基于高内涵分析技术表征磷酸三苯酯的肺细胞毒性

丰一兴; 王济洲; 段鹤君; 李伟红; 邵兵

doi:10.7524/AJE.1673-5897.20220914001

生态毒理学报

, , , ,

丰一兴, 王济洲, 段鹤君, 李伟红, 邵兵. 基于高内涵分析技术表征磷酸三苯酯的肺细胞毒性[J]. 生态毒理学报, 2023, 18(2): 259-268. doi: 10.7524/AJE.1673-5897.20220914001

引用本文:

Feng Yixing, Wang Jizhou, Duan Hejun, Li Weihong, Shao Bing. Characterizing Cytotoxicity of Lung Cells Induced by Triphenyl Phosphate Based on High Content Analysis[J]. Asian journal of ecotoxicology, 2023, 18(2): 259-268. doi: 10.7524/AJE.1673-5897.20220914001

Citation:

Feng Yixing, Wang Jizhou, Duan Hejun, Li Weihong, Shao Bing. Characterizing Cytotoxicity of Lung Cells Induced by Triphenyl Phosphate Based on High Content Analysis[J]. Asian journal of ecotoxicology, 2023, 18(2): 259-268. doi: 10.7524/AJE.1673-5897.20220914001

基于高内涵分析技术表征磷酸三苯酯的肺细胞毒性

1. 北京市疾病预防控制中心食物中毒诊断溯源技术北京市重点实验室, 北京 100013;
2. 中国科学院大学, 北京 100049

作者简介: 丰一兴(1979—)，女，副研究员，研究方向为生态毒理学，E-mail: fengyxioz@126.com

通讯作者: 邵兵, E-mail: shaobingch@sina.com

中图分类号: X171.5

Characterizing Cytotoxicity of Lung Cells Induced by Triphenyl Phosphate Based on High Content Analysis

1. Beijing Center for Disease Control and Prevention, Beijing Key Laboratory of Diagnostic and Traceability Technologies for Food Poisoning, Beijing 100013, China;
2. University of Chinese Academy of Sciences, Beijing 100049, China

Corresponding author: Shao Bing, shaobingch@sina.com

摘要: 磷酸三苯酯(triphenyl phosphate, TPhP)是环境中最常见的有机磷酸酯阻燃剂之一，具有较强的挥发性，每天持续摄入TPhP，可能对人体肺部组织产生不利影响。本研究以人源的非小细胞肺癌细胞系(A549)为研究对象，采用高内涵分析系统检测50、100和200 μmol·L^-1 TPhP对细胞核形态、核膜通透性、线粒体纹理结构、线粒体膜电位、细胞内氧自由基水平、磷酸化组蛋白H2AX(phosphorylated histones H2AX, pH2AX)、细胞色素C和细胞凋亡等参数的影响。结果显示，与对照组相比，随着TPhP暴露浓度的增加，A549细胞活性显著下降，细胞核面积增加、核亮度降低和核碎片化程度加剧，核膜通透性增高，细胞核的各参数在200 μmol·L^-1 TPhP组变化最显著；细胞内氧自由基水平的显著升高，进一步导致线粒体损伤和细胞色素C释放，当TPhP为100 μmol·L^-1时，线粒体损伤程度最严重；当TPhP为100 μmol·L^-1和200 μmol·L^-1时，pH2AX含量显著升高，表明DNA受到损伤；100 μmol·L^-1 TPhP主要诱导细胞发生早期凋亡，而200 μmol·L^-1 TPhP诱导细胞发生早期凋亡和晚期凋亡。TPhP对人肺细胞呈现明显的细胞毒性，可引起氧化应激介导的DNA损伤和线粒体损伤，并最终导致人肺细胞发生凋亡。本研究结果为科学评估TPhP对人体健康的危害提供了数据支持和理论依据。
- 磷酸三苯酯 /
- 人肺细胞 /
- 细胞毒性表型 /
- 高内涵筛选分析
Abstract: Triphenyl phosphate (TPhP) is one of the most commonly detected organophosphorus flame retardants (OPFRs) in the environment. Continuous daily exposure to TPhP may adversely impact human lung tissues due to its high volatility. In this study, human lung carcinoma cell line (A549) was selected as the cell model and the effects of 50, 100, and 200 μmol·L^-1 TPhP on nuclear morphology, nuclear membrane permeability, mitochondrial texture structure, mitochondrial membrane potential, intracellular reactive oxygen species, phosphorylated histones H2AX (pH2AX), cytochrome C and cell apoptosis were detected using the high-content screening (HCS) system. As the results shown, with the increased exposure concentration of TPhP, a decrease in cell viability, an increase in the nuclear area, a decrease in the nuclear intensity, an elevated degree of nuclear fragmentation, and an increase in the nuclear membrane permeability were observed. The changes in the nuclear multiple parameters were most significant in the 200 μmol·L^-1 TPhP group. Moreover, with the increase of intracellular reactive oxygen species levels, the damage of mitochondria was aggravated and cytochrome C was released. At the dosage of 100 μmol·L^-1 TPhP, the degree of mitochondrial damage was the most serious. In addition, the content of pH2AX was significantly increased at the dosage of 100 μmol·L^-1 and 200 μmol·L^-1 TPhP, which indicated the presence of DNA damage. Finally, early apoptosis was induced by 100 μmol·L^-1 TPhP treatment, while early and late apoptosis was induced after exposure to 100 μmol·L^-1 and 200 μmol·L^-1 TPhP. In conclusion, TPhP showed obvious cytotoxic effects on lung cells and caused oxidative stress-mediated DNA damage and mitochondrial impairment, thereby leading to the cell apoptosis of lung cells. The completion of this study provides theoretical basis and data supports for the scientific evaluation of human health risk induced by TPhP.
- triphenyl phosphate /
- human lung cells /
- cytotoxic phenotype /
- high content screening analysis

