纳米银颗粒及纳米二氧化钛颗粒的体外遗传毒性研究

doi:10.16288/j.yczz.20-161

遗传 ›› 2020, Vol. 42 ›› Issue (12): 1192-1200.doi: 10.16288/j.yczz.20-161

纳米银颗粒及纳米二氧化钛颗粒的体外遗传毒性研究

朱海美¹, 黄鹏程², 赵田田³, 周长慧^2,³, 李若婉³, 于春荣^2,³, 陈志勇², 顾林峰², 常艳^1,^2,³()

^1. 上海工程技术大学化学化工学院,上海 201620
^2. 上海益诺思生物技术股份有限公司,上海 201203
^3. 中国医药工业研究总院,上海 201201

收稿日期:2020-07-18 修回日期:2020-10-08 出版日期:2020-12-17 发布日期:2020-11-09
通讯作者: 常艳 E-mail:ychang@ncdser.com
作者简介:朱海美,在读硕士研究生,专业方向：毒理学方向,E-mail:1695051900@qq.com|黄鹏程,硕士,专业方向：毒理学方向,E-mail: 774121501@qq.com
朱海美和黄鹏程为并列第一作者。
基金资助:
“十三五”重大新药创制科技重大专项编号(2018ZX09201017-008);上海市科委研发公共服务平台专项资助编号(18DZ2290100)

In vitro genotoxicity study of silver nanoparticles and titanium dioxide nanoparticles

Haimei Zhu¹, Pengcheng Huang², Tiantian Zhao³, Changhui Zhou^2,³, Ruowan Li³, Chunrong Yu^2,³, Zhiyong Chen², Linfeng Gu², Yan Chang^1,^2,³()

^1. School of Chemistry and Chemical Engineer, Shanghai University of Engineering Science, Shanghai 201620, China
^2. Shanghai Innostar Bio-tech Co. Ltd, Shanghai 201203, China
^3. China State Institute of Pharmaceutical Industry, Shanghai 201201, China

Received:2020-07-18 Revised:2020-10-08 Online:2020-12-17 Published:2020-11-09
Contact: Chang Yan E-mail:ychang@ncdser.com
Supported by:
Supported by “the13th Five-Year plan” Major New Drug Innovation and Technology Major Project No(2018ZX09201017-008);the Shanghai Municipal Science and Technology Commission R&d Public Service Platform Project No(18DZ2290100)

摘要/Abstract

摘要：

纳米物质以其独特的物理性质广泛用于化妆品、医药和食品工业,但它们的安全性和遗传毒性仍然存在争议。本研究拟采用基于人成淋巴TK6细胞的体外彗星实验整合体外PIG-A基因突变实验对纳米银颗粒(AgNPs)和纳米二氧化钛颗粒(TiO₂NPs)的致DNA断裂作用和致突变性进行初步研究。本研究探究了最高至200 μmol/L浓度时,TK6细胞暴露于两种纳米物质4 h时的彗星实验和经过暴露24 h以及10 d基因突变表达期后的PIG-A基因突变结果。同时,本研究还检测了TK6细胞暴露于纳米颗粒4 h及24 h对纳米物质的摄取能力。在纳米物质摄取实验中,随着纳米物质浓度的升高,TK6细胞在流式细胞仪检测图中的侧向散射角(side scatter, SSC)呈现浓度与时间依赖性升高,表明TK6细胞可摄取两种纳米颗粒。在4 h彗星实验中,AgNPs可导致彗星尾DNA荧光%强度呈现浓度依赖性显著升高,为阳性结果。而TiO₂NPs则不能诱导彗星尾DNA荧光%强度呈现浓度依赖性显著升高,但最高浓度组尾DNA荧光%超过本实验室阴性历史对照数据,为可疑结果。而在体外PIG-A基因突变实验结果中,AgNPs和TiO₂NPs均不能诱导TK6细胞的PIG-A基因突变率出现阳性升高。AgNPs可导致TK6细胞出现DNA损伤,但不能诱导PIG-A基因突变率升高。TiO₂NPs既不能诱导TK6细胞出现DNA损伤,也不能诱导PIG-A基因突变率升高,TiO₂NPs的遗传毒性有待进一步验证。

关键词: 纳米银颗粒, 纳米二氧化钛颗粒, 彗星实验, PIG-A基因突变实验, 遗传毒性

Abstract:

