Somatic growth in vertebrates is mainly controlled by the growth hormone (GH)/insulin-like growth factor I (IGF-I) axis. The role of epigenetic mechanisms in regulating this axis in fish is far from being understood. This work aimed to optimize and evaluate the use of short-term culture of pituitary and liver explants from a farmed fish, the gilthead seabream Sparus aurata, for studying epigenetic mechanisms involved in GH/IGF-I axis regulation. Our results on viability, structure, proliferation, and functionality of explants support their use in short-term assays. Pituitary explants showed no variation in gh expression after exposure to the DNA methylation inhibitor decitabine (5-Aza-2\'-deoxycytidine; DAC), despite responding to DAC by changing dnmt3bb and tet1 expression, and TET activity, producing an increase in overall DNA hydroxymethylation. Conversely, in liver explants, DAC had no effects on dnmt s and tet s expression or activity, but modified the expression of genes from the GH-IGF-I axis. In particular, the expression of igfbp2a was increased and that of igfbp4, ghri and ghrii was decreased by DAC as well as by genistein, which is suggestive of impaired growth. While incubation of liver explants with S-adenosylmethionine (SAM) produced no clear effects, it is proposed that nutrients must ensure the methylation milieu within the liver in the fish to sustain proper growth, which need further in vivo verification. Pituitary and liver explants from S. aurata can be further used as described herein for the screening of inhibitors or activators of epigenetic regulators, as well as for assessing epigenetic mechanisms behind GH-IGF-I variation in farmed fish.

Heritability accounts for over 80% of adult human height, indicating that genetic variability is the main determinant of stature. The rapid technological development of Next-Generation Sequencing (NGS), particularly Whole Exome Sequencing (WES), has resulted in the characterization of several genetic conditions affecting growth and development. The greatest challenge of NGS remains the high number of candidate variants identified. In silico bioinformatic tools represent the first approach for classifying these variants. However, solving the complicated problem of variant interpretation requires the use of experimental approaches such as in vitro and, when needed, in vivo functional assays. In this review, we will discuss a rational approach to apply to the gene variants identified in children with growth and developmental defects including: (i) bioinformatic tools; (ii) in silico modeling tools; (iii) in vitro functional assays; and (iv) the development of in vivo models. While bioinformatic tools are useful for a preliminary selection of potentially pathogenic variants, in vitro-and sometimes also in vivo-functional assays are further required to unequivocally determine the pathogenicity of a novel genetic variant. This long, time-consuming, and expensive process is the only scientifically proven method to determine causality between a genetic variant and a human genetic disease.

The antidepressant venlafaxine can be found at levels nearing μg/L in waterways receiving municipal wastewater effluent, exposing non-target organisms, such as fish, to this chemical. We showed previously that zygotic exposure to venlafaxine alters neurodevelopment and behaviour in zebrafish (Danio rerio) larvae. Here, we tested the hypothesis that the zygotic deposition of venlafaxine disrupts endocrine pathways related to growth in zebrafish. This was carried out by microinjecting embryos (1-4 cell stage) with either 0, 1, or 10 ng venlafaxine. Zygotic venlafaxine deposition reduced the growth of fish after 30 days post-fertilization. Specific growth rate was particularly impacted by 1 ng venlafaxine. This growth retardation corresponded with the disruption of endocrine pathways involved in growth and metabolism. Venlafaxine exposed embryos displayed reduced transcript abundance of key genes involved in anabolic hormone action. Early-life venlafaxine exposure also reduced whole-body insulin and glucose content in juveniles. Target-tissue glucose uptake measurements indicated that high venlafaxine deposition preferentially increased glucose uptake to the brain. Zygotic venlafaxine did not affect feed intake nor altered the transcript abundance of key feeding-related peptides. Taken together, zygotic venlafaxine deposition compromises zebrafish growth by disrupting multiple endocrine pathways, and this study has identified key markers for potential use in risk assessment.