Yang J W, Zhao Y Y, Li M H, et al. A review of a class of emerging contaminants: The classification, distribution, intensity of consumption, synthesis routes, environmental effects and expectation of pollution abatement to organophosphate flame retardants (OPFRs)[J]. International Journal of Molecular Sciences, 2019, 20(12): 2874

Du J, Li H X, Xu S D, et al. A review of organophosphorus flame retardants (OPFRs): Occurrence, bioaccumulation, toxicity, and organism exposure[J]. Environmental Science and Pollution Research International, 2019, 26(22): 22126-22136

Wei G L, Li D Q, Zhuo M N, et al. Organophosphorus flame retardants and plasticizers: Sources, occurrence, toxicity and human exposure[J]. Environmental Pollution, 2015, 196: 29-46

Feng L P, Ouyang F X, Liu L P, et al. Levels of urinary metabolites of organophosphate flame retardants, TDCIPP, and TPHP, in pregnant women in Shanghai[J]. Journal of Environmental and Public Health, 2016, 2016: 9416054

He R W, Li Y Z, Xiang P, et al. Organophosphorus flame retardants and phthalate esters in indoor dust from different microenvironments: Bioaccessibility and risk assessment[J]. Chemosphere, 2016, 150: 528-535

Chen Y X, Liu Q Y, Ma J, et al. A review on organophosphate flame retardants in indoor dust from China: Implications for human exposure[J]. Chemosphere, 2020, 260: 127633

Li H R, La Guardia M J, Liu H H, et al. Brominated and organophosphate flame retardants along a sediment transect encompassing the Guiyu, China e-waste recycling zone[J]. The Science of the Total Environment, 2019, 646: 58-67

Wang R M, Tang J H, Xie Z Y, et al. Occurrence and spatial distribution of organophosphate ester flame retardants and plasticizers in 40 rivers draining into the Bohai Sea, North China[J]. Environmental Pollution, 2015, 198: 172-178

He C, Toms L L, Thai P, et al. Urinary metabolites of organophosphate esters: Concentrations and age trends in Australian children[J]. Environment International, 2018, 111: 124-130

Gao D T, Yang J, Bekele T G, et al. Organophosphate esters in human serum in Bohai Bay, North China[J]. Environmental Science and Pollution Research International, 2020, 27(3): 2721-2729

Chen X L, Zhao X Z, Shi Z X. Organophosphorus flame retardants in breast milk from Beijing, China: Occurrence, nursing infant's exposure and risk assessment[J]. Science of the Total Environment, 2021, 771: 145404

