采用体外染色体畸变试验检测锐钛型纳米二氧化钛的遗传毒性

doi:10.3969/j.issn.1004-616x.2022.01.013

摘要/Abstract

摘要： 目的： 按《OECD化学品测试准则473：体外哺乳动物染色体畸变测试》的要求进行试验，检测锐钛型纳米二氧化钛诱发体外培养的哺乳动物细胞染色体畸变的能力，以评价其是否属于致突变物。方法： 试验组受试物终浓度分别设置为0.007 8、0.031 2、0.125、0.5和2 mg/mL，同时设立溶剂对照组和阳性对照组。试验分为有代谢活化系统短时处理组（+S9，4 h）、无代谢活化系统短时处理组（-S9，4 h）和无代谢活化系统连续处理组（-S9，24 h）3种处理方式，将培养的中国仓鼠肺细胞（CHL）暴露于锐钛型纳米二氧化钛混悬液中，染毒相应时间后收获细胞，经低渗固定后，制片并染色。每个浓度组选取300个分散良好的中期分裂相细胞进行染色体畸变分析。结果： 锐钛型纳米二氧化钛在0.007 8、0.031 2、0.125、0.5和2 mg/mL浓度下，无代谢活化系统短时处理组不含裂隙的染色体畸变率分别为1.67%、1.33%、2.00%、1.00%和2.33%，有代谢活化系统短时处理组不含裂隙的染色体畸变率分别为1.33%、1.33%、2.00%、1.67%和2.00%，无代谢活化系统连续处理组不含裂隙的染色体畸变率分别为1.33%、1.67%、2.00%、1.67%和2.33%，与溶剂对照组相比，差异均无统计学意义（P>0.05）。结论： 在本实验条件下，锐钛型纳米二氧化钛在0.007 8~2 mg/mL浓度范围内，对CHL细胞染色体无致畸作用。

关键词: 锐钛型纳米二氧化钛, 体外哺乳动物细胞染色体畸变试验, 中国仓鼠肺细胞, 遗传毒性

Abstract: OBJECTIVE: According to the "OECD Guidelines for the testing of chemicals 473",the in vitro mammalian chromosomal aberration test was used to determine whether anatase titanium dioxide nanoparticles could induce chromosome aberrations was genotoxic or not. METHODS: The final concentrations of the test substance were set to be 0.007 8,0.031 2,0.125,0.5,and 2 mg/mL. In addition,a solvent control group and a positive control group were set up at the same time. The treatment included short-term treatment group with metabolic activation (+S9,4 h),short-term treatment group without metabolic activation (-S9,4 h) and continuous treatment group without metabolic activation (-S9,24 h). The cultured Chinese hamster lung cells (CHL) were exposed to anatase nano-titanium dioxide suspension,and the cells were harvested after a corresponding period of time,treated with hypotonic solution and then fixed. The cells were spread onto slides and stained. For each dose group,300 well-dispersed metaphase cells were selected for chromosomal aberration analysis. RESULTS: At the dose level of 0.007 8,0.031 2,0.125,0.5 and 2 mg/mL without metabolic activation,the induced chromosome aberration rates (excluding the gap) were 1.67%,1.33%,2.00%,1.00% and 2.33%,respectively. With metabolic activation,the rates were 1.33%,1.33%,2.00%,1.67% and 2.00%,respectively. After 24 hours treatment without metabolic activation,the rates were 1.33%,1.67%,2.00%,1.67% and 2.33%,respectively. Compared with the solvent control group,there was no statistically significant difference observed (P>0.05). CONCLUSION: Under the current test conditions,anatase titanium dioxide nanoparticles showed no evidence of genotoxic potential in the CHL at the tested dose levels of 0.007 8-2 mg/mL.

Key words: anatase titanium dioxide nanoparticles, in vitro mammalian chromosome aberration test, Chinese hamster lung cells, genotoxicity

中图分类号:

R394-33

杜秀明, 洪丽玲, 徐灵芝, 周庆云. 采用体外染色体畸变试验检测锐钛型纳米二氧化钛的遗传毒性[J]. 癌变·畸变·突变, 2022, 34(1): 67-71.

DU Xiuming, HONG Liling, XU Lingzhi, ZHOU Qingyun. In vitro chromosome aberration evaluation of anatase titanium dioxide nanoparticles[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2022, 34(1): 67-71.

