Institute for Environmental Sciences, University of Shizuoka , 52-1 Yada, Suruga-ku, Shizuoka-shi, Shizuoka 422-8526, Japan.
Environ Sci Technol. 2012 Jul 17;46(14):7629-36. doi: 10.1021/es300433x. Epub 2012 Jul 6.
We recently clarified that the side-scatter(ed) light (SSC) of flow cytometry (FCM) could be used as a guide to measure the uptake potential of nanoparticles [ Suzuki et al. Environ. Sci. Technol. 2007 , 41 , 3018 - 3024 ]. In this paper, the method was improved so as to be able to determine simultaneously the uptake potential of nanoparticles and the production of reactive oxygen species (ROS), and correlations with genotoxicity were evaluated. In the FCM analysis, SSC and fluorescence of 6-carboxy-2,7'-diclorodihydrofluorescein diacetate, di(acetoxy ester) based on ROS production were concurrently detected after treatments with ZnO, CuO, Fe(3)O(4), TiO(2), and Ag nanoparticles. The ZnO and CuO nanoparticles caused high ROS production, which was more significant in the cells with higher SSC intensity. The increase of SSC intensity was more significant for TiO(2) than ZnO and CuO, whereas ROS production was higher for ZnO and CuO than TiO(2), suggesting that the extent of ROS production based on the uptake of nanoparticles differed with each kind of nanoparticle. ROS production was correlated with generation of the phosphorylated histone H2AX (γ-H2AX), a marker of DNA damage, and an antioxidant, n-acetylcysteine, could partially suppress the γ-H2AX. This method makes it possible to predict not only uptake potential but also genotoxicity.
我们最近澄清了，流式细胞术（FCM）的侧向散射（ed）光可以作为一种指导，以测量纳米颗粒的摄取潜力[ Suzuki 等人。环境。Sci。技术。2007 年，41 期，3018-3024]。在本文中，该方法得到了改进，以便能够同时测定纳米颗粒的摄取潜力和活性氧物质（ROS）的产生，并评估与遗传毒性的相关性。在 FCM 分析中，用 ZnO、CuO、Fe(3)O(4)、TiO(2)和 Ag 纳米颗粒处理后，同时检测到基于 ROS 产生的 6-羧基-2,7'-二氯二氢荧光素二乙酸酯、二（乙酰氧基酯）的侧向散射（SSC）和荧光。ZnO 和 CuO 纳米颗粒引起了高 ROS 产生，在 SSC 强度较高的细胞中更为显著。与 ZnO 和 CuO 相比，TiO(2)的 SSC 强度增加更为显著，而对于 ROS 产生，ZnO 和 CuO 则高于 TiO(2)，表明基于纳米颗粒摄取的 ROS 产生程度因每种纳米颗粒而异。ROS 产生与磷酸化组蛋白 H2AX（γ-H2AX）的产生相关，γ-H2AX 是 DNA 损伤的标志物，抗氧化剂 N-乙酰半胱氨酸可以部分抑制 γ-H2AX。该方法不仅可以预测摄取潜力，还可以预测遗传毒性。
