UMaine scientists use zebrafish to study the building blocks of
innate disease resistance that could one day benefit human health
About the Photo:
Carol Kim's interest in applying molecular virology and microbiology
to benefit the biomedical field and aquaculture industry brought her
to UMaine in 1998, where she set up the state's first and largest
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Like their mammalian namesakes, zebrafish are striped for camouflage
protection. When moving as a herd or a school, the stripes of the
zebra and zebrafish make it difficult for predators to focus on a
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In the room saturated in white light
and the sound of rushing water, it's 82 degrees. In rows of shoebox-size
tanks stacked five high dart slivers of silver — tiny fish no bigger
than grains of rice, adults two inches long and all ages in between.
Even if it's been years since your last
childhood trip to the store to bring home a plastic bag bulging with
water containing an aquarium fish, the sight of more than 40,000
zebrafish in a laboratory at the University of Maine can still make your
heart skip a beat. It all comes flooding back: the excitement and wonder
of watching zebras native to India's Ganges River swimming just inches
from your face.
Today, what's even more remarkable
about this hardy, popular home aquarium fish is the splash it's making
in basic science. Zebrafish as model organisms are now comparable in
importance to mice and fruit flies in the study of genetics and disease
prevention. Evidence of the research potential of the little striped
vertebrates is in the prevalence of zebrafish labs at medical schools
nationwide and the explosion of the resulting scientific literature
Zebrafish are being studied to better
understand such human conditions as congenital disease, cancer and
aging. At UMaine, the zebrafish facility of Carol Kim, associate
professor of biochemistry, microbiology and molecular biology, is a hub
of activity, facilitating the initiatives of as many as five campus
scientists conducting research in such areas as microbiology,
toxicology, immunology, developmental biology and genetics. In her
research on innate immunity and infectious diseases, Kim collaborates
with scientists across the country and abroad.
Zebrafish hold the promise of very
basic scientific and applied research breakthroughs for Kim, a faculty
member in the Department of Biochemistry, Microbiology and Molecular
Biology, with affiliations with the Functional Genomics graduate program
offered by UMaine, Jackson Laboratory and Maine Medical Center Research
Institute, part of the Graduate School of Biomedical Science. In her
research, she is studying the biological factors that supplement and
prolong the body's immune response to infection. Working on the
molecular level, she is studying how cells respond to infection,
contributing basic knowledge that could one day lead to disease
prevention or more effective vaccines in humans, and in other mammals
and fish species.
"We're using the zebrafish as a model
for the immune response to infectious disease," Kim says. "The zebrafish
is a powerful model system that will allow us to better understand the
immune system and implement preventative measures against infection for
humans, as well as fish."
Zebrafish make ideal model organisms in
science for many of the same reasons they are popular in home aquariums.
They are easy to care for and breed, and they are resilient, tolerating
fluctuations in water temperatures. A female can lay up to 300 eggs each
week. Zebras can live for up to five years.
For researchers, two of the most
important characteristics of zebrafish are their rapid and viewable
development, and their biological traits that mimic those of humans.
Zebrafish eggs are transparent. Under a microscope, scientists can watch
the embryonic growth that occurs in two to four days following
fertilization. Development is so rapid that a single cell multiples to
take on a fish shape within 24 hours.
Zebrafish can serve as models for human
developmental biology, neurobiology, toxicology and genetic disease.
While they are lower vertebrates, their genes, developmental processes,
anatomy, physiology and behaviors bear similarities to those of humans,
according to ZFIN, the Web-based Zebrafish Information Network of the
Zebrafish International Resource Center at the University of Oregon.
"There are some differences in the
zebrafish model; namely, that it is not a mammal," says Kim. "In
UMaine's partnership with Jackson Lab (the world's largest mammalian
genetic research facility, where the mouse is studied as a model for
human disease), our zebrafish facility is complementary. I think that
the zebrafish model system soon will rival the mouse system as more
reagents, antibodies and cell lines are developed. It already rivals the
mouse in developmental biology and toxicology."
In recent years, scientists like Kim
have turned to zebrafish as models for the study of immunity, and
infectious viral and bacterial disease. Key to Kim's work is the
identification of the genes and molecular processes involved in innate
immunity — the natural ability of multicelled organisms to ward off
pathogens. Her studies of infectious disease in zebrafish bridge the
biomedical and applied application fields because they have the
potential to lead to a better understanding of disease development,
resistance, diagnosis and treatment in other vertebrates, including
humans and fish.
