Oregon State University

The

Linus Pauling
Institute
SPRING-SUMMER 2015 RESEARCH NEWSLETTER

From the Director

Fish Toxicology

Balz Frei, Ph.D.

An Interview with
Robert Tanguay, Ph.D.

LPI Director and Endowed Chair
Distinguished Professor of Biochemistry
and Biophysics
Joan H. Facey LPI Professor

T

he Linus Pauling Institute recently added four new
investigators to its faculty roster, who joined theInstitute as Adjunct members: Drs. Claudia Maier, Arup
Indra, Robert Tanguay, and Siva Kolluri, all from Oregon
State University. Their role as Adjunct Faculty is to supportand help advance LPI’s research program by collaboratingwith other LPI faculty on research projects,
grant proposals, and shared methodologies.
Dr. Maier is a Professor of Chemistry andan expert in mass spectrometry applied tothe analysis of biomolecules, including
proteins and metabolites (called proteomics
and metabolomics, respectively), and theClaudia Maierstudy of the interactions of these proteinsand metabolites in human blood and cells(for an article on Metabolism, see ourSpring/Summer 2013 Newsletter). Dr. Indra,an Associate Professor of Pharmaceutical
Sciences, is studying melanoma, inflammatory
skin diseases like atopic dermatitis, andArup Indranatural compounds for effective therapeuticinterventions. Dr. Kolluri, an Associate
Professor in the Department of Environmental
and Molecular Toxicology, is developing
novel agents that selectively kill cancer
cells, but not normal cells, and is devising
optimal combinations of therapeutic agents
Siva Kolluri
for the prevention and treatment ofdifferent types of cancers. You can readmore about Dr. Tanguay’s exciting researchin the world-class zebrafish facility that hebuilt at OSU in this Newsletter (at right)
and will hear more about the research in
the other Adjunct Faculty’s laboratories inRobert Tanguay
future issues of the Newsletter.
continued on page 2

LPI Adjunct Faculty

Q. You earned your Ph.D. from the University ofCalifornia-Riverside about 20 years ago.
What was your thesis?
A. Post-transcriptional regulation of gene expression.
Q. What does that mean?
A. Prior to that era, few scientists thought that gene expression
could be regulated outside the nucleus or after messenger
RNAs were produced. Messenger RNA (mRNA)
conveys information from DNA to the cellular machinery
where proteins are made. My advisor Daniel Gallie and
I proposed that there are elements within RNAs that
regulate how much protein is produced from a given
messenger RNA. More specifically, we proposed for the
first time a circular translation model where the ends ofmRNA were looped backed and intimately connected
through proteins. Our work and a number of otherpapers demonstrated that our hypothesis was correct,
and now it is well established that post-transcriptional
mechanisms are very important in all species. Ultimately,
my interest in gene expression really stemmed from theRNA universe I was studying as a graduate student.

Q. Where did you grow up?
A. I’m from the upper peninsula of Michigan. I lived there

until my junior year in high school, and then we moved to
California, where I finished high school, my undergraduate
training in biology at California State UniversitySan Bernardino, and graduate school in biochemistryat the University of California-Riverside. I did my
post-doctoral training for 3½ years at the University of
Wisconsin in Madison with Richard E. Peterson. That iswhere I developed the zebrafish model for use in toxicology
in 1996. I got my first faculty appointment at the
University of Colorado Health Sciences Center in thecontinued on page 2

Optimum Health conference. Re-designing the LPI
website and its Micronutrient Information Center (MIC)would not have been possible without the painstaking
work of my assistant, Barbara McVicar; LPI’s MIC
manager, Victoria Drake; and many others in theInstitute. I invite you to visit and explore our new
website (lpi.oregonstate.edu), including the MIC(lpi.oregonstate.edu/mic), and send us your feedback
and suggestions for further improvements!

Continued from cover — From the Director
I am also very pleased to report that
LPI’s Joe Beckman has recently been
awarded a $2.1 million grant fromthe Department of Defense, entitled“Development of Copper ATSM asa Therapeutic for SOD-Familial andSporadic ALS” (amyotrophic lateral
Joe Beckman
sclerosis, also known as Lou Gehrig’s
disease). This project is based on
Dr. Beckman’s recent breakthrough discovery that thecopper complex, Cu-ATSM, is exceptionally effective
in delaying the onset of ALS in a mouse model of thedisease. The DOD grant will allow Dr. Beckman todesign new chemical derivatives of Cu-ATSM targeted
specifically to the mitochondria of motor neurons,
where the copper is released and triggers the protective
mechanisms against the neurons’ death. This research will
help move Cu-ATSM and these novel derivatives towards
human clinical trials to test their safety and efficacy.
Although we still have a long road ahead of us, if
successful, this could become the first treatment everfor Lou Gehrig’s disease.

The 2015 Diet and Optimum Health conference, to beheld September 9-12 on the OSU campus in Corvallis,
Oregon, emphasizes dietary and lifestyle approaches,
including exercise and dietary supplementation, toadvance human healthspan and prevent age-relateddiseases. We are excited to have been able to get thecommitments of many outstanding scientists from aroundthe world to present their work at our conference on suchimportant topics as “Vitamin E: A Critical Nutrient forDevelopment and Health,” “Extending Healthspan:
Basic Concepts, Underlying Mechanisms, and Role ofDiet and Lifestyle,” and “Cancer Chemoprevention.”
The conference also features a session open to the public
focusing on “LPI’s Rx for Health: The Role of Diet,
Micronutrients, and Dietary Supplements in Human
Health and Disease.” The complete program of theconference can be found on pages 12-13 of thisNewsletter—which I now invite you to enjoy reading! LPI

Two other projects have kept us busy in the Linus
Pauling Institute in recent months—the re-design of
our website and the eighth installment of our Diet and

Q. About 15 years ago you developed an interestin zebrafish and trout as useful models in
toxicology. Why fish?
A. Good question. When I was offered an NIH fellowship

Continued from cover — Fish Toxicology
School of Pharmacy in 1999. I was then recruited todirect the Sinnhuber Aquatic Laboratory and to join
OSU’s Department of Environmental and Molecular
Toxicology as an Associate Professor in 2003.

to go to Wisconsin, I had a choice of 20 labs to join on
the training grant. I interviewed Richard Peterson, who
has a long track record in evaluating the toxicity ofenvironmental chemicals in applied research using lake
trout and rainbow trout. He determined how much ofa chemical was killing these fish in the Great Lakes.
This led to some dramatic changes in regulation andsaved fish populations in the Great Lakes. But at that time
the Peterson lab had limited experience with molecular
explorations in those models. When I went there, I
didn’t want to work with trout because I became awareof a new model—zebrafish—in toxicology and humanhealth. I didn’t know anything about toxicology. I had
never taken a toxicology course and still haven’t, but I
just saw there was a huge potential to solve problems intoxicology with this new model.

