摘要：纳米氧化锌(ZnO NPs)是应用前景广泛的一种新型多功能无机材料，通过呼吸道进入生物机体并对生物体造成损伤。茶多酚是一类能有效抑制或延缓氧化的物质，具有减缓脂质氧化反应等方面的作用，但其针对ZnO NPs诱导的呼吸道毒性的保护作用研究尚不广泛。因此本文选用大鼠气管上皮（RTE）细胞为实验对象，30nm和90nm纳米氧化锌为目标污染物，研究典型茶多酚物质表没食子儿茶素没食子酸酯（EGCG）的抗氧化效应。采用离体细胞体外诱导和暴露培养方法，通过CCK8（Cell Counting Kit-8）检测细胞增殖抑制率，同时测定活性氧（ROS）和丙二醛（MDA）水平对细胞毒性进行测定。结果显示，纳米氧化锌材料诱导RTE细胞毒性呈现浓度-依赖效应与粒径-依赖效应，30nm纳米氧化锌暴露组在高浓度（10mg/L）时，其细胞抑制率、ROS以及MDA水平分别是对照组的6.7倍、2.7倍和2.8倍，同时在1mg/L浓度暴露下，30nm组的细胞抑制率约为90nm组的3倍；而添加高浓度EGCG（1mg/L）后，与纳米氧化锌诱导对照组相比，90nm组细胞抑制率与MDA水平显著降低到对照组的4.5倍和2倍，30nm组ROS水平降低到对照组的1.6倍。以上结果均显示EGCG这种天然抗氧化剂具有良好的抗氧化作用。

ZnO NPs are a new type of multi-functional inorganic material with a very wide application prospect, which can enter into the organism bodies and cause toxic damage. Tea polyphenols is a class of substances that can effectively inhibit or delay oxidation, with the role of slowing the lipid oxidation reaction. However, there are lack of researches about the protective action of polyphenols to the toxicity induced by ZnO NPs in air way. In this paper, rat tracheal epithelial cells (RTE cells) were selected as experimental materials and ZnO NPs as the target pollutant to find out the toxic effect and  the antioxidant of epigallocatechin-3-gallate (EGCG), a typical tea polyphenol was utilized to identify the protective mechanism in RTE cells. The cell proliferation inhibition rate which was assayed by CCK8 (Cell Counting Kit-8), reactive oxygen species (ROS) detection and malondialdehyde (MDA) assay were detected. The results showed that the cytotoxicity in RTE cells induced by ZnO NPs was concentration-dependent and size-dependent. When the concentration reached to 10mg/L, the cell inhibitory rate, ROS and MDA levels of  ZnO NPs (30nm) exposure group was 6.7 times, 2.7 times and 2.8 times of the control group，respectively. After adding high concentration of EGCG (1mg/L), compared with nano-zinc-induced control group, the inhibition rate and MDA level in 90nm group were significantly decreased to 4.5-fold and 2-fold of the control group, and the ROS level in 30nm group was decreased to 1.6-fold .The above results showed that EGCG  protected RTE cells from the toxicity induced by ZnO NPs.

关键词：纳米氧化锌； RTE细胞； EGCG； 氧化损伤； 保护机制

Keyword: ZnO NPs; RTE cells; EGCG; oxidative damage; protective mechanism

目    录

1 引言 1

2 材料与方法 2

2.1 实验材料 2

2.1.1 试验细胞 2

2.1.2 毒素和试剂 2

2.2 实验方法 2

2.2.1 细胞培养与诱导 2

2.2.2 Cell Counting Kit-8（CCK8）测定及原理 2

2.2.3 活性氧ROS测定及原理 3

2.2.4 丙二醛MDA测定及原理 3

3 结果 3

3.1 不同粒径的纳米氧化锌对RTE细胞生长抑制率的影响 3

3.2 不同粒径的纳米氧化锌对RTE细胞ROS水平的影响 4

3.3 不同粒径的纳米氧化锌对RTE细胞MDA水平的影响 5

3.4 不同浓度的EGCG对纳米氧化锌暴露RTE细胞生长抑制率的影响 6

3.5 不同浓度的EGCG对纳米氧化锌暴露的RTE细胞ROS水平的影响 7

3.6 不同浓度的EGCG对纳米氧化锌暴露的RTE细胞MDA水平的影响 8

4 结论与讨论 10

参考文献 12

致谢 13