响尾蛇神经毒素诱导人大细胞肺癌NCI-H661细胞凋亡机制研究

doi:10.3760/cma.j.issn.1673-422X.2015.07.001

摘要/Abstract

摘要： 目的观察南美响尾蛇神经毒素(crotoxin)诱导人大细胞肺癌NCIH661细胞的凋亡，并初步探讨其作用机制。方法通过四唑盐还原法(CCK8)检测crotoxin对NCIH661细胞的生长抑制率，应用细胞集落平板克隆实验观察crotoxin对NCIH661细胞集落形成的影响。实验分4组：阴性对照组、crotoxin组（60 μg/ml crotoxin作用24 h）、crotoxin+SB203580组（5 μmol/L SB203580预处理1 h后，60 μg/ml crotoxin作用24 h）、SB203580组(5 μmol/L SB203580预处理1 h后，完全培养液培养)，应用流式细胞术检测crotoxin对NCIH661细胞周期及细胞凋亡的影响，并检测以SB203580抑制p38MAPK活性后细胞周期及细胞凋亡率的变化。结果当crotoxin浓度≥30 μg/ml时对NCIH661细胞生长和集落形成有抑制作用，且抑制率随着作用时间和给药浓度的增加而上升；流式细胞术检测crotoxin对NCIH661细胞周期及细胞凋亡影响结果显示crotoxin组、crotoxin+SB203580组凋亡率分别为(16.70±1.38)%、(2.15±0.54)%，与对照组(1.47±0.29)%相比，前者差异具有统计学意义，后者差异无统计学意义(t=-18.763，P=0.000；t=-1.935，P=0.125)；crotoxin组、crotoxin+SB203580组G1期细胞分别为(57.25±1.09)%、 (48.04±1.03)%，与对照组(47.46±0.69)%相比，前者差异具有统计学意义，后者差异无统计学意义(t=-13.124，P=0.000；t=-0.809，P=0.464)。结论crotoxin对人大细胞肺癌NCIH661细胞凋亡有促进作用，这一作用可能与其激发p38MAPK信号通路活性有关。

关键词: 神经毒素类, 癌, 非小细胞肺, 细胞凋亡, p38MAPK

Abstract: ObjectiveTo observe the apoptosis of human large cell lung cancer NCIH661 cells induced by crotoxin, and to explore its mechanism. MethodsThe growth suppression of crotoxin on the NCIH661 cells was detected by CCK8 colorimetry, and the formation of NCIH661 cells was observed by the plat colony experiment. This experiment included 4 groups: negative control group, crotoxin group (60 μg/ml crotoxin acted for 24 h), crotoxin +SB203580 group (pretreated cells using 5 μmol/L SB203580 for 1 h, then 60 μg/ml crotoxin acted for 24 h), SB203580 group (pretreated cells using 5 μmol/L SB203580 for 1 h, then cultivated cells using complete culture solution). They were detected that the cell cycle and apoptosis rate of NCIH661 cells treated with crotoxin by the flow cytometry. Additionally, they were tested that the change of the cell cycle and apoptosis rate after the NCIH661 cells were treated with crotoxin and the activity of p38MAPK was inhibited by SB203580. ResultsWhen the concentration of crotoxin was greater than or equal to 30 μg/ml, the inhibitory effect of crotoxin on the activity of NCIH661 cells and colony formation, and inhibition rate rose with increasing function of time and drug concentration. Flow cytometry showed that the apoptosis rate of crotoxin group and crotoxin+SB203580 group were (16.70±1.38)% and (2.15±0.54)%, compared to the control group (1.47±0.29)%, and the former difference was statistically significant and the latter was not statistically significant(t=-18.763, P=0.000; t=-1.935, P=0.125). The G1 period cells of crotoxin group and crotoxin+SB203580 group were (57.25±1.09)% and (48.04±1.03)%, compared to the control group (47.46±0.69)%, and the former difference was statistically significant and the latter was not statistically significant(t=-13.124, P=0.000; t=-0.809, P=0.464). ConclusionCrotoxin can promote the apoptosis of human large cell lung cancer NCIH661 cells, and this effect may be related to the excitation of p38MAPK signal pathway.

Key words: Neurotoxins, Carcinoma, nonsmallcell lung, Apoptosis, p38MAPK

李睿,何靖康,申健,施臻,李威,于晓军. 响尾蛇神经毒素诱导人大细胞肺癌NCI-H661细胞凋亡机制研究[J]. 国际肿瘤学杂志, 2015, 42(7): 481-484.

Li Rui, He Jingkang, Shen Jian, Shi Zhen, Li Wei, Yu Xiaojun. Study on the mechanism of human large cell lung cancer NCIH661 cells apoptosis induced by crotoxin[J]. Journal of International Oncology, 2015, 42(7): 481-484.

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