Effects of environmentally relevant concentrations of tris (2-chloroethyl) phosphate (TCEP) on early life stages of zebrafish (Danio rerio)

https://doi.org/10.1016/j.etap.2021.103600Get rights and content

Highlights

  • TCEP (20, 200 μg L−1) could lead to developmental toxicity, including decreased body length and delay of hatching.

  • TCEP exposure decreased whole-body T4 levels and altered transcriptional profiles of genes involved in HPT axis.

  • TCEP significantly affected transcriptional levels of key genes related to neurodevelopment.

Abstract

Tris (2-chloroethyl) phosphate (TCEP) has been received great concerns because of its increasing presence in various environmental compartments and toxicity. In the present study, zebrafish embryos were exposed to environmentally relevant concentrations of TCEP (0.2, 2, 20, 200 μg/L) from 3 to 120 h post-fertilization (hpf). The results showed that TCEP exposure (20, 200 μg/L) led to developmental toxicity including decreased body length and delay of hatching. Treatment with TCEP significantly decreased whole-body thyroxine (T4) levels and mRNA level of thyroglobulin (tg), and enhanced transcriptions of genes sodium/iodide symporter (nis), thyroid hormone receptor α (trα) and ugt1ab involved in thyroid synthesis and metabolism, respectively. Additionally, TCEP altered the transcription of α1-tubulin, gap43 and mbp related to nervous system development, even at relatively low concentrations. Overall, our results revealed that TCEP exposure can lead to developmental toxicity, thyroid endocrine disruption and neurotoxicity on early developmental stages of zebrafish.

Introduction

After the prohibition of using polybrominated diphenyl ethers (PBDEs), organophosphorus flame retardants (OPFRs) consumption increased greatly in the past few years. As one of the important OPFRs, Tris (2-chloroethyl) phosphate (TCEP) has been widely used in plastics, building materials, electronic devices, furniture, baby toys and other products. Given that TCEP is not covalently bound to the polymer matrices, it tends to be released into the environment through many pathways, such as volatilization, leaching and abrasion (Bollmann et al., 2012). According to recent reports, high concentrations of TCEP have been detected in various biological and environmental samples including surface water, atmosphere, street dust, soil, sediment, bird plasma and human serum (Kurtkarakus et al., 2017; Li et al., 2017; Hao et al., 2018; He et al., 2018; Guo et al., 2018; Zhong et al., 2018). Because of its high water solubility (7000 mg/L) (Hou et al., 2016; Van der Veen and de Boer, 2012), TCEP exists extensively in water. The highest concentration of the TCEP ever reported was up to 87.4 μg/L in raw water sampled near solid waste disposal site in Japanese (Kawagoshi et al., 1999). In Sweden, the concentrations of TCEP detected in sludge ranged from 6.6–110 μg/kg (Marklund et al., 2005).

Nowadays, the potential risk of OPFRs on environmental health has attracted a lot of attention due to their persistence, wide distribution and similar chemical structure with organophosphorus insecticides. In recent years, several literatures have shown that OPFRs exhibited embryotoxicity, reproduction toxicity, immunotoxicity, thyroid disruption, neurotoxicity and even carcinogenicity (Kim et al., 2015; Sun et al., 2016a, b; Canbaz et al., 2017; Li et al., 2019). For example, tris (2-chloropropyl) phosphate (TCPP) could impair the normal embryonic development process, resulting in deformed zebrafish larvae (Xia et al., 2020). After exposure to Triphenyl phosphate (TPP), both T3 and T4 levels in zebrafish larvae increased significantly and expression levels of the genes associated with thyroid hormone synthesis were changed (Kim et al., 2015). Given the important role thyroid hormones play in early developmental stages of zebrafish, modification of thyroid hormone homeostasis may lead to serious toxicological effects (Jarque and Piña, 2014; Spaan et al., 2019). However, to our knowledge, information about the endocrine disrupting effects aroused by TCEP and the underlying mechanism is poorly understood. Furthermore, it has been proved that TPP and TCEP can affect the transcriptional levels of several genes involved in nervous system development of zebrafish larvae (Peng et al., 2016; Li et al., 2019). Overall, these findings suggest that TCEP may have adverse effects on embryonic development, potential thyroid hormone toxicity and neurotoxicity. However, current knowledge of the toxicological effects of TCEP on aquatic organisms are largely limited. Moreover, exposure concentrations adopted in the previous works were much higher than environmental realistic concentration.