Q. What led you to Oregon State University?
A. Whenever you move there is push and draw. The push

was I had large plans to grow my program that could
not be accommodated in a typical School of Pharmacy.
The draw was that I saw tremendous potential everywhere on this campus, and I thought I could help leadadvancements. I’m the type of person who likes to buildand nucleate programs. I enjoy collaborating withothers, and OSU has a tremendous collaborative spiritthat can be addictive and rewarding.

Q. Some of your initial research was on plants.
What did you work on?
A. In plants, I was trying to understand how an RNA

Q. Is the zebrafish a good model for human diseases
like cancer and heart disease or is it limited to
toxicology?
A. I think that the field has advanced faster outside of thesequence could possibly modulate how much protein
is made. I’m a reductionist, really, and like tounderstand processes at the most fundamental level;
I had little inherent interest in plants but was reallyinterested in understanding evolutionarily conserved
molecular machinery. Funding also influenced mydecision to cross over into a vertebrate model.

S P RI NG /SUM M ER 2 0 15

toxicology realm. There are hundreds or thousands ofinvestigators using the zebrafish model for various

2

aspects of human disease, including heart disease andcancer, for example, in the discovery of tumor
suppressor genes, oncogenes, and therapeutics.

Q. What are the specific attributes of the zebrafish
model that make it more compelling than arodent model?
A. They are numerous, and we try to exploit all theadvantages and avoid the disadvantages with a lot of
discipline. For example, we now know that the zebrafish
genome is remarkably well conserved, sharing about
70% with humans, and of the human genes known to
be involved in disease, 84% are also found in zebrafish.
If you look at individual biochemical pathways, they are
all there. And then there are massive technical advantages.
We can investigate almost every human disease in thismodel at a fraction of the cost and a fraction of the time,with very few limitations. Most of the limitations are
truly just matter of perception or unintended biases byscientists unfamiliar with the strengths of the model.

Q. Is it more economical to use zebrafish rather
than rodents?
A. Absolutely. You still have the same animal care and use
considerations and the same compliance requirements,
but the cost of housing is dramatically lower. We haveseveral hundred thousand zebrafish that are supporting
dozens of researchers on this campus.

Q. I imagine that the low cost and high numbersof fish increase the statistical power of theseexperiments.
A. It certainly does, but more importantly, it frees yourimagination. If you talk to researchers who areprimarily using rodents for their research, the number
one consideration that stops them from doing an
exciting experiment is cost. I do not like barriers to
my imagination.

Zebrafish

Q. What kinds of toxins have you studied using
zebrafish?
A. We have tested thousands of compounds, including

I just saw there was ahuge potential to solve
problems in toxicology
with this new model.

pharmaceuticals, dietary constituents, pesticides, industrial
chemicals, herbicides, chemical libraries, environmental
contaminants, and real-world mixtures.

Q. Are the concentrations of chemicals to which you
expose the zebrafish embryos relevant to thosethat humans might be exposed to?
A. All of our studies are completed at relevant ranges, but

because of the low cost, we don’t limit it just to that. We
evaluate broad ranges; we may do a typical study with fiveorders of magnitude evaluations in large numbers of fish,giving massive statistical power. It allows comparative
analysis of different chemicals based on their structure.

continued on page 4

3

THE LINUS PAULING INSTITUTEWe do that because I’m interested in finding compounds
that affect development. By definition, chemicals that
affect development have the ability to interact withand alter evolutionarily conserved molecular pathways.
These chemicals may pose risks to humans, or they may
have therapeutic value. We use zebrafish as the ultimate
biological sensor.

Continued from page 3 — Fish Toxicology

Q. What kind of defects do you typically observein embryos that are treated with pesticides
or herbicides?
A. We’ve spent almost 20 years doing this, trying to identify

anything that’s different. That may be the length of theanimal, the diameter of the eye, the axis, the spinal cord,
vascular development, heart rate, or complex behaviors.
All these can be measured. Since we have studied over
100,000 compounds, we know that many chemicals
produce the same effects. What that means is thatdevelopment is so integrated and dynamic that there arealmost countless ways that you can disrupt it to lead tothe same problem. For example, in order for dioxin tocause cardiovascular failure, you need to have a certain

A five-day-old zebrafish in a microwell plate

Q. Could some of your results be applied toamphibians like salamanders and frogs, or are
they more sensitive than fish?
A. For some chemical classes, amphibians seem to be moresensitive. But my goal is not to translate my findings
directly to amphibians. Many of the biochemical pathways that we’ve discovered certainly exist in amphibians,
and I will let the amphibian experts translate my results
to their studies. Because I work with zebrafish, people
think I am a “fish guy,” but I really am not. Part of myresearch focus is to understand chemicals—how they
modify a biological target or pathway. I want to discover
the structural attributes of the chemicals so that we
can identify their targets. If we do this well, we caneventually predict the activity of all chemicals. As you
can imagine, there would be tremendous societal value
in reaching that goal. We do not do one chemical ata time or one pathway at a time but rather systems
approaches to tackle bigger problems.

24-hour-old zebrafish embryos

cellular receptor that is directly activated by dioxin. If
you delete that receptor in fish or other animals, dioxin
doesn’t cause problems.

