蛇床子素联合顺铂对人肺癌细胞的杀伤效应及机制

doi:10.3969/j.issn.1673-8640.2015.06.021

摘要/Abstract

摘要： 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、200 μmol/L)、顺铂(1.5、3.0、6.0 μmol/L)及两者联合(50 μmol/L蛇床子素+1.5 μmol/L顺铂)对NCI-H460细胞增殖的影响。采用膜联蛋白(Annexin Ⅴ)/碘化丙啶(PI)染色法、二氢乙啶(DHE)染色法检测蛇床子素(50 μmol/L)、顺铂(1.5 μmol/L)及两者联合(50 μmol/L蛇床子素+1.5 μmol/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(50 μmol/L)、顺铂(1.5 μmol/L)及两者联合(50 μmol/L蛇床子素+1.5 μmol/L顺铂)对NCI-H460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。50 μmol/L蛇床子素联合1.5 μmol/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。

关键词: 蛇床子素, 顺铂, 凋亡, 活性氧簇, 人肺癌细胞NCI-H460

Abstract: 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 and6.0 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.

Key words: Osthole, Cisplatin, Apoptosis, Reactive oxygen species, Human lung cancer cell NCI-H460

中图分类号:

R446.1

程丽霞, 张本宏. 蛇床子素联合顺铂对人肺癌细胞的杀伤效应及机制[J]. 检验医学, 2015, 30(6): 631-634.

CHENG Lixia, ZHANG Benhong. Anticancer activity and mechanism of osthole combined with cisplatin in human lung cancer cells[J]. Laboratory Medicine, 2015, 30(6): 631-634.

图/表 5

表1 不同浓度蛇床子素和顺铂对NCI-H460细胞增殖的抑制作用(x?±s)

组别	抑制率(%)
对照组	0
蛇床子素组(μmol/L)
50	15.4±2.2^*
100	41.8±4.9^*
200	78.2±5.5^*
顺铂组(μmol/L)
1.5	11.3±2.4^*
3.0	45.2±3.5^*
6.0	82.1±6.9^*

表2 50 μmol/L蛇床子素和1.5 μmol/L顺铂单独及联合作用对NCI-H460细胞增殖的抑制作用(x?±s)

组别	蛇床子素 (μmol/L)	顺铂 (μmol/L)	抑制率 (%)
对照组	0	0	0
蛇床子素组	50	0	16.1±2.6^*
顺铂组	0	1.5	12.8±2.7^*
蛇床子素+顺铂组	50	1.5	74.2±6. $1 *# Δ$

表2 50 μmol/L蛇床子素和1.5 μmol/L顺铂单独及联合作用对NCI-H460细胞增殖的抑制作用(x?±s)

组别	蛇床子素 (μmol/L)	顺铂 (μmol/L)	抑制率 (%)
对照组	0	0	0
蛇床子素组	50	0	16.1±2.6^*
顺铂组	0	1.5	12.8±2.7^*
蛇床子素+顺铂组	50	1.5	74.2±6. $1 *# Δ$

表3 50 μmol/L蛇床子素和1.5 μmol/L顺铂单独及联合作用对NCI-H460细胞ROS生成及凋亡的诱导效应(x?±s)

组别	蛇床子素 (μmol/L)	顺铂 (μmol/L)	ROS细胞阳性率(%)	凋亡率 (%)
对照组	0	0	1.8±0.4	2.9±0.8
蛇床子素组	50	0	12.6±1.5	14.3±1.8
顺铂组	0	1.5	10.8±2.0	9.4±1.6
蛇床子素+ 顺铂组	50	1.5	42.5±3.6^*#	49.7±4.9^*#

图1 DHE染色法检测蛇床子素、顺铂及二者联合作用后对NCI-H460细胞ROS产生的影响

表4 ROS对蛇床子素联合顺铂对诱导NCI-H460细胞凋亡的影响(x?±s)

组别	NAC (mmol/L)	蛇床子素 (μmol/L)	顺铂 (μmol/L)	ROS细胞阳性率 (%)	增殖抑制率 (%)	凋亡率 (%)
对照组	0	0	0	2.1±0.5	0	2.6±0.7
蛇床子素+顺铂组	0	50	1.5	46.7±4.1	77.2±6.9	51.3±4.2
NAC+蛇床子素+顺铂组	10	50	1.5	19.7±2.6^*	33.2±4.0^*	22.5±3.5^*

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