基于γH2AX和微核的流式细胞法检测NNK与AαC遗传毒性

发布时间：2019-05-28 07:46

【摘要】：目的探讨烟草特有亚硝胺NNK和杂环胺AαC的遗传毒性,对γH2AX焦点与微核分析结果进行比较。方法以0、25、50、100、200和400μg/ml的NNK,0、2.5、5、10、20和40μg/ml的AαC分别染毒中国仓鼠卵巢细胞(CHO细胞)24 h后,采用MTT试验检测细胞存活率,采用流式细胞仪检测NNK和AαC各剂量诱导的γH2AX焦点率和微核率,并比较γH2AX焦点率与微核率的差异。结果NNK各剂量的γH2AX焦点率分别为:(1.98±0.15)%、(2.28±0.04)%、(2.86±0.13)%、(3.14±0.09)%、(3.88±0.21)%和(4.34±0.20)%。与阴性对照相比,NNK剂量为25μg/ml时,γH2AX焦点率的差异有统计学意义(P0.05)。AαC各剂量的γH2AX焦点率分别为:(3.81±0.15)%、(3.24±0.25)%、(3.47±0.21)%、(3.90±0.14)%、(6.12±0.14)%和(11.98±0.24)%。与阴性对照相比,AαC剂量为10μg/ml时γH2AX焦点率的差异有统计学意义(P0.05)。NNK各剂量的微核率分别为:(1.50±0.22)%、(1.76±0.44)%、(3.26±0.27)%、(4.01±0.88)%、(7.50±0.70)%和(14.67±1.08)%;与阴性对照相比,NNK剂量为50μg/ml时微核率的差异有统计学意义(P0.05)。AαC各剂量微核率分别为:(1.42±0.50)%、(1.47±0.43)%、(1.42±0.35)%、(1.88±0.15)%、(5.88±1.15)%和(10.67±1.82)%。与阴性对照相比,AαC剂量为20μg/ml时微核率的差异有统计学意义(P0.05)。结论一定剂量的NNK和AαC可以诱发体外遗传毒性;与流式微核方法相比,流式γH2AX方法灵敏度更高。
[Abstract]:Objective to investigate the genotoxicity of tobacco specific nitrosamine NNK and heterocycloamines A 伪 C, and to compare the results of gamma H2AX focus with micronucleus analysis. Methods Chinese hamster ovarian cells (CHO cells) were exposed to NNK,0,2.5,5,10,20 of 0, 25, 50100200 and 400 渭 g / ml and A 伪 C of 40 渭 g / ml for 24 h, respectively. the survival rate of Chinese hamster ovarian cells was detected by MTT assay. The focal point rate and micronucleus rate of 纬 H2AX induced by NNK and A 伪 C were detected by flow cytometry, and the difference between focal point rate and micronucleus rate of 纬 H2AX was compared. Results the focal point rates of NNK were (1.98 卤0. 15)%, (2. 28 卤0. 04)%, (2. 86 卤0. 13)%, (3. 14 卤0. 09)%, (3. 88 卤0. 21)% and (4. 34 卤0. 20)%, respectively. Compared with the negative control, when the dose of NNK was 25 渭 g / ml, the focal point rate of 纬 H2AX was significantly different (P 0.05). The focal point rate of 纬 H2AX at each dose of A 伪 C was (3.81 卤0.15)%, (3.24 卤0.25)%, (3.47 卤0.21)%, respectively. (3.90 卤0.14)%, (6.12 卤0.14)% and (11.98 卤0.24)%. Compared with the negative control, there was significant difference in the focal point rate of 纬 H2AX when the dose of A 伪 C was 10 渭 g / ml (P 0.05). The micronucleus rate of each dose of NNK was (1.50 卤0.22)%, (1.76 卤0.44)% and (3.26 卤0.27)%, respectively. (4.01 卤0.88)%, (7.50 卤0.70)% and (14.67 卤1.08)%; Compared with the negative control, there was significant difference in micronucleus rate when the dose of NNK was 50 渭 g / ml (P 0.05). The micronucleus rate of each dose of A 伪 C was (1.42 卤0.50)%, (1.47 卤0.43)%, (1.42 卤0.35)%, respectively. (1.88 卤0.15)%, (5.88 卤1.15)% and (10.67 卤1.82)%. Compared with the negative control, there was significant difference in micronucleus rate when the dose of A 伪 C was 20 渭 g / ml (P 0.05). Conclusion certain doses of NNK and A 伪 C can induce genotoxicity in vitro, and the sensitivity of flow gamma H2AX method is higher than that of flow micronucleus method.
【作者单位】：中国烟草总公司郑州烟草研究院烟草化学重点实验室;军事医学科学院放射与辐射医学研究所放射毒理与辐射卫生防护研究室;上海烟草集团北京卷烟厂;
【基金】：国家科技支撑计划项目(2012BAK01B03) 郑州烟草研究院科技项目(322013CZ0600)
【分类号】：R99

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