Q. Do you usually treat zebrafish embryos withchemicals and watch for developmental defects?
A. We actually ask the embryos, in as many ways as we can

Q. Do you test these toxins in adult fish?
A. We certainly do some exposures in adult fish—overimagine, to tell us if a chemical has biological activity.
Does it cause changes in heart development, eyedevelopment, hearing, swimming behavior, learning andmemory, anxiety, social behavior, fear responses, skeletal
problems, gene expression (mRNAs, non-coding RNAs,
and proteins), or metabolism.

three months old—but you can’t do rapid screening
in adults. With the genetic tools at our disposal weare increasingly doing more adult studies, but the trueadvantages of zebrafish are the early life stages. Staywith the strengths of your model is something I preachto my students and other scientists.

Q. How long do you follow these embryos?
A. For the early life stage studies we typically follow them for

Q. How long do the fish live?
A. Zebrafish can live up to three years. They are typically

five days. In that five-day period, the embryo goes froma single cell to an animal that has all of the functioning
organs—the heart, the brain, the kidneys, the liver, etc.

S P RI NG /SUM M ER 2 0 15

in our facility about a year.

4

Q. You also investigated the effect of embryonic
exposure to alcohol and various flame retardants
that are ubiquitous these days. What did you
find in those experiments?
A. The ethanol work started when I was a post-doctoraland life stage. We have all the tools necessary to domuch more microRNA work, which I really enjoy as
this gets me back to my scientific roots in posttranscriptional regulation of gene expression.

Q. Do the behavioral effects caused by alcohol
persist into adulthood?
A. We haven’t done the behavioral analysis in adults that

fellow. I wanted to see whether or not I could use zebrafish to study fetal alcohol syndrome. Amazingly, we still
don’t fully understand how ethanol produces the typicalcranial facial malformations and, more importantly, thelearning and memory deficits. And we don’t understand
why there is such a variation of responses between people
exposed to similar doses of ethanol. We published anumber of papers demonstrating that the zebrafish model
is appropriate to study the developmental responses toethanol exposure. In particular, we are looking at the
involvement of some microRNAs that are regulated in
response to ethanol exposure. MicroRNAs are small,non-coding pieces of RNA that regulate gene expression.
They were discovered about 20 years ago. These 22nucleotide modulators can bind recognition sequences
through the genome and affect gene expression in any
number of interesting ways.

other collaborators have. In early development wetypically identify either hyper- or hypo-activity inswimming behavior. When we raise these fish to thejuvenile and young adult stage, we see hypo-activity inswimming that persists in adults. We developed a whole
roomful of new instruments to assess higher orders oflearning and memory. We can evaluate the effect ofchemicals on decision-making in fish.

Q. What about flame retardants?
A. The study of flame retardants is a very active area in mylab right now. We just completed the world’s largest
evaluation of broad classes of flame retardants inzebrafish. We looked at 44 different flame retardants
in four different chemical classes. Flame retardants
are required by law to be in many consumer products,
and there are hundreds of millions of tons used worldwide. But some of these chemicals are not as bad asothers. There are hundreds of different flame retardant
compounds used commercially, but many people groupthem all together based on their commercial use. Thisis equivalent to suggesting that all the food we consume
is equally healthy because it is all food. I maintain thatsome flame retardants are safer than others. I want to
identify which of these chemicals are intrinsicallybioactive and which ones aren’t and then advise theindustry so that they can make the safe and effective ones.

Q. What environmental pollutants or carcinogens
have you studied using zebrafish?
A. We have lookedat hundreds ofdifferent compounds—polycyclicaromatic hydrocarbons (PAHs),
dioxins, and PCBs.
There is much tobe learned. Not all
carcinogenic
compounds are
48-hour-old zebrafish embryo
mutagenic, so weare not looking formutagenicity in our model. Can we use this model toidentify mechanisms and relate those to the chemical
structure of the compound? If we test 50 compounds
that produce a similar response—for instance, molecular
networks are perturbed in a similar way—then we may
be able to predict the biological effects of many otherchemicals with similar structures. Again, we thinkpredictively, not one chemical at a time.
continued on page 6

Automated embryo plating system

Q. So they are regulators of protein synthesis?
A. Absolutely, they are. It’s amazing what these can do. And
they are incredibly well conserved evolutionarily acrossspecies. In fact, some of the microRNAs that we discoveredare 100% identical between humans and zebrafish.

Q. How many microRNAs are known?
A. In zebrafish, there are under 400. In humans, it’s more
like 800. But discovering them is one thing; identifying
what they do is another. What they do varies by tissue

5

THE LINUS PAULING INSTITUTE

A. Nanoparticles are touted as the foundation for the next

Continued from page 5 — Fish Toxicology

technological revolution. Nanomaterials are less than
100 nanometers, or about 100 billionths of a meter, in
one dimension. Nanoparticles called fullerenes are perfectspheres made from 60 atoms of carbon. We proposed in2005 that we could use the zebrafish embryo totest interactions between nanoparticles
and biology.
_

Q. With Maret Traber’s lab you published a numberof papers on induced vitamin E deficiency inzebrafish embryos. What did you find?
A. The first challenge was to generate afully defined zebrafish diet—to_
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3
reproductively functional as
Flexible
displays,next-generation
adults. That was the first big hurdle.
Functionalized gold nanoparticle
computers, batteries, solar cells, pesticides,
Together, we found that when we fed thefoodpackaging, diagnostic imaging, LEDzebrafish parents the vitamin E-deficient diets, theylighting,building
materials, and catalysts all use nanoproduced embryos that had poor viability. So there
particles.
In
medicine,
one of the huge advantages ofwere developmental defects in the embryos—the lack ofnanomaterials
isthesurfacearea that you gain. You
vitamin E prevented normal vertebrate development.
can take a sphere that is a millimeter in diameter anddecorate it with functional drugs. Now if you can do
Q. What were the effects in these embryos fromthis in nanoscale, the surface area and opportunities forparents that were depleted of vitamin E?
multi-functional, drug delivery devices become fantastic.
A. There was clear evidence that vitamin E was necessary
to accomplish normal vertebrate development. This had
Q.Your work is focused on gold and silver
been hypothesized in the literature on mammals, but it
nanoparticles, sometimes coated with various
was always difficult to separate out the effects of vitaminchemicals. What does the chemical treatment
E on the mother and the effects in utero.
achieve, and do you find that some chemical

N+

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treatments of these nanoparticles might
attenuate toxicity?