参考文献

[1] CHEN T, YAN J, LI Y. Genotoxicity of titanium dioxide nanoparticles[J]. J Food Drug Anal, 2014, 22(1):95-104.
[2] FOWLER P, MEURER K, HONARVAR N, et al. A review of the genotoxic potential of 1, 4-naphthoquinone[J]. Mutat Re Genet Toxicol Environ Mutagen, 2018, 834:6-17.
[3] YANG H, ZHANG X, LIU H, et al. Lanthanum nitrate genotoxicity evaluation:Ames test, mouse micronucleus assay, and chromosome aberration test[J]. Mutat Res Genet Toxicol Environ Mutagen, 2016, 810:1-5.
[4] SHI H, MAGAYE R, CASTRANOVA V, et al. Titanium dioxide nanoparticles:a review of current toxicological data[J]. Part Fibre Toxicol, 2013, 10:15.
[5] CHARLES S, JOMINI S, FESSARD V, et al. Assessment of the in vitro genotoxicity of TiO₂ nanoparticles in a regulatory context[J]. Nanotoxicology, 2018, 12(4):357-374.
[6] TROUILLER B, RELIENE R, WESTBROOK A, et al. Titanium dioxide nanoparticles induce DNA damage and genetic instability in vivo in mice[J]. Cancer Res, 2009, 69(22):8784-8789.
[7] LINDBERG H K, FALCK G C M, CATALÁN J, et al. Genotoxicity of inhaled nanosized TiO₂ in mice[J]. Mutat Res Genet Toxicol Environ Mutagen, 2012, 745(1/2):58-64.
[8] SHUKLA R K, SHARMA V, PANDEY A K, et al. ROS-mediated genotoxicity induced by titanium dioxide nanoparticles in human epidermal cells[J]. Toxicol Vitro, 2011, 25(1):231-241.
[9] PETKOVIĆ J, ZEGURA B, STEVANOVIĆ M, et al. DNA damage and alterations in expression of DNA damage responsive genes induced by TiO₂ nanoparticles in human hepatoma HepG2 cells[J]. Nanotoxicology, 2011, 5(3):341-353.
[10] GUICHARD Y, SCHMIT J, DARNE C, et al. Cytotoxicity and genotoxicity of nanosized and microsized titanium dioxide and iron oxide particles in Syrian hamster embryo cells[J]. Ann Occup Hyg, 2012, 56(5):631-644.
[11] WANG S, HUNTER L A, ARSLAN Z, et al. Chronic exposure to nanosized, anatase titanium dioxide is not cyto- or genotoxic to Chinese hamster ovary cells[J]. Environ Mol Mutagen, 2011, 52(8):614-622.
[12] LORGE E, MOORE M M, CLEMENTS J, et al. Standardized cell sources and recommendations for good cell culture practices in genotoxicity testing[J]. Mutat Res Genet Toxicol Environ Mutagen, 2016, 809:1-15.
[13] DU X, GAO S, HONG L, et al. Genotoxicity evaluation of titanium dioxide nanoparticles using the mouse lymphoma assay and the Ames test[J]. Mutat Res Genet Toxicol Environ Mutagen, 2019, 838:22-27.
[14] MORITA T, HONMA M, MORIKAWA K. Effect of reducing the top concentration used in the in vitro chromosomal aberration test in CHL cells on the evaluation of industrial chemical genotoxicity[J]. Mutat Res Toxicol Environ Mutagen, 2012, 741(1/2):32-56.
[15] WOODRUFF R S, LI Y, YAN J, et al. Genotoxicity evaluation of titanium dioxide nanoparticles using the Ames test and Comet assay[J]. J Appl Toxicol, 2012, 32(11):934-943.
[16] WARHEIT D B, HOKE R A, FINLAY C, et al. Development of a base set of toxicity tests using ultrafine TiO₂ particles as a component of nanoparticle risk management[J]. Toxicol Lett, 2007, 171(3):99-110.
[17] SUZUKI T, MIURA N, HOJO R, et al. Genotoxicity assessment of titanium dioxide nanoparticle accumulation of 90 days in the liver of gpt delta transgenic mice[J]. Genes Environ, 2020, 42:7.
[18] THEOGARAJ E, RILEY S, HUGHES L, et al. An investigation of the photo-clastogenic potential of ultrafine titanium dioxide particles[J]. Mutat Res Toxicol Environ Mutagen, 2007, 634(1/2):205-219.
[19] 马茂才, 黄萍, 晏辉, 等. 纳米二氧化钛和纳米氧化锌的Ames试验[J]. 癌变·畸变·突变, 2010, 22(4):302-304.
[20] DI VIRGILIO A L, REIGOSA M, ARNAL P M, et al. Comparative study of the cytotoxic and genotoxic effects of titanium oxide and aluminium oxide nanoparticles in Chinese hamster ovary (CHO-K1) cells[J]. J Hazard Mater, 2010, 177(1/2/3):711-718.
[21] LU P J, HO I C, LEE T C. Induction of sister chromatid exchanges and micronuclei by titanium dioxide in Chinese hamster ovary-K1 cells[J]. Mutat Res, 1998, 414(1/2/3):15-20.
[22] HASHEM M M, ABO-EL-SOOUD K, ABD-ELHAKIM Y M, et al. The long-term oral exposure to titanium dioxide impaired immune functions and triggered cytotoxic and genotoxic impacts in rats[J]. J Trace Elem Med Biol, 2020, 60:126473.