μmol/L蛇床子素+1.5μmol/L顺铂)对NCI-H460细胞增殖的影响。采用膜联蛋白(AnnexinⅤ)/碘化丙啶(PI)染色法、二氢乙啶(DHE)染色法检测蛇床子素(50μmol/L)、顺铂(1.5μmol/L)及两者联合(50μmol/L蛇床子素+1.5μmol/L顺铂)对NCI-H460细胞凋亡及活性氧簇(ROS)的影响。用ROS抑制剂N-乙酰半胱氨酸(NAC)进行预处理,然后检测蛇床子素(50μmol/L)、顺铂(1.5μmol/L)及两者联合(50μmol/L蛇床子素+1.5μmol/L顺铂)对NCIH460细胞的凋亡诱导能力。以未加药物的NCI-H460细胞作为对照组。结果蛇床子素和顺铂均呈剂量依赖性地抑制NCI-H460细胞的增殖。50μmol/L蛇床子素联合1.5μmol/L顺铂可明显抑制NCI-H460细胞的增殖并诱导其发生凋亡。与蛇床子素及顺铂单独作用比较,二者联合作用后ROS阳性细胞明显增加(P<0.05)。与对照组(未用NAC预处理)比较,用NAC预处理过的NCI-H460细胞在用蛇床子素联合顺铂作用后产生的ROS明显降低(P<0.05),且凋亡率和对细胞增殖的抑制作用也同样明显降低(P<0.05)。结论蛇床子素联合顺铂可通过诱导ROS的产生,引起人肺癌细胞NCI-H460发生凋亡。

Objective To investigate anticancer activity and mechanism of osthole combined with cisplatin in human lung cancer cell NCI-H460. Methods NCI-H460 cells were treated with osthole（50,100 and 200 μmol / L）,cisplatin（ 1. 5,3. 0 and 6. 0 μmol / L） and osthole combined with cisplatin（ 50 μmol / L osthole ＋ 1. 5 μmol / L cisplatin）,and the cell proliferation was determined by 3-（4,5-dimethyl-2-thiazolyl）-2,5-diphenyl tetrazolium bromide（MTT） assay. NCI-H460 cells were treated with osthole（50 μmol/L）,cisplatin（1. 5 μmol/L）and osthole combined with cisplatin（ 50 μmol / L osthole ＋ 1. 5 μmol / L cisplatin）,and then the induction of apoptosis was determined by Annexin Ⅴ / propidium iodide（ PI） staining,and the generation of reactive oxygen species（ ROS） was determined by dihydroethidium（ DHE） staining. Furthermore,N-acetylcysteine（ NAC） was used to determine the apoptosis induced by above treatments. Untreated NCI-H460 cells were taken as control group. Results Osthole and cisplatin showed the dose dependent inhibition of the NCI-H460 cell proliferation. The 50 μmol / L osthole combined with 1. 5 μmol / L cisplatin can significantly inhibit NCI-H460 cell proliferation and induce its apoptosis. Compared to single treatments,combined treatment can significantly increase the ROS positive cells（ P〈0. 05）. Compared to the control group,NCI-H460 cells treated with NAC produced obviously the decreasing of ROS,apoptosis and cell proliferation（ P〈0. 05）. Conclusions Osthole combined with cisplatin can cause the apoptosis of NCI-H460 cells by inducing production of ROS.