Q. Since vitamin E mainly functions as anantioxidant, oxidative stress probably played arole in those defects that you observed.
A. More than likely, but that doesn’t necessarily mean that

A. This is relevant to our “green” chemistry focus and “green”
nanotechnology with my collaborator, Jim Hutchison,at the University of Oregon. The gold nanoparticles area good model. We want to learn something about thefeature of a particle that might predict what it can do.
That knowledge will help us to develop better
performing products that are also safer. For example,
we can put different functional chemical groups on
nanoparticles and ask what impact does thissubstitution have on bioactivity? Is it the core, the size,or the actual chemical group? We’ve learned that thefunctionality of the surfaces of gold nanoparticles, inoxidative damage is the only factor. We now understandoxidative signaling, and since there are developmentally
important genes, it’s my hypothesis that by changing theoxidative environment you change signaling in subtle
ways that affect gene expression. Additionally, changes
in membrane integrity or function may be main drivers.

Q. You’ve also had a long-term interest in the
toxicology of nanoparticles, including fullerenes.
What are nanoparticles?
S P RI NG /SUM M ER 2 0 15

6

particular, is the most important. We can now playaround with those functionalities. If you are
manufacturing a compound for industrial purposes orfor clinical use, understanding these rules is very
important. We are trying to come up with the generalrules for building safe but functional particles.

Q. Your major focus has been on the disruption ofdevelopment in zebrafish embryos treated with
nanoparticles and the persistence of those defects
into adulthood. Have you checked nanoparticle
toxicity in adult fish?

Q. You found that the treatment of zebrafish withgold nanoparticles causes eye and neuronal
problems, as well as defects in adult swimming
behavior. Did you figure out why the gold
nanoparticles caused these effects?
A. Not yet, but we are still working on it. We know thatstage. We have done a little bit of work with adultexposures, but the cost of materials is huge because you
have to use tanks of water instead of miniscule amounts
needed for embryos. We only need 50 or 100 microliter
volumes with just a few micrograms of particles to doa full bioactivity profile with embryos. With our new
instruments we can measure learning and memory,social behavior, respiratory function, and cardiovascularfunction, as well as pathology.

A. The strength of our model is in the developmental

gold nanoparticles induce changes in neuronaldevelopment and apoptosis, or cellular death.

Q. With apoptosis, you might be losing cells that are
critical to development.
A. Exactly.

Q. Are results from zebrafish experiments likely to be
applicable to humans?
A. Absolutely. We have dozens and dozens of papers to

Q. Are zebrafish the best model to screen for
nanoparticle toxicology?
A. I would say zebrafish is the best model by far for many

indicate that. The zebrafish field in general has put thisquestion to rest, which explains the exponential rise infunded zebrafish research and a decline in funding forother models.

of the reasons I’ve discussed. The work we are doing
absolutely cannot be done in laboratory rodent models.
Again, stay with the strength of your model.

Q. Are people exposed environmentally tonanoparticles, or would exposure be limited to acertain context like manufacturing?
A. Right now, there is limited environmental exposure to

Q. How do you treat the embryos with nanoparticles?
Are they injected?
A. Typically, we put nanoparticles in their water or inject them.

nanomaterials. I’m part of a large grant at ArizonaState University where we’re trying to assess the likely
routes of exposure. For example, you might make atennis racket with carbon nanotubes, but the likelihoodof a carbon nanotube being liberated during a tennis
game is pretty small. However, if you worked at anaircraft-manufacturing facility and part of the fuselage
is a carbon nanotube material, sanding surfaces would
create a fair amount of dust. The good news is that
research has demonstrated that personal protective
equipment can be very effective in the work place.

We can modulate the aqueous environment in such a waythat it affects the dispersion of nanomaterials, which is a
powerful advantage. When you put nanoparticles in an
ion-containing medium, they sometimes aggregate, which
affects bioavailability. Some of the data from mammaliancell-based models are complicated by the media in which
the cells are grown. And we don’t have that problem. We
can run our assays under various ionic conditions to confirm
the dispersion of the particles before we do the studies.

Q. Have nanoparticles found their way into
commercial products that people commonly use?
A. There are hundreds of commercial applications ofnanomaterials already in the cosmetics industry inconsumer products like creams and sunscreens,and also in paints, coatings, building materials,
airplanes, fabrics, and food.

continued on page 8

Carbon nanotube

7

THE LINUS PAULING INSTITUTE

Q.What do you think are the important lessons toapply to future work with nanoparticles?
A. I think getting a strong focus on green chemistry andContinued from page 7 — Fish Toxicology

Q. Would nanoparticles be absorbed through theskin, ingested, or inhaled?
A. Absolutely. Some applications in medicine, food, andgreen nanotechnology in the beginning to avoid mistakes
is important—trying to identify hazardous nano featuresprior to releasing 100,000,000 metric tons of a product.
And that is working. We’ve tested hundreds of different
nanomaterials, and most appear biologically benign.

cosmetics are designed to do so. Most of the exposure,
however, I suspect will be from manufacturing or release
into the environment at the end of product life.

Q. People are concerned about safety and arewilling to apply some guidelines to thedevelopment and production of some of thesecompounds. Historically, many chemicals weresynthesized and released into the environment
without establishing an adequate safety profile.
A. I totally agree. We’ve been fortunate none of theQ. There is a possibility that nanoparticles will findtheir way into the water supply. Are common
water-treatment processes adequate to remove
nanoparticles?
A. No, they’re not completely adequate. There are a lotof arguments about this in the field. The bulk of nanomaterials don’t dissolve. Silver nanoparticles are themost well-studied and widely used. Silver nanoparticles
have antimicrobial uses in clothing, wound dressing,band aids, and the linings of washing machines. Thereis a very slow release of ionic silver, and ionic silver isantimicrobial. If the silver nanoparticle gets into theenvironment, one of three things will happen. It’s either
going to completely dissolve into ionic silver, in whichcase it’s not a nanoparticle any longer. It may stay inthe silver form and get chemically modified by bacteria,
which changes the dissolution rate. Or third, it’s justgoing to sediment with other stuff in water treatment assludge. Sludge is removed from our drinking water but
sometimes dispersed back into the environment.

nanomaterials are likely to be really nasty. The biggest
application of nanotechnology has been in diagnostics—
special chips, diagnostic tools, and biomarker detection.

