Neoplasia and Neoplasm Associated Lesions in Laboratory Colonies of Zebrafish Emphasizing Key Influences of Diet and Aquaculture System Design

During the past decade the zebrafish has emerged as a leading model for mechanistic cancer research due to its sophisticated genetic and genomic resources, its tractability for tissue targeting of transgene expression, its efficiency for forward genetic approaches to cancer model development, and its cost-effectiveness for enhancer and suppressor screens once a cancer model is established. However, in contrast to other laboratory animal species widely used as cancer models, much basic cancer biology information is lacking in zebrafish. As yet data are not published regarding dietary influences on neoplasm incidences in zebrafish. Little information is available regarding spontaneous tumor incidences or histologic types in wild-type (wt) lines of zebrafish. So far a comprehensive database documenting the full spectrum of neoplasia in various organ systems and tissues in not available for zebrafish as it is for other intensely studied laboratory animal species. This manuscript confirms that as in other species diet and husbandry can profoundly influence tumor incidences and histologic spectra in zebrafish. We show that in many laboratory colonies wt lines of zebrafish exhibit elevated neoplasm incidences and neoplasm associated lesions such as heptocyte megalocytosis. We present experimental evidence showing that certain diet and water management regimens can result in high incidences of neoplasia and neoplasm associated lesions. We document the wide array of benign and malignant neoplasms affecting nearly every organ, tissue and cell type in zebrafish, in some cases as a spontaneous aging change, and in other cases due to carcinogen treatment or genetic manipulation.Keywords: Zebrafish, Hepatocyte megalocytosis, Diet, Non-protocol induced variation, Naturally occurring carcinogen, Danio rerio, Husbandry, NeoplasiaKeywords: Zebrafish, Hepatocyte megalocytosis, Diet, Non-protocol induced variation, Naturally occurring carcinogen, Danio rerio, Husbandry, Neoplasi

Effects of 3,3\u27,4,4\u27,5,5\u27-Hexachlorobiphenyl on Cytochrome P4501A and Estrogen-Induced Vitellogenesis in Rainbow Trout (Oncorhynchus mykiss)

Estrogen-regulated synthesis of vitellogenin (Vg), a yolk-protein precursor required for reproduction, was monitored to explore the potential antiestrogenic effects of the coplanar polychlorinated biphenyl (PCB), 3,3\u27,4,4\u27,5,5\u27-hexachlorobiphenyl (3,4,5-HCB), in juvenile rainbow trout (Oncorhynchus mykiss). The effects of 17β-estradiol on 3,4,5-HCB induction of cytochrome P4501A (CYP1A) were also examined. Trout were injected with 3,4,5-HCB (0.25, 2.5, 25, 50, or 100 mg/kg) or a vehicle control, and after 10 weeks, they were sampled or injected with 17β-estradiol (0.1 mg/kg). Markers of vitellogenesis, such as liver somatic index, hepatic estrogen- binding sites, and plasma Vg concentrations, in 17β-estradiol-treated fish were not affected by 3,4,5-HCB. Maximal induction of CYP1A protein and mRNA occurred at doses above 2.5 mg/kg, and 17β-estradiol reduced CYP1A protein content at a single dose (0.25 mg 3,4,5-HCB/kg). Ethoxyresorufin-O-deethylase (EROD) activity was induced by 3,4,5-HCB doses of between 0.25 and 2.5 mg/kg, but induction was reduced at higher doses, indicating that 3,4,5-HCB suppressed CYP1A catalytic activity. In 3,4,5-HCB/17β-estradiol-treated fish, plasma estradiol was significantly reduced at 100 mg 3,4,5-HCB/kg, but the depression was not associated with CYP1A induction or with other antiestrogenic effects. Although CYP1A was induced, 3,4,5-HCB did not interfere with vitellogenesis, which suggests that the PCB congener is not a potent antiestrogen in rainbow trout

CYP1B1 Knockdown Does Not Alter Synergistic Developmental Toxicity of Polycyclic Aromatic Hydrocarbons in Zebrafish (Danio rerio)

Polycyclic aromatic hydrocarbons (PAHs) are contaminants increasing in the environment largely due to burning of fossil fuels. Our previous work identified a synergistic toxicity interaction in zebrafish embryos occurring when PAHs that are agonists for the aryl hydrocarbon receptor (AHR) co-occur with PAHs that are CYP1A inhibitors. This toxicity is mediated by the AHR2, and morpholino knockdown of CYP1A exacerbated toxicity. This study tested two hypotheses: 1) in the absence of functional CYP1A, metabolism of PAHs is shunted towards CYP1B1, which has been shown in mammals to produce more reactive metabolites of PAHs; alternatively 2) CYP1B1 serves a protective role similar to CYP1A. We used a morpholino approach to knockdown CYP1B1 alone and in co-knockdown with CYP1A to determine whether we could alter deformities caused by synergistic toxicity of PAHs. CYP1B1 knockdown was not different from non-injected controls; nor were CYP1B1+CYP1A co-knockdown deformities different from CYP1A knockdown alone. These data suggest that CYP1B1 is not a significant factor in causing synergistic toxicity of PAHs, nor, in contrast to CYP1A, in providing protection

Dieldrin Pretreatment Alters [\u3csup\u3e14\u3c/sup\u3eC]Dieldrin and \u3csub\u3e3\u3c/sub\u3eH]7,12-Dimethylbenz[a]Anthracene Uptake in Rainbow Trout Liver Slices

We previously demonstrated that pretreatment of rainbow trout with the organochlorine insecticide dieldrin altered in vivo disposition of a subsequent [14C]dieldrin dose. This was not explained by changes in total lipid content or the activity of common xenobiotic metabolizing enzymes. We hypothesized that dieldrin induced hepatic proteins responsible for organochlorine (OC) sequestration, transport, or excretion and that these changes reflected an adaptive response of trout to OC exposure. Here, uptake of 1.18 μM [14C]dieldrin by precision cut liver slices was increased by dieldrin pretreatment of rainbow trout. Uptake of 0.118 and 1.18 μM [3H]7,12-dimethylbenz[a]anthracene DMBA) and efflux of 0.118 μM [3H]DMBA were significantly increased in slices from dieldrin pretreated trout. Liver slice uptake of 10 but not 1.18 μM [3H]estradiol and [3H]cholic acid was significantly increased by dieldrin pretreatment. There were no such significant differences for [3H]cholesterol, [3H]cholesterol-oleate, or [3H]oleic acid uptake. Dieldrin pretreatment did not alter hepatic microsomal metabolism of [3H]DMBA or [14C]benzo[a]pyrene or content of six cytochrome P450 isozymes, as quantitated by Western blot analysis. These results provide further evidence that altered disposition of [14C]dieldrin and [3H]DMBA in dieldrin-pretreated trout was not explained by microsomal enzyme induction but reflected altered processes integral to hepatocellular transmembrane kinetics. These changes may have important implications for OC bioaccumulation by rainbow trout and demonstrate an interaction between dieldrin and DMBA in the absence of cytochrome P450 system induction