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Environmental Factor, October 2012

Models take spotlight at council science talks

By Ernie Hood

Robert Tanguay, Ph.D.

Tanguay and his group, whose motto is “Thinking outside the tank,” have pioneered new methods allowing zebrafish assays to be run in high-throughput equipment. “We realized that there were a lot of ways to impact the environmental health sciences field, if we could automate several of the steps in the process,” he told Council. (Photo courtesy of Steve McCaw)

Zebrafish

Thanks to innovations by Tanguay’s lab and others, the humble zebrafish has achieved supermodel status in the scientific world. (Photo courtesy of Robert Tanguay)

Advances in animal models were the focus of two scientific presentations at the Sept. 11 National Advisory Environmental Health Sciences Council meeting. Zebrafish and mice have been model organisms in environmental health sciences for a long time, but today researchers are crafting bold innovations to advance their usefulness as vehicles for scientific inquiry into toxicity and disease.

Council members were treated to exciting, informative talks by NIEHS Superfund Research Program grantee Robert Tanguay, Ph.D., (http://emt.oregonstate.edu/roberttanguay)  a distinguished professor of molecular toxicology and head of the Sinnhuber Aquatic Research Laboratory at Oregon State University; and lead researcher Jef French, Ph.D., head of the NTP Host Susceptibility Group within the Biomolecular Screening Branch.

Harnessing the power of zebrafish

In Tanguay’s lab, the goal is to link phenotype with genotype, using zebrafish, which closely resemble humans in terms of responses to environmental insults and disease traits. Several years ago, Tanguay realized there were major bottlenecks in the process. He and his team used funding from the American Recovery and Reinvestment Act of 2009, to focus intensive efforts on streamlining and accelerating the lab’s ability to produce the fish and run them efficiently through toxic exposure assays.

“We needed to crank it up,” he told the Council audience. “We identified that if we were going to take full advantage of this model, we needed to massively increase the robustness of phenotype discovery, to link exposure to phenotype, to identify a mechanism.”

Crank it up they did, achieving a quantum leap in experimental productivity by engineering several ingenious new processes to break the bottlenecks, including highly automated methods to allow unlimited production of zebrafish embryos, nondestructive handling of the embryos, and chorion removal, where the embryo’s outer membrane is excised to permit direct exposure of the developing embryo to challenge toxicants. Those innovations allowed the use of high-throughput technology to run experiments designed to gauge the impact of exposures, including mixtures, on the very early development of the zebrafish, with measurable endpoints in just five days.

Today, thanks to his group’s technological advances, the zebrafish is poised to take on supermodel status. “We can translate the data in many different ways, such as identifying inherently hazardous compounds,” said Tanguay. The approach also supports a process he calls binning. “Once you have the ability to massively do exposures with lots of different compounds, with known structures or known mixture composition, you then have the ability to associate the chemical structure and the composition with the biological responses you get, and start binning the compounds based on the commonalities of responses and structure. And, then, you can drill down from representative members of these families to define the actual cause of toxicity.”

Mouse models have it both ways

French shared the results of recent experiments with two types of mouse models — a fixed-genotype, inbred, 18-strain panel known as the NTP panel, and a highly genetically diverse mouse population called the Diversity Outbred (J:DO) mouse, where random outbreeding was used to scramble the genomes of eight laboratory and wild-derived mouse inbred strains into a new wild-type mouse population, where every mouse is genetically different from every other mouse.

He described assays exposing both models to benzene, a ubiquitous toxicant in the environment with a large amount of existing data on the effect on metabolism, toxicity, and carcinogenicity, which are remarkably similar between humans and rodents.

A haplotype association mapping approach was used with the inbred strains to allow characterization of their benzene absorption, distribution, metabolism, and excretion phenotypes.

Experiments with the J:DO mice were designed to assess benzene-induced hematotoxicity and genotoxicity, following 28 days of controlled inhalation exposures at low levels.

According to French, both approaches — measuring inter-individual variability and population-level variability — are valuable in the effort to link genome-by-environment interactions with human outcomes. They will also help to quantify the uncertainty factors in correlations between mouse and human.

“Based on the studies performed using the J:DO mouse population, we conclude that population-based models are warranted in in vivo toxicology, in order to determine the role of inter-individual variability, and identify resistance and susceptibility alleles associated with population-based outcomes,” French said. “I emphasize that we have identified a number of candidate genes that may be major determinants of these phenotypes, but after this discovery phase, using the forward genetics approach, we must return to reverse genetics and molecular biology, to validate their function and the mechanistic basis for the observed toxicity.”

(Ernie Hood is a contract writer with the NIEHS Office of Communications and Public Liaison.)


Slide form Jef French's presentation

Representative genomic structure maps, and photos of an inbred Collaborative Cross mouse, left, and outbred J:DO siblings, underscore the extent of diversity found in the outbred population.(Photo courtesy of Jef French)


Jef French, Ph.D.

French said he appreciated the opportunity to update Council on advances in experimental mouse models. French has built upon insights from the NIEHS Mouse Genome Resequencing and SNP Discovery Project, in the search for a mouse model with genetic diversity more similar to that found in humans.(Photo courtesy of Steve McCaw)




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