Q. You were recently invited to join LPI as an
adjunct professor. Why does this positioninterest you?
A. I have so many collaborations on this campus, and manyof them are with LPI investigators. As I mentioned earlier,
I’ve been working with Maret Traber for about eight
years now, and I have strong collaborations with LPI’s
Dave Williams and Emily Ho.

Linus Pauling DayA◊

s has been done in prior years, the Governor of Oregon
issued a proclamation that declared Linus Pauling’s
birthday, February 28, “Linus Pauling Day.”

Pauling was lauded for his two Nobel Prizes and his“courageous efforts for world peace and to ban the testingof nuclear weapons,” as well as for his seminal andmomentous scientific accomplishments that included
establishing the basis of modern chemistry; the discoveryof the cause of sickle-cell anemia—the first disease to be
described as a molecular disease; and setting the stage forthe science of molecular biology with his discovery of thealpha helix—a central structural theme of proteins—andadvancing the concept of biological specificity.
As an essay in Nature, one of the world’s pre-eminentscientific journals, opined in 2000, “Linus Pauling ranks
with Galileo, Da Vinci, Shakespeare, Newton, Bach,Faraday, Freud and Einstein as one of the great thinkers
and visionaries of the millennium. Truly he was not of thisage, but for all time.”

S P RI NG /SUM M ER 2 0 15

8

Q. What is the nature of those collaborations?
A. Dave and I co-directed the Superfund Research Center,prevalent compounds at numerous Superfund sites.
We want to bring in world leaders in environmental
chemistry who study PAHs in water, air, soil, and foodand then look at the impact of PAHs in people. The
main strength of our center is our interactions—we have
five projects and six cores that work incredibly welltogether and are centrally focused on identifying thestructural characteristics of PAHs that adversely affectand we published one paper together. Emily has a longstanding interest in trying to understand the role of zincin early development and also in susceptibility to disease.
We have an NIH grant that was recently funded tosupport this collaboration. I think our zebrafish model isideal to identify how zinc deficiency affects disease

Our focus is on PAHs
because they are one of themost prevalent compounds
at numerous superfund sites.
We want to bring in world
leaders in environmental
chemistry who study PAHs
in water, air, soil, and foodand then look at the impactof PAHs in people.

human health. Currently, most of the research in thefield is focused on less than two dozen PAHs and theirrole in cancer. Growing evidence suggests that PAHs
affect early development, learning and memory, andcardiovascular disease later in life. Kim Anderson andStacy Simonich at OSU are discovering, for the firsttime, abundant PAHs in our environment that may be
the real causes of human disease. And Dave Williams
and Rick Corley at Pacific Northwest National Labatory
are defining, for the first time, how these chemicals movethrough the human body. Together with our zebrafish
work, our center is poised to have a substantial positive
impact on human health.

and susceptibility to other environmental stressors. We
can use our model and unique infrastructure to produce
tens of thousands of zinc-deficient embryos and then ask
global questions that nobody else can answer.

Q. What are the objectives of the Superfund
Research Program?
A. It’s funded by the National Institute of Environmental

Health Sciences. I became the director at the end of
2014. The main goal is to use a perfect blend ofenvironmental and human health-focused research toidentify polycyclic aromatic hydrocarbons (PAHs) inthe environment, to characterize their toxicity, and to
determine the levels of those chemicals in the environment
below which they pose no threat to human health.

Q. What do you like to do in your free time?
A. I don’t have a lot of free time, but when I do, I spend

Q. These are sites that have a lot of industrial
contamination?
A. Yes. A Superfund site is a heavily contaminated area

it with my family—my wife Shari and daughter Lilli.
I have a huge lab and am really fortunate to have theopportunity to interact with students and scientists fromdifferent backgrounds every day—I guess that’s my mostconsistent social life. Managing this complex group andguiding career development are more fun to me thanhaving a dinner party or playing poker with the guys.

where the responsible companies are out of businessand have left it to the government to clean up. The
EPA’s goal is to try to find the resources to make thesesites clean enough for humans and other organisms.
Our focus is on PAHs because they are one of the most

LPI

9

THE LINUS PAULING INSTITUTENew Technologies Track
Human Metabolism of Polycyclic
Aromatic Hydrocarbons,an Important Class ofEnvironmental Pollutants
Erin Madeen, LPI Graduate Student

D

r. David Williams’
laboratory is applying
novel technology to track
human absorption, metabolism,and excretion of polycyclicaromatic hydrocarbons (PAHs),
universal dietary pollutants
and carcinogens. PAHs areproducts of combustion foundDavid Williams
in essentially all foods but
present at highest levels in smoked and charcoalgrilled foods, due to environmental contaminationfrom sources like burning of coal, wood, and fossil
fuels. Previous work in Williams’ and otherlaboratories has demonstrated that some of these
PAHs induce cancer in laboratory rodents throughbinding to DNA (adduction) and generation of reactive
oxygen species. The most well studied of these
PAHs is benzo(a)pyrene (BaP); 95% or more of our
exposure (non-smokers) to BaP is through our diet.

and elimination of DBC. To track the DBC that wasadministered, it was labeled with the radioactive
isotope 14C. Collaborators at LLNL utilize the Center
for Accelerator Mass Spectrometry to analyze theplasma and urine with a carbon-dating instrument(an accelerator mass spectrometer or AMS) that has
been modified to detect carbon isotopes in biologicalsamples. Because this spectrometer is so sensitive, we
can safely give very small quantities of radioactive DBC.
This sensitivity permits detection of environmentally
relevant doses of PAHs. The AMS is capable of thedetection of DBC and metabolites in blood or urine
at concentrations equivalent to one drop of water in
4,000 Olympic-size swimming pools. From thecollected data, a collaborator at PNNL and I calculated
the maximum concentration of DBC in plasma and its
elimination in urine. These results are used by risk
assessors and risk modelers who generate data usedin policy decisions related to pollutant emissions andacceptable exposure recommendations.

