Hepatic Transcriptomic Responses to Ethinylestradiol in Two Life Stages of Japanese Quail

Abstract

Chemical risk assessment for avian species typically depends on information from toxicity tests performed in adult birds. Early-life stage (ELS) toxicity tests have been proposed as an alternative, but incorporation of these data into existing frameworks will require knowledge about the similarities/differences between ELS and adult responses. The present study uses transcriptomics to assess hepatic gene expression in ELS and adult Japanese quail following exposure to ethinylestradiol (EE2). Prior to incubation, ELS quail were dosed with measured EE2 concentrations of 0.54, 6.3, and 54.2 µg/g egg weight via air cell injection. Adult quail were fed a single dose of EE2 at nominal concentrations of 0, 0.5, and 5 mg/kg body weight by gavage. Liver tissue was collected from five to six individuals per dose group at mid-incubation for ELS quail and 4 days after dosing for adults. A total of 283 and 111 differentially expressed genes (DEGs) were detected in ELS and adult quail, respectively, 16 of which were shared across life stages. Shared DEGs included estrogenic biomarkers such as vitellogenin genes and apovitellenin-1. For the dose groups that resulted in the highest number of DEGs (ELS, 6.3 µg/g; adult, 5 mg/kg), 21 and 35 Kyoto Encyclopedia of Genes and Genomes pathways were enriched, respectively. Ten of these pathways were shared between life stages, including pathways involved with signaling molecules and interaction and the endocrine system. Taken together, our results suggest conserved mechanisms of action following estrogenic exposure across two life stages, with evidence from differential expression of key biomarker genes and enriched pathways. The present study contributes to the development and evaluation of ELS tests and toxicogenomic approaches and highlights their combined potential for screening estrogenic chemicals.

Hepatic Transcriptomic Responses to Ethinylestradiol in Embryonic Japanese Quail and Double‐Crested Cormorant

“…Even though sex was not determined in the avian early life stage toxicity test protocol used in our study (Farhat et al, 2020), a strong transcriptomic signal of estrogenic exposure was robustly detected in both species in the present study and in both embryonic (sex unknown) and adult (known sex) JQ in a previous study (Jeon et al, 2022). However, sexually dimorphic responses to contaminants are well described in the literature (Lee et al, 2015; Martyniuk et al, 2013; Mittal et al, 2021).…”

“…This species difference in transcriptomic response was consistent with the previously reported sensitivity differences at the organismal level (Crump et al, 2021). A consistent positive relationship between the number of DEGs and the number or severity of organismal effects has not been firmly established in the literature; however, a similar pattern across biological scales was observed in a life stage comparison study of embryonic and adult JQ exposed to EE2 (Jeon et al, 2022) and a multi‐omics study in embryonic JQ and DCCO following chlorpyrifos exposure (Desforges et al, 2021). Increased DEGs at higher concentrations with greater corresponding key event responses were also observed in fathead minnow exposed to different doses of EE2 (Alcaraz et al, 2021).…”

“…The up‐regulation of vitellogenin and apovitellenin genes in liver has long been associated with estrogenic exposure (Denslow et al, 1999; Elbrecht et al, 1984; Sharin et al, 2021). Vitellogenin and apovitellenin were robustly dysregulated in embryonic chickens (Elbrecht et al, 1984; Li et al, 2014) and JQ at different life stages (Hanafy et al, 2006; Ichikawa et al, 2003; Jeon et al, 2022) following exposure to estrogenic chemicals. Estrogen exposure also dysregulates genes involved in the hepatic production of other yolk proteins such as riboflavin‐binding protein gene (Clagett et al, 1970; Jeon et al, 2022).…”

“…Because ZP genes are used as estrogen exposure biomarkers in fish (Gunnarsson et al, 2007), they might be equally useful in birds. Indeed, ZP1 expression increased in JQ after intraperitoneal, maternal, dietary, or embryonic exposure to an estrogenic compound (Hanafy et al, 2007; Jeon et al, 2022). Whereas ZP1 (ZPB) and ZP2 (ZPA; ZPY1) were up‐regulated in JQ (Table 3), ZP3 (ZPC) was up‐regulated in DCCO (Supporting Information, Table S1).…”

“…The RNA sequencing data for JQ were previously analyzed using a reference transcriptome-based method and published as a comparison of the EE2 responses between the adult and early life stage JQ (Jeon et al, 2022). We re-analyzed the early life stage JQ data using a reference-free method, Seq2Fun (Liu et al, 2021), to compare data from the two species using the same bioinformatics pipeline (Figure 2).…”