二种晶型纳米二氧化钛诱导细胞氧化和遗传毒性的差异分析  

Differential analysis of cellular oxidation and genotoxicity induced by two crystalline forms of titanium dioxide nanoparticles

作　　者：徐云东 周滔[1,2] 倪娟[1,2] 汪旭[1,2,3] 王晗 XU Yun-dong;ZHOU Tao;NI Juan;WANG Xu;WANG Han(School of life sciences Yunnan normal university,Kunming,Yunnan 650500,China;Engineering Research Center of Sustainable Development and Utilization of Biomass Energy,Ministry of Education,Yunnan Normal University,Kunming,Yunnan 650500,China;Yeda Institute of Gene and Cell Therapy,Taizhou,Zhejiang 318000,China)

机构地区：[1]云南师范大学生命科学学院,云南昆明650500 [2]云南师范大学生物能源持续开发与利用教育部工程研究中心,云南昆明650500 [3]耶大基因与细胞治疗研究院,浙江台州318000

出　　处：《毒理学杂志》2024年第6期444-448,454,共6页Journal of Toxicology

基　　金：云南省教育厅科学研究基金项目(2020J0089);云南省科技厅科技计划项目(202101AU070047);云南师范大学博士科研启动项目;云南师范大学研究生科研创新基金项目。

摘　　要：目的探究金红石型纳米二氧化钛(TiO_(2)-NPs-R)及锐钛矿型纳米二氧化钛(TiO_(2)-NPs-A)对人正常肝细胞L-02的遗传毒性差异,并分析其可能机制。方法对数生长期的L-02细胞暴露于80μg/ml的TiO_(2)-NPs-R或TiO_(2)-NPs-A 48 h后,胞质分裂阻断微核细胞组试验(CBMN-Cyt)评估受试细胞的染色体不稳定性(CIN);酶标仪荧光法检测细胞内活性氧(ROS)和丙二醛(MDA)含量;流式细胞仪检测侧向散射角(SSC)检测细胞摄取颗粒数量;RT-qPCR法检测细胞膜转运蛋白相关基因小窝蛋白1(CAV-1)和网格蛋白重链(CLTC)的mRNA水平;紫外-分光光度计检测无细胞系中TiO_(2)-NPs-R和TiO_(2)-NPs-A悬液在505 nm波长下的吸光度(A)值评估颗粒沉降速率。结果与TiO_(2)-NPs-R相比,80μg/ml的TiO_(2)-NPs-A暴露48 h诱导L-02细胞产生更高的CIN(F=17.79,P<0.001)、ROS(F=50.49,P<0.001)、MDA(F=31.15,P<0.001)及SSC值(F=164.97,P<0.001),且伴随着CAV-1 mRNA表达水平的升高(F=33.01,P<0.001)。结论TiO_(2)-NPs-A因沉降速率较大,更易附着在L-02细胞表面,并诱导CAV-1基因表达水平升高,使其较TiO_(2)-NPs-R更易被摄取并诱导细胞产生更多的活性氧进而造成细胞氧化损伤,诱发染色体不稳定性升高。Objective To investigate the differences in genotoxicity of rutile-type titanium dioxide(TiO_(2)-NPs-R)and anatase nanotitanium dioxide(TiO_(2)-NPs-A)on human normal hepatocytes L-02 and to analyze the possible mechanisms.Methods After exposure of logarithmic growth stage L-02 cells to 80μg/ml of TiO_(2)-NPs-R or TiO_(2)-NPs-A for 48 h,the cytokinesis-blocked micronucleus cytome assay(CBMN-Cyt)was performed to assess the chromosomal instability(CIN)of the test cells.The intracellular reactive oxygen species(ROS)and malondialdehyde(MDA)levels were detected by enzyme-labeled fluorimetry.Flow cytometry was used to detect the number of particles taken up by the cells by measuring the side scatter(SSC);RT-qPCR was used to detect the mRNA levels of cell membrane transport protein-related genes,cavitin 1(CAV-1)and clathrin heavy chain(CLTC);UV-spectrophotometer was used to measure the TiO_(2)-NPs-R and TiO_(2)-NPs-A suspensions in cell-free lines at 505 nm A value to assess the particle sedimentation rate.Results Compared with TiO_(2)-NPs-R,80μg/ml of TiO_(2)-NPs-A exposure for 48 h induced higher CIN(F=17.79,P<0.001),ROS(F=50.49,P<0.001),MDA(F=31.15,P<0.001)and SSC values(F=164.97,P<0.001)in L-02 cells,accompanied by increased mRNA expression levels of CAV-1(F=33.01,P<0.001).Conclusion TiO_(2)-NPs-A,due to its higher sedimentation rate,is more likely to attach to the surface of L-02 cells and induce an increase in the expression level of CAV-1 gene,making it more susceptible to uptake and induce more reactive oxygen species production than TiO_(2)-NPs-R,which in turn causes oxidative damage to cells and induces increased chromosome instability.

关 键 词：金红石型纳米二氧化钛 锐钛矿型纳米二氧化钛 L-02细胞 细胞摄取 

分 类 号：R114[医药卫生—卫生毒理学] R99[医药卫生—公共卫生与预防医学]