Superfund Research Program
Developmental Toxicity of non-Dioxin-like PCBs and Chemical Mixtures
During 2010, Dr. John Stegeman’s research group continued to address the regulation of cytochrome P450 (CYP) genes that code for enzymes that metabolize environmental chemicals in zebrafish, its primary developmental model, and killifish from New Bedford Harbor, which have a heritable resistance to the toxicity of PCBs.
In zebrafish, the research group published a major work on the total complement of 94 CYP genes in this model. That included the results of microarray studies on the expression of the full complement of genes in developing zebrafish, showing unanticipated waves of CYP gene expression over the first 48 hrs after fertilization. The studies to examine selected CYP genes for potential involvement in developmental toxicity also have begun to focus on CYP expression in very early development, before the zygote begins to express genes. The researchers have found that a number of CYP genes are present as mRNA in unfertilized eggs. The results indicate that there is abundant deposition of maternal CYP transcripts in the egg and implies that there could be involvement of these CYPs in normal developmental processes or in effects of chemicals on early development.
The researchers have continued to study the regulation of CYP genes via activation of the pregnane X receptor (PXR) in both zebrafish and killifish. In zebrafish adult liver, they have confirmed that the known PXR agonists pregnenalone (P) and pregenalone carbonitrile (PCN) are able to induce both CYP2AA2, and of PXR itself. The reseachers also found that PCB153 (the most abundant ortho-substituted PCB congener) is able to induce CYP2AA2 and PXR, inducting that it, too, is a PXR agonist in zebrafish. A combined treatment with PCB153 and PCN showed an additive effect in the induction of CYP2AA2 and the PXR, indicating that this PCB does not likely have any partial PXR antagonist activity.
Studies with killifish have identified new aspects of developmental regulation of the planar (non-ortho-substituted) inducible CYP1s (the CYP1As, CYP1B1, CYP1C1, CYP1C2) and the constitutive CYP1D1 in the killifish. Analysis of the expression of these genes in three-hour intervals after hatching shows that there is up-regulation that is associated with the hatching process itself, but that the patterns differ for the five genes.
Homology modeling of the five CYP1s in zebrafish has continued to be refined. The models are being used to identify the ligand properties of these CYP1s with all 209 PCB congeners. This information will be important in determining which congeners might possibly exert toxicity via metabolites, or via uncoupling of several of the CYP1s. The predictions for metabolism or uncoupling will be addressed using the CYP1 enzymes that the research group has expressed in yeast.
These studies in both zebrafish and killifish are disclosing new insights into molecular processes that may be involved in chemical effects, especially of PCBs, in these models, and especially in developing fish.