An J, Hu J W, Shang Y, et al. The cytotoxicity of organophosphate flame retardants on HepG2, A549 and Caco-2 cells[J]. Journal of Environmental Science and Health Part A, Toxic/Hazardous Substances & Environmental Engineering, 2016, 51(11): 980-988

Canbaz D, Logiantara A, van Ree R, et al. Immunotoxicity of organophosphate flame retardants TPHP and TDCIPP on murine dendritic cells in vitro[J]. Chemosphere, 2017, 177: 56-64

Belcher S M, Cookman C J, Patisaul H B, et al. In vitro assessment of human nuclear hormone receptor activity and cytotoxicity of the flame retardant mixture FM 550 and its triarylphosphate and brominated components[J]. Toxicology Letters, 2014, 228(2): 93-102

Wang X Q, Li F, Liu J L, et al. New insights into the mechanism of hepatocyte apoptosis induced by typical organophosphate ester: An integrated in vitro and in silico approach[J]. Ecotoxicology and Environmental Safety, 2021, 219: 112342

Bonner W M, Redon C E, Dickey J S, et al. GammaH2AX and cancer[J]. Nature Reviews Cancer, 2008, 8(12): 957-967

Eisner V, Picard M, Hajnóczky G. Mitochondrial dynamics in adaptive and maladaptive cellular stress responses[J]. Nature Cell Biology, 2018, 20(7): 755-765

Le Y F, Shen H P, Yang Z, et al. Comprehensive analysis of organophosphorus flame retardant-induced mitochondrial abnormalities: Potential role in lipid accumulation[J]. Environmental Pollution, 2021, 274: 116541

Perdiz D, Oziol L, Poüs C. Early mitochondrial fragmentation is a potential in vitro biomarker of environmental stress[J]. Chemosphere, 2019, 223: 577-587

Li R W, Zhou P J, Guo Y Y, et al. Tris (1,3-dichloro-2-propyl) phosphate induces apoptosis and autophagy in SH-SY₅Y cells: Involvement of ROS-mediated AMPK/mTOR/ULK1 pathways[J]. Food and Chemical Toxicology, 2017, 100: 183-196

Yu X L, Yin H, Peng H, et al. OPFRs and BFRs induced A549 cell apoptosis by caspase-dependent mitochondrial pathway[J]. Chemosphere, 2019, 221: 693-702

Kluck R M, Esposti M D, Perkins G, et al. The pro-apoptotic proteins, Bid and Bax, cause a limited permeabilization of the mitochondrial outer membrane that is enhanced by cytosol[J]. The Journal of Cell Biology, 1999, 147(4): 809-822

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Citation:

基于高内涵分析技术表征磷酸三苯酯的肺细胞毒性

通讯作者: 邵兵, E-mail: shaobingch@sina.com

作者简介: 丰一兴(1979—)，女，副研究员，研究方向为生态毒理学，E-mail: fengyxioz@126.com
1. 北京市疾病预防控制中心食物中毒诊断溯源技术北京市重点实验室, 北京 100013;

2. 中国科学院大学, 北京 100049

收稿日期: 2022-09-14

关键词:

摘要: 磷酸三苯酯(triphenyl phosphate, TPhP)是环境中最常见的有机磷酸酯阻燃剂之一，具有较强的挥发性，每天持续摄入TPhP，可能对人体肺部组织产生不利影响。本研究以人源的非小细胞肺癌细胞系(A549)为研究对象，采用高内涵分析系统检测50、100和200 μmol·L^-1 TPhP对细胞核形态、核膜通透性、线粒体纹理结构、线粒体膜电位、细胞内氧自由基水平、磷酸化组蛋白H2AX(phosphorylated histones H2AX, pH2AX)、细胞色素C和细胞凋亡等参数的影响。结果显示，与对照组相比，随着TPhP暴露浓度的增加，A549细胞活性显著下降，细胞核面积增加、核亮度降低和核碎片化程度加剧，核膜通透性增高，细胞核的各参数在200 μmol·L^-1 TPhP组变化最显著；细胞内氧自由基水平的显著升高，进一步导致线粒体损伤和细胞色素C释放，当TPhP为100 μmol·L^-1时，线粒体损伤程度最严重；当TPhP为100 μmol·L^-1和200 μmol·L^-1时，pH2AX含量显著升高，表明DNA受到损伤；100 μmol·L^-1 TPhP主要诱导细胞发生早期凋亡，而200 μmol·L^-1 TPhP诱导细胞发生早期凋亡和晚期凋亡。TPhP对人肺细胞呈现明显的细胞毒性，可引起氧化应激介导的DNA损伤和线粒体损伤，并最终导致人肺细胞发生凋亡。本研究结果为科学评估TPhP对人体健康的危害提供了数据支持和理论依据。