In December 2014, colleagues from OSU, thePacific Northwest National Laboratory (PNNL), theLawrence Livermore National Laboratory (LLNL),
and I published the first phase of these studies.
Human volunteers were given a capsule of the PAH
dibenzo(def,p)chrysene (DBC) containing a 29nanogram dose, which was chosen to represent the
dosage of PAHs that the average person consumes
daily in food. This dose was calculated to contributeno appreciable disease risk to volunteers.

Phase II of this work is focused on determining whichof several enzymes carry out the specific metabolism
of DBC. DBC and other PAHs are pro-carcinogens,meaning that they are not capable of DNA adduction or
generating reactive oxygen species without first being
modified to a reactive form by a cytochrome P450
enzyme. Two additional reactions generate the ultimate
carcinogen, a dihydrodiol-epoxide. Understanding
DBC metabolite generation will provide a better
understanding of risk from ingestion of thesecompounds. LLNL has developed a novel coupling of
ultrahigh-performance liquid chromatography (UHPLC)to AMS for detection of biological samples containing
14
C. The phase II samples are the first to be analyzed

To assess the range of human metabolism, healthy,
nonsmoking male and female volunteers were included
in a range of BMI and ages. Volunteers donated bloodand urine over four days to be analyzed for the presence

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10

with this new technology. Preliminary tests are able todetect the diol, tetrol, and DBC for calculation ofpharmacodynamics (dose-response effects in thebody) of DBC metabolism. Phase II is expected to becompleted by June 2015.
Building on the success of applying the sensitivity
and selectivity of AMS to PAH metabolism, the Williams
lab has begun recruitment of healthy nonsmoking
volunteers for a new project to analyze the metabolism
in humans of a dietarily relevant dose of another PAH,
benzo(a)pyrene (BaP), alone or in combination withsmoked salmon from the Confederated Tribe of the
Umatilla Indian Reservation in Washington. Mixtures ofvarious PAHs form on food during the traditionalsmoking process. The risk from ingestion of any PAH
mixture is estimatedby adding theconcentration
multiplied by the
carcinogenicity ofeach compound in
the mixture, dubbed
the Relative Potency
Factor, whichassumes additivityof individual PAHs in complex mixtures. The accuracyof this approach has been challenged because not all
PAHs are carcinogenic. Non-carcinogenic and weakly
carcinogenic PAH fractions in a mixture compete
for the same enzyme systems, potentially reducing
the activation of carcinogenic PAHs in the mixture.
Volunteers agree to exclude smoked meats, cheese,and charcoal-grilled meats from their diet during thestudy. Study participants ingest 46 nanograms of
BaP, labeled with 14C, for UHPLC-AMS assessment ofpharmacokinetics (absorption, metabolism, andexcretion) and pharmacodynamics of BaP. This doseis about 10-fold less than the estimated amount of BaP
consumed by the average non-smoking American every
day, so the study is being done at an environmentally
relevant dose. A subset of volunteers will be given a
serving of smoked salmon in addition to the 14C-BaP
dose. If the pharmacodynamics of 14C-BaP metabolites
are altered by the addition of the salmon, the Relative
Potency Factor approach will not be supported.
Conversely, if the detected 14C-BaP metabolites remain
the same, this approach will be supported. Results are
expected to affect calculations of risk from exposure to
PAHs and regulations concerning allowable limits of
PAH emissions. LPI

Traditional salmon smoking

Collaborators
at Lawrence
Livermore National
Laboratory utilize
the Center for
Accelerator Mass
Spectrometry toanalyze the plasma
and urine witha carbon-dating
instrument.

11

THE LINUS PAULING INSTITUTE

Diet andThursday, September 10, 2015

Optimum

Organized Walk/Run

7:30—8:30

Breakfast provided

EXTENDING HEALTHSPAN:
BASIC CONCEPTS, UNDERLYING MECHANISMS,
AND ROLE OF DIET AND LIFESTYLE

HealthSeptember 9 - 12, 2015

6:00 AMSESSION 1

Risk Factors that Limit Healthspan
Chairs: Tory Hagen, PhD and Viviana Pérez, PhD
Linus Pauling Institute, Oregon State University
8:30—9:00

Basic biology of aging, healthspan,and geroscience
Felipe Sierra, PhD
National Institute on Aging
National Institutes of Health, Bethesda, MD

9:00—9:30

Global burden of disease
Mohammad Forouzanfar, MD, PhD
University of Washington, Seattle, WA

9:30—10:00

What comparative biology can tell us
about healthspanSteven Austad, PhD
University of Alabama, Birmingham, AL

10:00—10:30

Coffee/Tea BreakCorvallis, Oregon

Wednesday, September 9, 2015
12:00 PM

Registration begins

1:45—2:00

Welcome and Opening Remarks
Balz Frei, PhD, Linus Pauling Institute,Oregon State University

Inflammation, Redox Biology, and Healthy Aging
Chairs: Fritz Gombart, PhD
and Kathy Magnusson, DVM, PhD
Linus Pauling Institute, Oregon State University

VITAMIN E: A CRITICAL NUTRIENT FOR
DEVELOPMENT AND HEALTH
Chair: Maret G. Traber, PhD
Linus Pauling Institute, Oregon State University
Vitamin E, necessary nutrient for the brain
Maret G. Traber, PhD
Linus Pauling InstituteOregon State University, Corvallis, OR		
2:30—3:00 Vitamin E deficiency: An unfolding public healthconcern in poor rural societies in South Asia
Keith West, PhD
Johns Hopkins Bloomberg School of Public Health
Baltimore, MD

