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[目的]研究单壁碳纳米管(single-walled carbon nanotubes,SWCNTs)对人源支气管上皮细胞株(BEAS-2B)氧化应激和细胞凋亡的影响。[方法]采用生长状态良好的BEAS-2B,分别以含0、10、50、100、200μg/mL SWCNTs的培养液处理24、48h。采用四甲基偶氮唑盐(MTT)比色法检测SWCNTs对BEAS-2B细胞活力的影响;采用荧光探针2’,7’-二氯荧光素二乙酸(DCFH-DA)进行活性氧(ROS)检测;采用黄嘌呤氧化酶法测定超氧化物歧化酶(SOD)活性;采用可见光法测定过氧化氢酶(CAT)活性;采用硫代巴比妥酸(TBA)法测定丙二醛(MDA)的含量;采用二硫代二硝基苯甲酸(DTNB)法测定谷胱甘肽过氧化物酶(GSH-Px)活性;采用阳离子荧光染料罗丹明123测定线粒体膜电位;采用pNA法检测细胞内caspase-3的蛋白活性;Real-time PCR测定细胞内bax和bcl-2基因的表达变化。[结果]当处理浓度大于10μg/mL时,SWCNTs可诱导BEAS-2B细胞存活率下降,细胞内ROS含量增加,线粒体膜电位减低,MDA升高,抗氧化酶系(SOD、CAT、GSH-Px)酶活性降低,caspase-3蛋白活性增加,bax在转录水平上表达量增加,bcl-2表达量降低,各实验结果均呈效应随浓度变化的趋势(P<0.05)。[结论]SWCNTs能够诱导BEAS-2B细胞的氧化损伤和细胞凋亡。
[Objective] To investigate the effects of single-walled carbon nanotubes (SWCNTs) on oxidative stress and apoptosis in human bronchial epithelial cell line (BEAS-2B). [Method] The BEAS-2B with good growth condition was treated with 0, 10, 50, 100 and 200 μg / mL SWCNTs for 24 and 48 h respectively. The effect of SWCNTs on the viability of BEAS-2B cells was detected by MTT colorimetric assay. The effects of SWCNTs on the viability of BEAS-2B cells were examined by using fluorescent probe 2 ’, 7’-dichlorofluorescein diacetic acid (DCFH-DA) ROS). The activity of superoxide dismutase (SOD) was measured by xanthine oxidase method. The activity of catalase (CAT) was measured by visible light method. The content of malondialdehyde (MDA) was measured by thiobarbituric acid MDA). The glutathione peroxidase (GSH-Px) activity was measured by DTNB method. Mitochondrial membrane potential was measured by cationic fluorescent dye rhodamine 123. The pNA assay was used to detect the content of glutathione peroxidase The protein expression of caspase-3 in the cells was detected by real-time PCR. The expression of bax and bcl-2 in the cells was detected by Real-time PCR. [Results] The results showed that SWCNTs could decrease the viability of BEAS-2B cells, increase the content of intracellular ROS, decrease the mitochondrial membrane potential and increase the MDA levels at the concentrations above 10μg / mL. The activities of antioxidant enzymes SOD, CAT, GSH-Px ) Enzyme activity decreased, caspase-3 protein activity increased, bax expression increased at the transcription level, bcl-2 expression decreased, the experimental results showed a trend with the effect of concentration (P <0.05). [Conclusion] SWCNTs can induce oxidative damage and apoptosis in BEAS-2B cells.