陸杰， 應佳麗， 張婷*
（ 東南大學公共衛生學院，環境醫學工程教育部重點實驗室，江蘇省生物材料與器件重點實驗室，南京 210009； ）
摘要： 目的：探討γ-Fe2O3和納米氧化鋅顆粒（ZnO NPs）對人肝癌細胞（HepG2）的毒性作用及其影響因素，為評價納米金屬氧化物對生物體的肝毒性提供基本的科學依據。方法：利用透射電鏡和馬爾文激光粒度儀表征兩種納米材料的尺寸，水合粒徑和Zeta電位；利用原子吸收光譜法和火焰原子吸收法分別測定培養基中金屬氧化物的含量和細胞內鐵元素和鋅元素的含量；采用針對鐵離子和鋅離子的熒光探針方法檢測細胞內鐵離子和鋅離子的含量；MTT法檢測兩種納米材料對HepG2細胞活性的影響；Annexin-V/PI雙染法檢測細胞凋亡的情況；利用熒光探針DCFH-DA檢測細胞活性氧（ROS）的含量。結果：電鏡表征結果顯示Fe2O3和ZnO的粒徑分別為8.70±2.6nm和22.71±4.70nm；培養基中金屬離子隨著染毒濃度增高而升高，但隨時間延長而減少；細胞內兩種元素含量則具有正向的時間和劑量依賴性關系。MTT結果顯示兩種金屬氧化物均具有劑量依賴性關系，并且同等濃度下ZnO NPs抑制率比Fe2O3高，這與細胞凋亡的結果相一致。ROS檢測結果顯示ZnO組細胞ROS的含量具有明顯劑量依賴性關系，而Fe2O3組ROS含量隨劑量增大而增大，但趨勢不如ZnO組明顯。結論：兩種金屬納米材料對HepG2細胞的毒性的可能原因是進入細胞后釋放金屬離子導致ROS水平升高所致，造成兩種金屬納米材料毒性差異的原因是組成化學物的化學性質不同，化學性質活潑者毒性越大，如更容易釋放金屬離子，而粒徑等因素并不是主要因素。
Lu Jie， Ying JiaLi， Zhang Ting*
（ Key Laboratory of Environmental Medicine Engineering, Jiangsu Key Laboratory for Biomaterials and Devices, School of Public Health, Southeast University, Nanjing 210009； ）
Abstract： Objective: To investigate the toxic effects of γ-Fe2O3 and ZnO nanoparticles (ZnO NPs) on human hepatocellular carcinoma cells (HepG2), and to provide basic scientific evidence for evaluating hepatotoxicity of nano-metal oxides on organism. Method: The size, hydration size and zeta potential of the two nanomaterials were characterized by transmission electron microscopy and Malvern laser particle size analyzer. The contents of metal oxides and intracellular iron contents were determined by atomic absorption spectrometry and flame atomic absorption spectrometry, respectively. The contents of iron and zinc in the cells were measured by the fluorescent probe method. The MTT assay was used to detect the effect of the two kinds of nano-materials on the activity of HepG2 cells. Annexin-V / PI double staining (ROS) was detected by fluorescent probe DCFH-DA. Results: The results of electron microscopy showed that the particle size of Fe2O3 and ZnO was 8.70 2.6nm and 22.71±4.70nm, respectively. The metal ions in the medium increased with the increase of the concentration of Fe2O3 and ZnO, but decreased with the prolongation of time. And the content was in a time-dependent and dose-dependent manner. MTT results showed that both metal oxides had a dose-dependent relationship, and the inhibitory rate of ZnO NPs was higher than that of Fe2O3 at the same concentration, which was consistent with the results of apoptosis. The results of ROS showed that the content of ROS in the cells of ZnO group was dose-dependent, while the content of ROS in Fe2O3 group increased with the dose increasing, but the trend was not as obvious as in ZnO group. Conclusion: The toxicity of the two metal nanomaterials to HepG2 cells was due to the release of metal ions into the cells and the increase of ROS level. The difference of toxicity between the two metal nanomaterials were attributed to the different chemical properties of the chemical constituents, The greater the toxicity of lively, such as easier to release metal ions, and particle size and other factors are not the main factors.
Keywords： Nano Toxicology; metal oxides; toxicity; Influencing factors