Use of a Rainbow Trout Oligonucleotide Microarray to Determine Transcriptional Patterns in Aflatoxin B1-Induced Hepatocellular Carcinoma Compared to Adjacent Liver

Department of Environmental & Molecular Toxicology, Oregon State University, Corvallis, 97331, USA.
Toxicological Sciences (Impact Factor: 3.85). 01/2006; 88(2):319-30. DOI: 10.1093/toxsci/kfi309
Source: PubMed


Hepatocellular carcinoma (HCC) is one of the most common malignant tumors worldwide, and its occurrence is associated with a number of environmental factors including ingestion of the dietary contaminant aflatoxin B(1) (AFB(1)). Research over the last 40 years has revealed rainbow trout (Oncorhynchus mykiss) to be an excellent research model for study of AFB(1)-induced hepatocarcinogenesis; however, little is known about changes at the molecular level in trout tumors. We have developed a rainbow trout oligonucleotide array containing 1672 elements representing over 1400 genes of known or probable relevance to toxicology, comparative immunology, carcinogenesis, endocrinology, and stress physiology. In this study, we applied microarray technology to examine gene expression of AFB(1)-induced HCC in the rainbow trout tumor model. Carcinogenesis was initiated in trout embryos with 50 ppb AFB(1), and after 13 months control livers, tumors, and tumor-adjacent liver tissues were isolated from juvenile fish. Global gene expression was determined in histologically confirmed HCCs compared to noncancerous adjacent tissue and sham-initiated control liver. We observed distinct gene regulation patterns in HCC compared to noncancerous tissue including upregulation of genes important for cell cycle control, transcription, cytoskeletal formation, and the acute phase response and downregulation of genes involved in drug metabolism, lipid metabolism, and retinol metabolism. Interestingly, the expression profiles observed in trout HCC are similar to the transcriptional signatures found in human and rodent HCC, further supporting the validity of the model. Overall, these findings contribute to a better understanding of the mechanism of AFB(1)-induced hepatocarcinogenesis in trout and identify conserved genes important for carcinogenesis in species separated evolutionarily by more than 400 million years.

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    • "Rainbow trout (Oncorhynchus mykiss) are widely distributed and cultured aquaculture species used as a food and sport fish (Thorgaard et al., 2002). Additionally, this organism is used as a model species in many different fields of research such as cancer biology (Tilton et al., 2005), toxicology (Köllner et al., 2002), nutrition (Wong et al., 2013), evolutionary biology (Taylor et al., 2011), and immunology (Nya and Austin, 2011). Rainbow trout are larger than other fish model species, making them a source of large quantities of specific tissues and cells for immunological, biochemical and molecular analyses. "
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    ABSTRACT: Resistance against diseases affects profitability of rainbow trout. Limited information is available about functions and mechanisms of teleost immune pathways. Immunogenomics provides powerful tools to determine disease resistance genes/gene pathways and develop genetic markers for genomic selection. RNA-Seq sequencing of the rainbow trout spleen yielded 93,532,200 reads (100 bp). High quality reads were assembled into 43,047 contigs. 26,333 (61.17%) of the contigs had hits to the NR protein database and 7024 (16.32%) had hits to the KEGG database. Gene ontology showed significant percentages of transcripts assigned to binding (51%), signaling (7%), response to stimuli (9%) and receptor activity (4%) suggesting existence of many immune-related genes. KEGG annotation revealed 2825 sequences belonging to "organismal systems" with the highest number of sequences, 842 (29.81%), assigned to immune system. A number of sequences were identified for the first time in rainbow trout belonging to Toll-like receptor signaling (35), B cell receptor signaling pathway (44), T cell receptor signaling pathway (56), chemokine signaling pathway (73), Fc gamma R-mediated phagocytosis (52), leukocyte transendothelial migration (60) and NK cell mediated cytotoxicity (42). In addition, 51 transcripts were identified as spleen-specific genes. The list includes 277 full-length cDNAs. The presence of a large number of immune-related genes and pathways similar to other vertebrates suggests that innate and adaptive immunity in fish are conserved. This study provides deep-sequence data of rainbow trout spleen transcriptome and identifies many new immune-related genes and full-length cDNAs. This data will help identify allelic variations suitable for genomic selection and genetic manipulation in aquaculture.
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    • "Omic-based approaches have been reported for other fish, identifying key factors in the pathways of diseases or in carcinogenesis. For example, in O. mykiss, oligonucleotide microarray studies identified the mechanism of aflatoxin B 1 -induced hepatocarcinogenesis (Tilton et al., 2005). Gene expression profiling from cDNA arrays revealed that the tumor-promoting activities of perfluorooctanoic acid were related to estrogenic signaling pathways in animals fed 200–1800 ppm perfluorooctanoic acid (Tilton et al., 2008). "

    Full-text · Article · Apr 2014 · Marine Pollution Bulletin
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    • "Because PFOS trout were treated at a later age, a separate time-matched reference RNA pool was prepared for competitive hybridization of PFOS samples. Details on the development, manufacture, and quality control assessment of the OSUrbt version 5.0 microarray have been provided previously (Benninghoff and Williams, 2008; Tilton et al., 2005) (Gene Expression Omnibus [GEO] platform accession ID: GPL5478). For detection of gene expression on the OSUrbt-v5 array, the Genisphere 3DNA Array 900 kit (Hatfield, PA) was used according to the supplier's protocol in a standard dye-swap reference sample design as previously described (Benninghoff and Williams, 2008). "
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    ABSTRACT: Previously, we reported that perfluorooctanoic acid (PFOA) promotes liver cancer in a manner similar to that of 17β-estradiol (E2) in rainbow trout. Also, other perfluoroalkyl acids (PFAAs) are weakly estrogenic in trout and bind the trout liver estrogen receptor. The primary objective of this study was to determine whether multiple PFAAs enhance hepatic tumorigenesis in trout, an animal model that represents human insensitivity to peroxisome proliferation. A two-stage chemical carcinogenesis model was employed in trout to evaluate PFOA, perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), perfluorooctane sulfonate (PFOS), and 8:2 fluorotelomer alcohol (8:2FtOH) as complete carcinogens or promoters of aflatoxin B(1) (AFB(1))- and/or N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-induced liver cancer. A custom trout DNA microarray was used to assess hepatic transcriptional response to these dietary treatments in comparison with E2 and the classic peroxisome proliferator, clofibrate (CLOF). Incidence, multiplicity, and size of liver tumors in trout fed diets containing E2, PFOA, PFNA, and PFDA were significantly higher compared with AFB(1)-initiated animals fed control diet, whereas PFOS caused a minor increase in liver tumor incidence. E2 and PFOA also enhanced MNNG-initiated hepatocarcinogenesis. Pearson correlation analyses, unsupervised hierarchical clustering, and principal components analyses showed that the hepatic gene expression profiles for E2 and PFOA, PFNA, PFDA, and PFOS were overall highly similar, though distinct patterns of gene expression were evident for each treatment, particularly for PFNA. Overall, these data suggest that multiple PFAAs can promote liver cancer and that the mechanism of promotion may be similar to that of E2.
    Preview · Article · Jan 2012 · Toxicological Sciences
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