Nanoparticles are widely used in cosmetic, pharmaceutical, and food industries, but their safety and genetic toxicity are still unclear. In this study, the genotoxicity of silver nanoparticles (AgNPs) and titanium dioxide nanoparticles (titanium dioxide nanoparticles) were evaluated by in vitro comet assay and PIG-A assay in TK6 cells. We exposed TK6 cells to two types of nanoparticles at the highest concentration of 200 μmol/L for 4 h and conducted the in vitro comet assay. We examined the mutation results of PIG-A gene in vitro after 4 h, 24 ho and 10 days of exposure, respectively. We also examined the endocytosis of nanoparticles in TK6 cells exposed to nanoparticles for 24 h. In the endocytosis assay, with the increase of nano-material concentration, the side scatter (SSC) of TK6 cells in flow cytometry showed a concentration-dependent and time-dependent increase, indicating that TK6 cells could uptake both types of nanoparticles. In the comet assay, AgNPs could induce a concentration-dependent increase in DNA tail intensity. However, titanium dioxide NPs could not induce the concentration-dependent increase of DNA fluorescence intensity of comet tail. In the PIG-A assay, both AgNPs and TiO₂NPs did not induce PIG-A gene mutation frequency in TK6 cells. The results showed that AgNPs could induce DNA damage in TK6 cells, but could not induce increase of PIG-A gene mutation frequency. TiO₂NPs neither induce DNA damage in TK6 cells nor increase PIG-A mutation frequency. Further tests are needed to determine whether TiO₂NPs are genotoxic.

Key words: silver nanoparticles, titanium dioxide nanoparticles, comet assay, in vitro PIG-A assay, genotoxicity

朱海美, 黄鹏程, 赵田田, 周长慧, 李若婉, 于春荣, 陈志勇, 顾林峰, 常艳. 纳米银颗粒及纳米二氧化钛颗粒的体外遗传毒性研究[J]. 遗传, 2020, 42(12): 1192-1200.

Haimei Zhu, Pengcheng Huang, Tiantian Zhao, Changhui Zhou, Ruowan Li, Chunrong Yu, Zhiyong Chen, Linfeng Gu, Yan Chang. In vitro genotoxicity study of silver nanoparticles and titanium dioxide nanoparticles[J]. Hereditas(Beijing), 2020, 42(12): 1192-1200.

