Abstract
Dietary and xenobiotic compounds may alter endocrine signaling and lipid homeostasis, thus inducing obesity. We describe a short-term assay method, the zebrafish obesogenic (ZO) test, for examining the effects of diet, drugs, and environmental contaminants, singly or in combination, on white adipose tissue (WAT) dynamics in live larvae. The ZO test is an intermediate step in obesity research, between in vitro and rodent assays, and may be also used to study the effect of environmental toxicants on the adiposity of aquatic species. The procedure, using Nile Red (NR) fluorescent probe to reveal adipocyte lipid droplets, is suitable for pharmaceutical or toxicological screening. Larvae treated at an environmentally-relevant concentration of tributyltin chloride (TBT), an environmental obesogen, exhibited a remarkable increase in adiposity, irrespective of the lipid composition of the background diet. Exogenous compounds, e.g., rosiglitazone or TBT, known to increase adiposity in the fasting state, were classified as obesogenic. Anti-obesogenic compounds favored a decrease in adiposity in the fasting state. The ZO test, using adipocyte lipid droplet size and adiposity as its endpoints, is a whole-organism alternative testing assay for obesogenic and anti-obesogenic compounds and mixtures and provides relevant information for environmental and human risk assessments.
Highlights
Dietary and xenobiotic compounds may alter endocrine signaling and lipid homeostasis, inducing obesity
In the experiment combining rosiglitazone or tributyltin chloride (TBT) treatment with high-fat diet (HFD), we used the univariate general linear model to check the individual effect of each factor and to determine whether there was any interaction between factors
The first step was to perform in vivo staining of zebrafish adipocytes with vital Nile Red (NR) using a rigorous feeding protocol (Fig. 1)
Summary
All zebrafish experiments were conducted in conformity with the Public Health Service Policy on Humane Care and Use of Laboratory Animals using protocols approved by the Institutional Animal Care and Use Committee of the Université Bordeaux 1 (UFR Biologie). Wild-type adult zebrafish (Danio rerio) were initially purchased from a commercial source (Exomarc, Lormont, France). Embryos and larvae were obtained by natural mating and raised in embryo water (90 μg/ml Instant Ocean [Aquarium Systems, Sarrebourg, France], 0.58 mM CaSO4, 2H2O, dissolved in reverseosmosis purified water) at 28.5°C with an 11L:13D photoperiod. Animal stages were recorded according to standard length (SL), i.e., the distance from the rostral tip of the larva to the base of the caudal fin. From five days postfertilization until they were used in the ZO test, larvae were maintained in static 5 l tanks at a density of around 30 larvae per liter and fed ad libitum with a commercial formulated food adapted for zebrafish larvae (ZF Biolabs, Tres Cantos, Spain)
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