English Abstract

Characterizing Cytotoxicity of Lung Cells Induced by Triphenyl Phosphate Based on High Content Analysis

Corresponding author: Shao Bing, shaobingch@sina.com

1. Beijing Center for Disease Control and Prevention, Beijing Key Laboratory of Diagnostic and Traceability Technologies for Food Poisoning, Beijing 100013, China;
2. University of Chinese Academy of Sciences, Beijing 100049, China

Received Date: 2022-09-14

Keywords:

Abstract: Triphenyl phosphate (TPhP) is one of the most commonly detected organophosphorus flame retardants (OPFRs) in the environment. Continuous daily exposure to TPhP may adversely impact human lung tissues due to its high volatility. In this study, human lung carcinoma cell line (A549) was selected as the cell model and the effects of 50, 100, and 200 μmol·L^-1 TPhP on nuclear morphology, nuclear membrane permeability, mitochondrial texture structure, mitochondrial membrane potential, intracellular reactive oxygen species, phosphorylated histones H2AX (pH2AX), cytochrome C and cell apoptosis were detected using the high-content screening (HCS) system. As the results shown, with the increased exposure concentration of TPhP, a decrease in cell viability, an increase in the nuclear area, a decrease in the nuclear intensity, an elevated degree of nuclear fragmentation, and an increase in the nuclear membrane permeability were observed. The changes in the nuclear multiple parameters were most significant in the 200 μmol·L^-1 TPhP group. Moreover, with the increase of intracellular reactive oxygen species levels, the damage of mitochondria was aggravated and cytochrome C was released. At the dosage of 100 μmol·L^-1 TPhP, the degree of mitochondrial damage was the most serious. In addition, the content of pH2AX was significantly increased at the dosage of 100 μmol·L^-1 and 200 μmol·L^-1 TPhP, which indicated the presence of DNA damage. Finally, early apoptosis was induced by 100 μmol·L^-1 TPhP treatment, while early and late apoptosis was induced after exposure to 100 μmol·L^-1 and 200 μmol·L^-1 TPhP. In conclusion, TPhP showed obvious cytotoxic effects on lung cells and caused oxidative stress-mediated DNA damage and mitochondrial impairment, thereby leading to the cell apoptosis of lung cells. The completion of this study provides theoretical basis and data supports for the scientific evaluation of human health risk induced by TPhP.

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基于高内涵分析技术表征磷酸三苯酯的肺细胞毒性

作者简介: 丰一兴(1979—)，女，副研究员，研究方向为生态毒理学，E-mail: fengyxioz@126.com

通讯作者: 邵兵, E-mail: shaobingch@sina.com

Characterizing Cytotoxicity of Lung Cells Induced by Triphenyl Phosphate Based on High Content Analysis

Corresponding author: Shao Bing, shaobingch@sina.com

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基于高内涵分析技术表征磷酸三苯酯的肺细胞毒性

通讯作者: 邵兵, E-mail: shaobingch@sina.com

作者简介: 丰一兴(1979—)，女，副研究员，研究方向为生态毒理学，E-mail: fengyxioz@126.com 1. 北京市疾病预防控制中心食物中毒诊断溯源技术北京市重点实验室, 北京 100013; 2. 中国科学院大学, 北京 100049

English Abstract

Characterizing Cytotoxicity of Lung Cells Induced by Triphenyl Phosphate Based on High Content Analysis

Corresponding author: Shao Bing, shaobingch@sina.com

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作者简介: 丰一兴(1979—)，女，副研究员，研究方向为生态毒理学，E-mail: fengyxioz@126.com
1. 北京市疾病预防控制中心食物中毒诊断溯源技术北京市重点实验室, 北京 100013;

2. 中国科学院大学, 北京 100049