Abstract
Zebrafish are used widely in biomedical, toxicological, and developmental research, but information on their xenobiotic metabolism is limited. Here, we characterized the expression of 14 xenobiotic cytochrome P450 (CYP) subtypes in whole embryos and larvae of zebrafish (4 to 144 h post-fertilization (hpf)) and the metabolic activities of several representative human CYP substrates. The 14 CYPs showed various changes in expression patterns during development. Many CYP transcripts abruptly increased at about 96 hpf, when the hepatic outgrowth progresses; however, the expression of some cyp1s (1b1, 1c1, 1c2, 1d1) and cyp2r1 peaked at 48 or 72 hpf, before full liver development. Whole-mount in situ hybridization revealed cyp2y3, 2r1, and 3a65 transcripts in larvae at 55 hpf after exposure to rifampicin, phenobarbital, or 2,3,7,8-tetrachlorodibenzo-p-dioxin from 30 hpf onward. Marked conversions of diclofenac to 4′-hydroxydiclofenac and 5-hydroxydiclofenac, and of caffeine to 1,7-dimethylxanthine, were detected as early as 24 or 50 hpf. The rate of metabolism to 4’-hydroxydiclofenac was more marked at 48 and 72 hpf than at 120 hpf, after the liver had become almost fully developed. These findings reveal the expression of various CYPs involved in chemical metabolism in developing zebrafish, even before full liver development.
Highlights
In recent years, zebrafish (Danio rerio) embryos and larvae have been used as alternative in vivo toxicity screening models early in the drug discovery process because of their low cost, the need for only small amounts of test drugs, and their high throughputs [1,2,3,4]
Our results revealed the transcriptional changes in 14 cytochrome P450 (CYP) isoforms, most of which are homologous to human metabolic CYP isoforms [27] in the whole zebrafish body from just after fertilization to 144 hpf, in comparison with the expression of these isoforms in the zebrafish adult liver
We found that the transcript levels of some CYP subtypes—cyp2ad2, 2k18, 2n13, 3a65, 3c/3, and 3c4 tended to rise from 96 hpf, while xenobiotic metabolism are fully operational by 5 dpf [25], suggesting that these CYP subtypes were localized in the liver
Summary
Zebrafish (Danio rerio) embryos and larvae have been used as alternative in vivo toxicity screening models early in the drug discovery process because of their low cost, the need for only small amounts of test drugs, and their high throughputs [1,2,3,4]. Several phenotype-based alternative methods for examining developmental toxicity by using zebrafish embryos have been reported and have shown high concordance (81% to 90%) with the findings of in vivo mammalian studies [6,7,8,9] Despite these advances, a problem remains in achieving more accurate predictions of the toxicity of various drugs to humans and other mammals: We still have insufficient information about the uptake and metabolism of drugs in zebrafish embryos and larvae. A lack of intrinsic capacity in zebrafish to metabolize proteratogens may lead to false negative results in teratogenicity assays of these compounds Despite these potential issues, there have been few comparisons of drug metabolism between zebrafish embryos or larvae and humans or other mammals [12]
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