Abstract
Cetaceans have been found to bioaccumulate high aromatic hydrocarbon tissue residues, and elevated levels of PCB residues in tissues are proposed to have occurred concurrently with recent epizootic deaths of dolphins. The objectives of this study were: (1) to develop and characterize an epithelial cell line derived from dolphin tissues, (2) to investigate the effects of hydrocarbon pollutants on those cells, and (3) to analyze the toxicity of hydrocarbon pollutants on cetacean tissues in vitro. An epithelial cell line, Carvan dolphin kidney (CDK), isolated from a spontaneously aborted female bottlenose dolphin, Tursiops truncatus, grew rapidly with high plating and colony forming efficiencies. These cells were neither transformed nor immortal, displaying contact inhibition, anchorage dependence, the requirement for high concentrations of fetal bovine serum for growth and a stable 2N = 44 karyotype. Velocity sedimentation analysis showed that CDK cells contained nuclear aryl hydrocarbon receptor, suggestive of cytochrome P450 inducibility. CDK cells were exposed in vitro to BaP with/without TCDD and/or the cytochrome P450 inhibitor α-naphthoflavone. BaP inhibited mitosis in CDK cells in a dose-dependent manner. Data indicate that CDK cells metabolize BaP, that BaP metabolites bind to cellular DNA initiating unscheduled DNA synthesis, and that the inhibition of cytochrome P450 metabolism decreased the BaP-associated inhibition of mitosis in dolphin cells. The data also suggest that TCDD acts synergistically to increase the levels of DNA damage by the procarcinogen BaP. Cetacean liver microsomes were isolated and evaluated for the presence of cytochrome P450 proteins by SDS-PAGE, apparent minimum molecular weight determination, and immunoblot analysis. None of the putative P450 proteins hybridized with monoclonal antibodies that recognize other mammalian P450s. P450 activity was induced in cetacean tissue samples and CDK cells by exposure in vitro to one of several cytochrome P450-inducing chemicals. The data suggest that cetacean tissues and cells can be utilized to study the in vitro induction of cytochrome P450, resultant metabolism of xenobiotic contaminants, and the subsequent cellular and molecular responses. However, the identity of specific P450 isozymes involved in this process will remain undetermined until monoclonal antibodies that recognize cetacean P450s can be generated.
Carvan, Michael John (1993). In vitro toxicity of polycyclic aromatic hyrocarbons and halogenated aromatic hydrocarbons to cetacean cells and tissues. Texas A&M University. Texas A&M University. Libraries. Available electronically from
https : / /hdl .handle .net /1969 .1 /DISSERTATIONS -1523715.