纳米氧化硅对RAW264.7细胞的DNA损伤作用

doi:10.3969/j.issn.1004-616x.2016.04.001

摘要/Abstract

摘要： 目的：研究纳米氧化硅(SiO₂)对小鼠RAW264.7细胞的DNA损伤作用，探讨其可能的毒作用机制。方法：分别采用浓度为31.25、125、500 μg/mL的两种纳米SiO₂(平均粒径分别为20、60 nm)和500 μg/mL的微米SiO₂粒子悬液对RAW264.7细胞染毒24 h，彗星试验检测细胞DNA单链断裂情况，CASP软件分析彗星图像，观察指标为彗星率(r_彗星)、尾长(TL)、尾部DNA百分率(r_尾部DNA)和OT尾矩(OTM)；分别用硝酸还原酶法、TBA法、Fenton法、黄嘌呤氧化酸法、钼酸法、考马斯亮兰法测定各组染毒细胞匀浆上清液中细胞氧化应激指标丙二醛(MDA)、羟自由基(^·OH)、超氧阴离子(O₂·)、过氧化氢(H₂O₂) 、一氧化氮(NO)和蛋白含量。结果：与阴性对照组比较，500 μg/mL 微米SiO₂染毒组和125、500 μg/mL的两种纳米SiO₂染毒RAW264.7细胞时，r_彗星、TL、r_尾部DNA、OTM均升高(P<0.01)；且500 μg/mL微米SiO₂组的r_彗星、r_尾部DNA、OTM高于同浓度两个纳米SiO₂组(P<0.01)。两种纳米SiO₂和微米SiO₂染毒细胞内脂质过氧化产物MDA、^·OH、O₂·、NO及H₂O₂的水平均高于阴性对照组(P<0.01)。结论：两种纳米SiO₂与微米SiO₂均可引起RAW264.7细胞DNA断裂和氧化损伤，氧化应激自由基的大量产生在DNA损伤过程中可能起到一定的作用。

关键词: 纳米SiO₂, RAW264.7细胞, 自由基, 遗传毒性, 单细胞凝胶电

Abstract: OBJECTIVE: RAW264.7 cells were exposed to different doses ofsilica nanoparticles and micron silicon dioxide (SiO₂), and induction of DNA damage was investigated. METHODS: RAW264.7 cells were treated with 20 nm, 60 nm silica nanoparticles at dosage of 31.25-500 μg/mL and micron SiO₂ at dosage of 500 μg/mL for 24 h. DNA single-strand break was detected by the Comet single cell gel electrophoresis (SCGE) and all the indexes of the assay:rate of comet (r_comet), tail length (TL) of DNA, rate of tail DNA (r_{tail DNA}), and Olive tail moment (OTM) were analyzed by the Comet Assay Software Project (CASP). In addition, levels of malondialdehyde (MDA), hydroxyl radicals (^·OH), superoxide anion radical (O₂·), hydrogen peroxide (H₂O₂) and nitric oxide (NO) in homogenate supernatantfrom treatedcells were measured. RESULTS: r_comet, TL, r_{tail DNA} and OTM induced by microsized SiO₂ were significantly increased compared with the negative control at the concentration of 500 μg/mL (P<0.01). Cells exposed to 125 and 500 μg/mL of the two kinds of silica nanoparticles showed a distinct increase of r_comet, TL, r_{tail DNA} and OTM compared with the negative control (P<0.01). r_comet, r_{tail DNA} and OTM caused by microsized SiO₂ were significantly increased compared with the silica nanoparticles groups(P<0.01) at the same concentrations. The increase of lipid peroxidation products of MDA and free radicals of ^·OH, O₂·, H₂O₂ and NO were in different degrees the result from silica nanoparticle and micron SiO₂ in comparison to the negative control. CONCLUSION: Both silica nanoparticle and microsized SiO₂ can cause DNA single-strand break and oxidative injury in RAW264.7 cells. The generation of free radicals of oxidative stress may have played a role in induction of DNA-damage.

Key words: silica nanoparticle, RAW264.7 cells, free radical, DNA-damage, single cell gel electrophoresis

中图分类号:

Q355

杨红, 吴秋云, 劳灿山, 李明月, 李文超. 纳米氧化硅对RAW264.7细胞的DNA损伤作用[J]. 癌变·畸变·突变, 2016, 28(4): 249-254.

YANG Hong, WU Qiuyun, LAO Canshan, LI Mingyue, LI Wenchao. DNA-damage in RAW264.7 cells exposed to different sizes of silica nanoparticles[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2016, 28(4): 249-254.

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