图/表 6

表1

图1

图2

图3

图4

图5

参考文献 25

[1]	Tortella GR, Rubilar O, Durán N, Diez MC, Martínez M, Parada J, Seabra AB . Silver nanoparticles: toxicity in model organisms as an overview of its hazard for human health and the environment. J Hazard Mater, 2019,390:121974.
[2]	Calderón-Jiménez B, Johnson ME, Montoro Bustos AR, Murphy KE, Winchester MR, Vega Baudrit JR . Silver nanoparticles: technological advances, societal impacts, and metrological challenges. Front Chem, 2017,5:6.
[3]	Patel S, Patel P, Bakshi SR . Titanium dioxide nanoparticles: an in vitro study of DNA binding, chromosome aberration assay, and comet assay. Cytotechnology, 2017,69(2):245-263.
[4]	Zhang R, Wu AQ, Shi LL , A review of the performance evaluation and application of nano-silver antibacterial agents. J Yangtze Univ (Nat Sci Ed), 2015,12(25):23-25.
	张然, 吴安琪, 施伶俐 . 纳米银抗菌剂性能评价和应用现状综述. 长江大学学报(自科版), 2015,12(25):23-25.
[5]	Gamucci O, Bertero A, Malvindi MA, Sabella S, Pompa PP, Mazzolai B, Bardi G . Detection of fluorescent nanoparticle interactions with primary immune cell subpopulations by flow cytometry. J Vis Exp, 2014, ( 85):51345.
[6]	Singh NP, Mccoy MT, Tice RR, Schneider EL . A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res, 1988,175(1):184-191.
[7]	Olive PL, Banáth JP, Durand RE . Heterogeneity in radiation-induced DNA damage and repair in tumor and normal cells measured using the "comet" assay. Radiat Res, 1990,122(1):86-94.
[8]	Kwon JY, Lee SY, Koedrith P, Lee JY, Kim KM, Lee JK, Jeong J, Maeng EH, Lee BJ, Seo YR . Lack of genotoxic potential of ZnO nanoparticles in in vitro and in vivo tests. Mutat Res Genet Toxicol Environ Mutagen, 2014,761:1-9.
[9]	Krüger CT, Fischer BM, Armant O, Morath V, Strähle U, Hartwig A . The in vitro PIG-A gene mutation assay: glycosylphosphatidylinositol (GPI)-related genotype-to- phenotype relationship in TK6 cells. Arch Toxicol, 2016,90(7):1729-1736.
[10]	Rees BJ, Tate M, Lynch AM, Thornton CA, Jenkins GJ, Walmsley RM, Johnson GE . Development of an in vitro PIG-A gene mutation assay in human cells. Mutagenesis, 2017,32(2):283-297.
[11]	Magdolenova Z, Collins A, Kumar A, Dhawan A, Stone V, Dusinska M . Mechanisms of genotoxicity. A review of in vitro and in vivo studies with engineered nanoparticles. Nanotoxicology, 2014,8(3):233-278.
[12]	Sheng LP, Wang ZD, Zhou PK . The genetic toxicity and toxicology mechanism of metal nano materials. Chin J Prev Med, 2015, ( 9):831-834.
	沈丽萍, 王治东, 周平坤 . 金属纳米材料的遗传毒性及遗传毒理机制. 中华预防医学杂志, 2015, ( 9):831-834.
[13]	Du XM, Gao SX, Hong LL, Zheng X, Zhou QY, Wu JH . Genotoxicity evaluation of titanium dioxide nanoparticles using the mouse lymphoma assay and the Ames test. Mutat Res, 2019,838:22-27.
[14]	Franci G, Falanga A, Galdiero S, Palomba L, Rai M, Morelli G, Galdiero M . Silver nanoparticles as potential antibacterial agents. Molecules, 2015,20(5):8856-8874.
[15]	Lubick N . Nanosilver toxicity: ions, nanoparticles--or both?. Environ Sci Technol, 2008,42(23):8617.
[16]	Xin Q, Zhang Q, Cheng JP . Effects of silver nanoparticles and silver ions on the early development of zebrafish embryos and toxicity mechanisms. Asian J Ecotoxicol, 2015,10(4):55-64.
	辛琦, 章强, 程金平 . 纳米银和银离子对斑马鱼胚胎早期生长发育的影响及作用机制. 生态毒理学报, 2015,10(4):55-64.
[17]	Kawata K, Osawa M, Okabe S . In vitro toxicity of silver nanoparticles at noncytotoxic doses to HepG2 human hepatoma cells. Environ Sci Technol, 2009,43(15):6046-6051.
[18]	Hsiao IL, Hsieh YK, Wang CF, Chen IC, Huang YJ . Trojan-horse mechanism in the cellular uptake of silver nanoparticles verified by direct intra- and extracellular silver speciation analysis. Environ Sci Technol, 2015,49(6):3813-3821.
[19]	Liu W, Wu Y, Wang C, Li HC, Wang T, Liao CY, Cui L, Zhou QF, Yan B, Jiang GB . Impact of silver nanoparticles on human cells:effect of particle size. Nanotoxicology, 2010,4(3):319-330.
[20]	Tavares P, Balbinot F, de Oliveira HM, Fagundes GE, Venâncio M, Ronconi JVV, Merlini A, Streck EL, da Silva Paula MM, de Andrade VM. Evaluation of genotoxic effect of silver nanoparticles (Ag-Nps) in vitro and in vivo. J Nanopart Res, 2012,14(4):791-800.
[21]	Gong Y, Ji YJ, Liu F, Li J, Cao Y . Cytotoxicity, oxidative stress and inflammation induced by ZnO nanoparticles in endothelial cells: interaction with palmitate or lipopolysaccharide. J Appl Toxicol, 2017,37(8):895-901.
[22]	Dobrzyńska MM, Gajowik A, Radzikowska J, Lankoff A, Dušinská M, Kruszewski M . Genotoxicity of silver and titanium dioxide nanoparticles in bone marrow cells of rats in vivo. Toxicology, 2014,315(1):86-91.
[23]	Proquin H, Rodríguez-Ibarra C, Moonen CGJ, Ortega IMU, Briedé JJ, de Kok TM, van Loveren H, Chirino YI. Titanium dioxide food additive (E171) induces ROS formation and genotoxicity: contribution of micro and nano- sized fractions. Mutagenesis, 2017,32(1):139-149.
[24]	Wen HR, Shao AL, Chen L, Dan M, Hu YP, Wang X, Xu LM . Selection of a suitable method for the evaluation of genetic toxicity of nanomaterials. Carcinog Teratog Mutag, 2018,30(4):326-331.
	文海若, 邵安良, 陈亮, 淡墨, 胡燕平, 王雪, 徐丽明 . 适合纳米材料遗传毒性评价方法的选择. 癌变·畸变·突变, 2018,30(4):326-331.
[25]	Karlsson HL, Di Bucchianico S, Collins AR, Dusinska M . Can the comet assay be used reliably to detect nanoparticle- induced genotoxicity? Environ Mol Mutagen, 2015,56(2):82-96.

编辑推荐

Metrics

www.chinagene.cn
备案号：京ICP备09063187号-4
总访问:,今日访问:,当前在线:

纳米物质	纯度	粒径范围(nm)	比表面积(m²/g)	振实密度(g/cm³)	外观描述	受试浓度(μmol/L)
AgNPs	＞99.9%	60~80	≥3	≥2.5	黑色粉末	20、60、200
TiO₂NPs	99%	15~25	77.37	-	白色蓬松粉末	20、60、200

纳米银颗粒及纳米二氧化钛颗粒的体外遗传毒性研究

In vitro genotoxicity study of silver nanoparticles and titanium dioxide nanoparticles

RichHTML

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

图/表 6

参考文献 25

相关文章 2

编辑推荐

Metrics

[1]	屈艾，李宗芸，朱卫中，朱明明，王秀琴，温洪宇，汪承润. 稀土多元复合肥和三种稀土元素的遗传毒性研究[J]. 遗传, 2001, 23(3): 243-246.
[2]	马明福，李新生，蔡敏，李练兵，曾维三，廖明阳，王治乔. 基因重组人碱性成纤维细胞生长因子的遗传毒性研究[J]. 遗传, 2000, 22(1): 15-18.