10:30—11:00

Metagenomics and metabolomics incentenarians and their offspring: Two newpieces of the inflamm-aging puzzle
Claudio Franceschi, MD
University of Bologna, Italy

11:00—11:30

Chronic inflammation and cell senescenceJames Kirkland, MD, PhD
Mayo Clinic, Rochester, MN

11:30—12:00

The redox code: Toward an understandingof genome—exposome interactions andhealthy aging
Dean Jones, PhD
Emory University, Atlanta, GA

2:00—2:30

3:00—3:30

3:30—4:00

Stroke and vitamin E
Chandan Sen, PhD
The Ohio State University Medical Center
Columbus, OH

12:00—1:30 PM Lunch provided

Oral Abstracts

Vitamin E requirements in metabolic syndrome
Rich Bruno, PhD, RD
The Ohio State University, Columbus, OH

4:00—4:30 Interactions of vitamins E and C in diabetics
Mark Levine, MD
National Institutes of Health, Bethesda, MD
6:30

Chair: TBA

Welcome Reception

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12

1:30—2:50

Talks

2:50—3:15

Coffee/Tea Break

3:15—4:55

Talks

5:00—7:00

Poster Session(hors d’oeuvres and beverages provided)

CELEBRATING THE LIFE AND WORK OF GEORGE BAILEY

Friday, September 11, 2015
6:00 AM

Organized Walk/Run

7:30—8:30

Breakfast provided

SESSION 1
Chairs: Rod Dashwood, PhD
Texas A&M Health Science Center and
Tom Kensler, PhD, University of Pittsburgh

EXTENDING HEALTHSPAN: BASIC CONCEPTS,
UNDERLYING MECHANISMS, AND ROLE OF DIET
AND LIFESTYLE

1:00—1:30

George Bailey’s career at Oregon State University
Richard Scanlan, PhD
Emeritus Dean of Research
Oregon State University, Corvallis, OR

1:30—2:00

Contamination of rainbow trout diet anddevelopment of rainbow trout as a cancer model
Roger Coulombe, PhD
Utah State University, Logan, UT

2:00—2:30

Indole-3-carbinol: Its mode(s) of action asa chemopreventative agent—and why it may
cause drug interactions
David Stresser, PhD
Corning Incorporated, Woburn, MA

SESSION 2

Strategies to Maintain Cognitive Function
Chairs: Kathy Magnusson, DVM, PhD
and Fritz Gombart, PhD
Linus Pauling Institute, Oregon State University
8:30—9:00

This is your brain on aging
Kathy Magnusson, DVM, PhD
Linus Pauling InstituteOregon State University, Corvallis, OR

9:00—9:30

Calorie restriction mimetics: Promiseand pitfalls
Donald Ingram, PhD
Pennington Biomedical Research Center
Louisiana State University System
Baton Rouge, LA

2:30—3:00

Large trout studies (ED001) and application
for risk assessment
Ashok Reddy, PhD
Oregon Health & Science University, Portland, OR

3:00—3:30

Coffee/Tea Break

Nutraceutical interventions for cognitive
functions throughout life
Andrew Scholey, PhD
Swinburne University of Technology
Melbourne, Australia

CELEBRATING THE LIFE AND WORK OF GEORGE BAILEY

9:30—10:00

SESSION 2
Chairs: Sharon Krueger, PhD and Emily Ho, PhD,
Linus Pauling Institute, Oregon State University

10:00—10:30 Coffee/Tea Break

3:30—4:00

Mechanistic and preclinical studies withnatural and synthetic chlorophylls
Rod Dashwood, PhD,
Texas A&M Health Science Center, Houston, TX

4:00—4:30“Green” chemoprevention in humans:
Chlorophyllin leads the way
Tom Kensler, PhD
University of Pittsburgh, Pittsburgh, PA

4:30—5:00

Low-dose accelerator mass spectrometry
studies of carcinogens and anti-carcinogens
in human volunteers
David Williams, PhD
Linus Pauling InstituteOregon State University, Corvallis, OR

5:00—5:30

International impact of George Bailey’s
scientific body of work
John Groopman, PhD
Johns Hopkins Bloomberg School of
Public Health, Baltimore, MD

6:30

Reception

7:30

Banquet DinnerSarcopenia, Frailty, and Exercise
Chairs: Viviana Pérez, PhD and Tory Hagen, PhD
Linus Pauling Institute, Oregon State University
10:30—11:00 Cardiovascular risk and kidney function
in older adults: The role of frailty
Michelle Odden, PhD
Oregon State University, Corvallis, OR
11:00—11:30

TBA

11:30—12:00 Risk factors and preventative strategies
for accidental falls and bone fractures inolder adults
Lynn Marshall, Sc.D.
Oregon Health & Science University
Portland, OR
12:00—1:00

Lunch provided

Saturday, September 12, 2015

PUBLIC SYMPOSIUM
9:00—11:00 AM LPI’s Rx for Health: The role of diet, micronutrients,and supplements in human health and disease

13

THE LINUS PAULING INSTITUTEUndergraduate Students
in the Lab
Learning While Advancing ScienceIn addition to the faculty researchers and graduate students
who are determining the impact of supplements on healthand disease and the relevant cellular mechanisms, the Linus
Pauling Institute has many undergraduate students working
alongside them each term.

One of these students is Jackilyn Toftner, a junior who ismajoring in chemistry with a biochemistry emphasis. Toftner
decided early on a medical career in disease prevention andJackilyn Toftnertreatment and is strongly motivated to help individuals and theirfamilies stay healthy to avoid the suffering that accompanies chronic diseases.
She chose OSU because it is known for strong science and engineering programs
and opportunities for undergraduates to work in a research lab.
Last year Toftner reached out to a number of faculty at the university to see if any
were accepting undergraduate students in their laboratories. She was especiallyinterested in working with Professor Fred Stevens in the Linus Pauling Institute,“because I am very passionate and interested in the potential applications ofchemistry to other disciplines. I want to use my ability tolearn and my thirst for knowledge and lab techniques tohelp others in any way I can.”Photo by Clint ShockToftner is working on a project with Stevens to determine
the properties of fagonia, a plant native to Pakistan that isused by people in the region to treat breast cancer.

