Molecular Biomarkers of Chemically-Mediated Hepatocarcinogenesis in Laboratory-Reared and Wild Teleost PopulationsEPA Grant Number: FP916422
Title: Molecular Biomarkers of Chemically-Mediated Hepatocarcinogenesis in Laboratory-Reared and Wild Teleost Populations
Investigators: Volz, David C.
Institution: Duke University
EPA Project Officer: Manty, Dale
Project Period: January 1, 2004 through December 31, 2006
Project Amount: $111,172
RFA: STAR Graduate Fellowships (2004) RFA Text | Recipients Lists
Research Category: Fellowship - Toxicology , Academic Fellowships , Health Effects
Genome-wide expression analysis has emerged as a powerful tool in fish ecotoxicology. Although ecotoxicogenomics will undoubtedly contribute to future ecological risk assessment, laboratory-based chronic exposure studies are needed to characterize transcriptome signatures unique to many classes of xenobiotics, as well as to uncover novel biomarkers applicable for field-based surveys of reference and contaminant-exposed feral fish populations. As hepatic neoplasms are commonly observed in fish inhabiting chemically contaminated sites, the overall objective of this research is to link gene expression responses with well-known pathological indicators of toxicant-induced hepatocarcinogenesis (foci of cellular alteration, adenoma, and carcinoma) in developing teleost models. To meet this objective, we initially will exploit wild-type and see-through Japanese medaka ( Oryzias latipes ) as laboratory-based models to link gene expression profiles with histopathological data and to identify robust and conserved molecular biomarkers for hepatocarcinogenesis in fish. Medaka is a well-established model for cancer research in fish, and genetic information is becoming increasingly available for molecular-based investigations, including global gene expression analysis. Cancer-related molecular biomarkers identified from medaka then will be tested and validated in field-caught fish inhabiting contaminated aquatic systems.
Using the ubiquitous environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin ( TCDD, a non-genotoxic chemical), we will promote tumorigenesis in developing medaka following initiation by the carcinogen diethylnitrosamine (a genotoxic chemical). Although TCDD does not induce DNA damage, low doses of TCDD have been shown to promote liver tumors in mammalian (rat) models; however, the mechanisms involved in toxicant-induced tumor promotion are not clearly understood. To this end, we think that wild-type and see-through medaka are ideal for investigating genome-wide expression responses associated with hepatocarcinogenesis. For example, we recently exploited suppression subtractive hybridization (SSH) to assess gene expression changes in male medaka liver exposed for 48 hours to TCDD (10 μg/kg) ; 112 TCDD-responsive genes were identified and mainly clustered with metabolism (23%), signal transduction (12%), immune response (9%), and blood coagulation (9%). Similarly, SSH will be used to specifically target gene expression signatures associated with discrete stages of hepatocarcinogenesis in medaka following long-term, low-dose TCDD exposure. Custom small-scale gene arrays, real-time RT-PCR, in situ hybridization, Western blots, and immunohistochemistry will be used to confirm, quantify, and localize significant gene- and protein-level changes associated with hepatocarcinogenesis. In addition, following identification of strongly responsive genes associated with these histopathologic stages, focused approaches such as drug-based inhibition or morpholino knock-down will be used to identify (onco)genes critical for development of hepatic neoplasms. Based on these studies, robust biomarkers identified in medaka will be monitored in field-caught, nonmigratory Atlantic Coast killifish (Fundulus heteroclitus) resident to both reference and contaminated estuarine systems such as Elizabeth River, VA, or Diesel Creek, SC. Pollutant-resistant killifish populations are commonly found in PAH-, PCB-, and dioxin-contaminated estuaries and typically have a high prevalence of hepatic neoplasms. Collectively, these data will provide a foundation for correlating gene expression data with toxicologic histopathology and for identifying reliable and predictive molecular biomarkers for hepatocarcinogenesis in feral fish populations.