The possible hormetic effects of fluorene-9-bisphenol on regulating hypothalamic-pituitary-thyroid axis in zebrafish

Highlights

• BHPF causes developmental toxicity and injured locomotion in a dose-dependent manner.

• The possible hormetic effects of BHPF on regulating HPT axis in zebrafish.

• BHPF disrupts myelination through affecting HPT axis, causing neurotoxicity.

Abstract

Fluorene-9-bisphenol (BHPF) is a bisphenol A substitute, which has been introduced for the production of so-called ‘bisphenol A (BPA)-free’ plastics. However, it has been reported that BHPF can enter living organisms through using commercial plastic bottles and cause adverse effects. To date, the majority of the toxicologic study of BHPF focused on investigating its doses above the toxicological threshold. Here, we studied the effects of BHPF on development, locomotion, neuron differentiation of the central nervous system (CNS), and the expression of genes in the hypothalamic-pituitary-thyroid (HPT) axis in zebrafish exposed to different doses of BHPF ranging from 1/5 of LD1 to LD50 (300, 500, 750, 1500, 3000, and 4500 nM). As a result, the possible hormetic effects of BHPF on regulating the HPT axis were revealed, in which low-dose BHPF positively affected the HPT axis while this regulation was inhibited as the dose increased. Underlying mechanism investigation suggested that BHPF disrupted myelination through affecting HPT axis including related genes expression and TH levels, thus causing neurotoxic characteristics. Collectively, this study provides the full understanding of the environmental impact of BHPF and its toxicity on living organisms, highlighting a substantial and generalized ongoing dose-response relationship with great implications for the usage and risk assessment of BHPF.

Introduction

As one of bisphenol A (BPA) substitutes, Fluorene-9-bisphenol (BHPF) has been widely used in plastic production (so-called ‘BPA-free’ plastics) because of its excellent thermal stability (den Braver-Sewradj et al., 2020). The kind and number of products containing BHPF is increasingly growing, including structural adhesives, insulation materials, protective coatings, and epoxy floor coatings, in the daily life and industries. However, like BPA, the adverse effects of BHPF are becoming a pressing issue and attracting a broadly attention. Previous findings reported that BHPF has anti-oestrogenic activity, causing harmful pregnancy outcomes in mice via affecting the homeostasis in the endocrine system (Zhang et al., 2017b). In this study, the mean level of BHPF in the water from plastic bottles is 19.51 ± 29.55 ng/L while the maximal concentration is 81.47 ng/L. BHPF is found in 7 out of 100 serum samples of human volunteers, with the average concentration of 0.34 ± 0.21 ng/mL. These results revealed the presence of BHPF in commercial plastic bottles and BHPF can enter living organisms. In another study, BHPF induces cytotoxicity in mice and porcine oocytes and causes ovarian damage (Jia et al., 2019; Jiao et al., 2020). In addition to the toxic studies in rodents, it has been reported that acute BHPF treatment in zebrafish damages courtship and sleep/wake behavior, as well as induce anxiety/depression-like behavior (Mi et al., 2019a; Mi et al., 2019b). Acute and chronic exposure alters the development and lipid accumulation in larval and adult zebrafish (Sun et al., 2020). In addition, zebrafish larvae treated with 1000–10,000 nM of BHPF shows developmental toxicity and cardiotoxicity. However, whether BHPF affecting hypothalamic-pituitary-thyroid (HPT) axis remains largely unknown. Importantly, limited data is available on the effects of low-dose of BHPF on organisms.

As a multicomponent neuroendocrine system, the HPT axis is responsible for maintaining homeostasis of the thyroid hormone (TH), which plays pivotal roles in teleost fish development, including brain and body development, yolk absorption, and regulation of cardiovascular, bone, liver, and nerve function (Admati et al., 2020; Blanton and Specker, 2007; Nelson and Habibi, 2009; Tomkins et al., 2018). In recent years, newly emerging compounds show the ability to interrupt the HPT axis. For example, transcription analysis of rat brains showed that BPA alters the expression of genes in the HPT (Zoeller et al., 2005). Moreover, TH is a typical example of biphasic dose-response under the medical conditions, which shows both hypothyroidism and hyperthyroidism (Donzelli et al., 2016). Thyroxine (T4) is the main TH secreted by the thyroid follicles of teleost fish, while 3,5,30-triiodothyronine (T3) is the biologically active form that is under the control of peripheral tissues, which is regulated by the iodothyronine deiodinases (Dio) (Power et al., 2001). It has been reported that TH disrupting chemicals including pesticides, PCBs, PBDEs, perchlorate, phthalates, and PBA are able to increase or decrease T4 and T3 levels in zebrafish (Chen et al., 2012b; Huang et al., 2016; Liang et al., 2015; Liu et al., 2011; Tang et al., 2015; Tu et al., 2016; Wang et al., 2013). Thus, it is worth to investigate the effects of BHPF on regulating the HPT axis. In addition, as a key factor responsible for regulating systemic thyroid hormone levels (Fekete and Lechan, 2014; Fliers et al., 2014; Zoeller et al., 2007), HPT axis is involved in variety of diseases including lipid and glucose metabolism disorder, obesity, and the CNS–related disorders (Calza et al., 2010; Fernandez et al., 2004; Klieverik et al., 2009; Krotkiewski, 2002; Muller et al., 1990; Sanyal and Raychaudhuri, 2016; Zhang et al., 2016).

Hormesis is a phenomenon characterized by biphasic dose-response transformation, which is for low-dose environmental stress-inducing factors. It shows a stimulation at low-dose while an inhibition at high-dose (Calabrese, 2008; Stebbing, 2002; Stebbing, 2009). The hormesis in aquatic and terrestrial organisms subjected to various environmental stressor, including global pollutants and contaminants, is observed in a wide range of endpoints like carcinogenicity, life span, growth, cell and tissue repair, and behavior (Arumugam et al., 2006; Calabrese and Mattson, 2017; Ng et al., 2016). Nonetheless, the relationship between the hormetic response and the regulation of HPT axis needs further explored.

In this study, the hormetic effects on BHPF-induced neurotoxicity were studied by using zebrafish as a model. Zebrafish were exposed to BHPF at different doses, ranging from 1/5 of LD1 to LD50 (300, 500, 750, 1500, 3000, and 4500 nM). This dose/response phenomenon represents an evolutionarily conserved process of adaptive, providing a novel perspective to understand organismic stress and how BHPF affect organisms at doses below the toxicological threshold.