Fagonia plant

Toftner is working on a
project with Stevens todetermine the propertiesof fagonia, a plant native
to Pakistan that is usedby people in the region
to treat breast cancer.
Stevens’ lab is part of theLinus Pauling Institute’s
Cardiometabolic Disease
Prevention program.

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“In America, there has been controversy about the
medicinal properties of this plant,” explains Toftner. “Myproject is focused on extracting the bioactive components
out of a dried, ground sample of the plant to make achemical profile. The goal is to then test the bioactive
components individually on breast cancer cells to see ifthey trigger apoptosis, or cell death, in the cancer cells.”
Fred Stevens

Stevens’ lab is part of the Linus Pauling Institute’s
Cardiometabolic Disease Prevention program, which is focused on a better
understanding of the molecular and cellular mechanisms of cardiometabolic
diseases and the role of oxidative stress and inflammation. Scientists in this area also
are investigating the protective effects of micronutrients and other dietary factors or
supplements, such as vitamins C and E, lipoic acid, fish oil, and flavonoids.
In addition to Stevens, Toftner works with graduate student Elizabeth Axton.
She notes that both have been helpful with not only her lab techniques but also inbrainstorming solutions to problems that she encounters in her experiments.
“Working in a lab is a lot different from being in chemistry lab class where you learn
how to use the general applications from class. Working in a research lab not only
applies those classroom techniques to real life experiments but requires the ability tothink independently and to make adjustments to those experiments when problems
arise,” Toftner adds. “It’s a great experience for students, and even if you aren’t
considering a career in research, the practical experience is invaluable.” LPI

14

DEVELOPMENTS
A Lasting Legacy of Nutritional ScienceW

hen the Linus Pauling Institute moved from
Palo Alto, California, to Corvallis, Oregon, in 1996,
it focused on advancing knowledge in areas of interest
to Dr. Linus Pauling, with an emphasis on the role ofnutritional factors in health and disease prevention.
Almost 20 years later, the Institute has 12 principal
investigators and is nationally recognized for its workin helping us all live longer, better lives.

many, many individuals, including more than 100 who
have made gifts to support our research through theirestate plans. We are so grateful to all of you who havechosen to make a gift to the Linus Pauling Institute—
your generosity enables all that we do.
If you are interested in supporting the Linus Pauling
Institute in a way that wouldn’t affect your current
lifestyle or your family’s security, you can make theInstitute part of your estate plans by including special
language in your will or trust. If this is something you
are considering or have already done, please let us knowof your intentions so that we can better steward yourgift according to your wishes.

Researchers in the Institute are driven to understand
not only the causes of diseases, but also how we canminimize our risk through eating right and supplementing
a healthy diet with vitamins and minerals. Researchspans a number of disciplines: from the impact ofenvironmental factors like pollution and toxic chemicals,
to the ways in which our bodies absorb (or don’t, insome cases) the nutrients that we need, to determining
the beneficial effects of flavonoids in plants.

I enjoy talking with supporters of the Linus Pauling
Institute and would love to hear how you came toknow of the Institute and its work. Many of you had
a chance to meet Dr. Pauling, as well—some at Caltech
or Stanford, some through professional societies, andsome who worked with him. If you have a story about
Dr. Pauling that you’d like to share with us, please
don’t hesitate to call (541-737-0055) or email me atmarlys.amundson@oregonstate.edu.

Although the Linus Pauling Institute is located in asmall town in Oregon, the Institute’s reach is global.
The Micronutrient Information Center website gets visitorsfrom around the world, and this year we have received
gifts from 45 states and four countries outside the US.
Since its inception, the Institute’s research on vitamins
and other nutritional factors has been supported by

If you’re online, the Linus Pauling Institute’s Facebook
site is a great way to keep up with news from the Institute:
https://www.facebook.com/LinusPaulingInstitute. LPI

Marlys Amundson
Director of Development
Photo by Jim Carroll Photography

LPI is grateful for the bequests
we have received from thefollowing friends this past year:
Sylvia Arenson
Gerald D. Carney

Marie K. Frank
Elaine W. Hindin
John F. Holterhoff
Marion T. Tsefalas

See us on facebook

Visit the LPI Blog

facebook.com/linuspaulinginstituteblogs.oregonstate.edu/linuspaulinginstitute15

THE LINUS PAULING INSTITUTE

Linus Pauling Institute

Non-Profit Org.
U.S. Postage
PAID
Portland, OR
Permit No. 1006

Stephen Lawson, Research Newsletter Editor
Oregon State University
307 Linus Pauling Science Center
Corvallis, Oregon 97331
phone: 541-737-5075
fax: 541-737-5077
email: lpi@oregonstate.edu
website: lpi.oregonstate.edu

Printed on recyclable paper using soy inks

Look for these informative
articles inside!
Page 1 ....... From the Director
Page 1 ....... Fish Toxicology — An Interview with
Robert Tanguay, Ph.D.
Page 10 ......New Technologies Track Human
Metabolism of Polycyclic Aromatic
Hydrocarbons, an Important Classof Environmental Pollutants
Page 12 ......Diet and Optimum HealthConference 2015
Page 14 ......Undergraduate Students in the Lab
Page 15 ......Developments

GIVING to theLinus Pauling InstituteGifts in support of research efforts can be madeat any time. Checks should be payable to OSUFoundation for Linus Pauling Institute. Information
on giving is available through the OSU Foundation,
1-800-354-7281, or by writing to the Institute.

CREATE YOUR LEGACY
Including the Linus Pauling Institute in your willor trust is an easy way to make a big difference for
future generations.
Contact us for a free estate planning guide or for
more information on bequests or life-income gifts.
Julie Irmer • OSU Foundation, Office of Gift Planning
800-336-8217 • Gift.Planning@oregonstate.edu
osufoundation.org/giftplanning

Special thanks to Barbara McVicar for editorial assistance and photographs
and to authors of signed articles.