Potential adverse outcome pathway of neurodevelopmental toxicity, inflammatory response, and oxidative stress induction mediated by three alkyl organophosphate flame retardants in zebrafish larvae.

Following restrictions on polybrominated flame retardants, trimethyl phosphate (TMP), triethyl phosphate (TEP), and tris(2-butoxyethyl) phosphate (TBEP) have been frequently used as plasticizers for fire-resistant plastics. This study investigated the neurodevelopmental effects, inflammatory response, and oxidative stress induction of three alkyl organophosphate flame retardants using a zebrafish embryo/larvae model. After exposure of zebrafish embryos to TMP, TEP, and TBEP (0, 0.02, 0.2, 2, 20, and 200 µg L-1) for 96 h, survival, development, swimming behavior, changes in acetylcholinesterase (AChE) activity, dopamine, tumor necrosis factor-alpha (TNF-α), interleukin (IL), reactive oxygen species (ROS), and antioxidant enzyme activities were observed. Concentrations of TMP, TEP, and TBEP were also measured in the whole body of exposed larvae. Our results showed that exposure to 200 µg L-1 TEP and ≥20 µg L-1 TBEP significantly reduced larval body length; however, TMP had no significant effects on developmental parameters up to 200 µg L-1. After 96 h of exposure to TBEP, total distance moved, mean velocity, AChE, and dopamine concentrations were significantly decreased. Exposure to TEP and TBEP decreased the expression of genes that regulate central nervous system development (e.g. gap43 and mbpa), whereas ROS, antioxidant enzymes, TNF-α, and IL-1ß concentrations were significantly increased. Notably, pretreatment with an antioxidant N-acetylcysteine reduced neurotoxicity and oxidative stress caused by TEP and TBEP. The results of this study demonstrated that exposure to TEP and TBEP causes oxidative stress and has adverse effects on the neurobehavioral and immune system of zebrafish, leading to hypoactivity and ultimately impairing development.

Effects of two alternative plasticizers on the growth hormone-related endocrine system, neurodevelopment, and oxidative stress of zebrafish larvae.

In response to the restriction of phthalate plasticizers, acetyl tributyl citrate (ATBC) and acetyl triethyl citrate (ATEC) have been used in medical devices and food packaging. In the present study, the effects of ATBC and ATEC on the development, behavior, growth hormone (GH)-related endocrine system, neurotransmitters, and oxidative stress of zebrafish embryo or larvae were investigated. After exposure of zebrafish to ATBC and ATEC (0, 0.03, 0.3, 3, 30, and 300 µg/L) for 96 h, developmental toxicity, behavioral changes under light/dark condition, changes in hormones and genes involved in GH/insulin-like growth factors (IGFs) axis, changes in hormone, enzyme, and genes related to neurodevelopment, antioxidant enzymes activities were determined. Larvae exposed to 30 or 300 µg/L ATBC showed significant reductions in body length and moving distance and speed, whereas no significant effects on development and locomotor behavior were observed in larvae exposed to ATEC. The contents of GH and IGF-I were significantly reduced in larvae exposed to 3, 30, and 300 µg/L ATBC. Hormonal changes in fish exposed to ATBC are well supported by regulation of genes related to GH (gh1) and the activity of IGF-I (igf1). In fish exposed to ATBC, reduced acetylcholinesterase activity and down-regulation of genes related to the central nervous system development (ache, gap43, mbpa, and syn21) were observed. ATBC increased the production of reactive oxygen species and the levels of superoxide dismutase, catalase, and glutathione peroxidase. Notably, pre-treatment with the classic antioxidant N-acetylcysteine alleviated ATBC-induced GH-related endocrine disruption and neurotoxicity. Our observations showed that exposure to low levels of ATBC could disturb the regulatory systems of GH/IGFs axis and neurobehavior, ultimately leading to developmental inhibition and hypoactivity, and that increased oxidative stress plays a major role in these toxicities.

Effects of di-(2-ethylhexyl) terephthalate on hypothalamus-pituitary-gonad axis in adult zebrafish.

Di-(2-ethylhexyl) terephthalate (DEHTP) is frequently used in food packaging and medical devices as an alternative to di-(2-ethylhexyl) phthalate (DEHP). In this study, zebrafish pairs were exposed to DEHTP for 21 d and the effects on fertility, sex hormone levels, vitellogenin levels, and transcription of genes along the hypothalamic-pituitary-gonad axis were evaluated. Results showed that mean egg numbers were significantly reduced in the 30 and 300 µg/L DEHTP groups. The adverse effects of DEHTP on hormones and gene transcripts were more prominent in males than in females. In male fish, the gonadosomatic index, hepatosomatic index, and vitellogenin concentration were significantly increased. The results of a significant decrease in testosterone (T) and an increase in the 17ß-estradiol (E2)/T ratio in males exposed to 3-300 µg/L DEHTP suggest that the endocrine potential of DEHTP is similar that of DEHP. In females, genes related to gonadotropin-releasing hormone and gonadotropin were up-regulated while E2 was significantly down-regulated. These findings suggest that positive E2 feedback mechanisms in the hypothalamus and pituitary gland are activated to balance sex hormones. The effects of chronic exposure to DEHTP on the neuroendocrine system require further investigation.