Zebrafish has become a valuable vertebrate model organism for chemical toxicity assessment duo to its multiple advantages including easy maintenance, short life cycle, high fecundity, rapid development and sensitivity to the environmental toxins (Dai et al., 2014). In the present work, zebrafish embryo and larvae were used as models to investigate the influences of TCEP on embryonic development, thyroid hormone disruption and neurodevelopment in its early life stages. Multiple indicators including developmental parameters (hatching rate, heartbeat, malformation rate and body length), thyroid hormones levels, and the transcriptional changes of several representative genes involved in thyroid hormone homeostasis and neurodevelopment were analyzed, which aimed to provide insights into the underlying mechanisms and enhance our awareness of the potential risk of TCEP for aquatic organisms.

Section snippets

Chemicals

Tris (2-chlorethyl) phosphate (TCEP, CAS: 115-96-8; >97% purity) was purchased from Sigma-Aldrich Chemical Co. (St. Louis, USA). TCEP stock solution were prepared in dimethyl sulphoxide (DMSO; >99.9% purity;Sigma-Aldrich, USA) to form a series of stock solutions. The final concentration of DMSO was 0.01% (v/v) in the exposure media. All other reagents used in this study were of analytical grade.

Experimental design

Adult zebrafish (TU strain) were acclimated in 40 L glass aquariums containing 30 L dechlorinated and

Developmental toxicity

A significant drop of heartbeat rate was only observed in 200 μg/L TCEP treated group at 48 hpf, while TCEP showed no significant effects on heartbeat rate in other exposure groups at 48 hpf and all treatment groups at 72 hpf and 96 hpf compared with the control (Fig. 1A). Body length declined in 20 and 200 μg/L TCEP treated groups in comparison to the control at 72 hpf and 120 hpf (Fig. 1B). The hatching rates were remarkably inhibited in 20 and 200 μg/L TCEP treated groups at 72hpf, but not

Discussion

Early-life stage toxicity test has been frequently adopted to evaluate adverse effects of environmental contaminants on fish, since they are extremely sensitive to chemical exposure during this stage. Several lethal and nonlethal endpoints such as survival, heartbeat number, hatching rate, malformation and body length were commonly used for assessing developmental toxicity. Here, malformations including pericardial edema, yolk sac edema, notochord distortion and tail deformation were observed

Conclusion

In summary, the present work indicated that exposure to TCEP (above 20 μg/L) can lead to developmental toxicity including decreased body length and delay of hatching.

Treatment with TCEP can result in thyroid hormone disruption by decreasing T4 levels and changing transcriptions of multiple genes related to thyroid hormone homeostasis. The present study also demonstrated that TCEP altered the expression of crucial genes (α1-tubulin, syn2a, gap43, elavl3, mbp) related to central nervous system

CRediT authorship contribution statement

Fengxiao Hu: Conceptualization, Formal analysis, Methodology, Writing - review & editing. Yixin Zhao: Conceptualization, Investigation, Methodology, Visualization, Writing - original draft. Yuan Yuan: Formal analysis, Investigation, Visualization. Li Yin: Investigation. Feilong Dong: Investigation. Weini Zhang: Resources. Xinhua Chen: Supervision.

Declaration of Competing Interest

We declare that we do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted.

Acknowledgements

This work was supported by National Natural Science Foundation of China (No. 31702394) and Natural Science Foundation of Fujian Province (2016J05062).

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    These authors have Zhao contributed equally to this work.

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