Kim's interest in applying molecular virology and microbiology to
benefit the biomedical field and aquaculture industry brought her to
UMaine in 1998, where she set up the state's first and now largest
zebrafish facility. Up to that point, much of the genetically based
research involving zebrafish focused on developmental biology and
neurobiology. Kim was among the first to use the zebrafish to study
What started as a small laboratory with
250 brood stock has grown to a climate-controlled facility in the new
wing of Hitchner Hall, housing more than 40,000 zebrafish at all stages
Kim's focus is on the role of toll-like
receptor (TLR) signal pathways that are key to innate immunity. First
identified in the fruit fly, TLRs are proteins found on the surface of
certain cells. The receptors act as defense mechanisms, recognizing and
binding with molecules of bacteria or viruses, and signaling the cell
nucleus of the invading microbial infection. The result is an innate
immune response — the release of infection-fighting molecules, such an
Such innate response is a primitive
physiological feature still shared among insects and vertebrates like
the mouse, zebrafish and human. Unlike adaptive immunity that depends on
virus- or bacteria-specific antibodies or vaccines, innate immunity as
the body's first line of defense provides an immediate, vigorous,
nonspecific inflammatory response to pathogens. Indeed, the stronger the
innate immune response in the mouse, zebrafish or human, the more
vigorous the adapted immune response.
In their efforts to better understand
how the immune system responds to viral infection, Kim and UMaine
researchers Stephen Altmann, Mark Mellon and Daniel Distel had a major
breakthrough in 2003. The scientists isolated and confirmed the function
of a zebrafish gene that produces interferon, an infection-fighting
protein known for its ability to inhibit the growth of virus.
The UMaine researchers were the first
to document the presence of interferon in any fish species. According to
the Web of Science, the report of their discovery in the Journal of
Virology is among the top 1 percent of papers cited by other scientists
in that field in the last two years.
Since then, Kim has been cloning in
zebrafish the immunogene known as Mx, which is activated by interferon.
First discovered in mice with an inborn resistance to influenza virus,
Mx bears a 50 percent resemblance to antiviral Mx proteins in humans.
The important diagnostic tool in assessing interferon activity has been
cloned in a variety of mammal, bird and fish species, but not in
zebrafish until research was completed by a team of scientists from
UMaine, Cornell and Boston's Brigham and Women's and Children's
In an effort to understand
disease-fighting responses in humans, more immune-related genes in the
zebrafish need to be identified. Earlier this year, Kim and another
research team focusing on the Mx and interferon proteins were the first
to describe how an experimental infection of snakehead rhabdovirus
developed and elicited an antiviral response in zebrafish. They focused
on the symptoms of disease and the immune response in zebrafish embryos
Targeted gene disruptions can be used
in conjunction with pathogen challenge to alter immunity to infection,
according to the research team of scientists from UMaine, the University
of Hamburg in Germany and Dalhousie University in Canada, writing in the
February 2005 issue of the Journal of Virology. Differences in mortality
rates, pathogenesis and gene expression may provide clues about the role
of genes linked to immunity.
Kim is now collaborating with Nick Trede at the Huntsman Cancer Institute at the University of Utah to
establish a transgenic (genetically modified) line of zebrafish that
would have fluorescence to indicate activation of the TLR signaling
pathways. When viewed using a special microscope, the mutant fish and
their embryos have the potential to show scientists how the different
proteins along the pathway function when the organism is compromised by
Researchers also hope to identify genes
that could improve or exaggerate the response of the TLR pathway.
"To identify what genes are responsible
for such changes could mean that one day, we can identify humans with —
or who are more susceptible to — disease," Kim says. "Using animal
models, we're hoping to mimic the abnormality."
That approach is at the heart of Kim's
most recent research project, funded by a more than $405,000 grant from
the National Institutes of Health (NIH). She is collaborating with
Dartmouth Cystic Fibrosis Research Development Program researchers to
develop a zebrafish model for studying cystic fibrosis.
According to NIH's National Human
Genome Research Institute, cystic fibrosis is the most common, fatal
genetic disease in the United States. About 30,000 people in the U.S.
have the disease, which is caused by a single mutated gene — the Cystic
Fibrosis Transmembrane Regulator (CFTR).
In normal cells, the CFTR protein
serves as a channel, allowing cells to release chloride as part of the
immune response system. However, in people with cystic fibrosis, the
protein is defective and the cells do not release the chloride,
resulting in an improper salt balance and production of thick mucus.
In her lab, Kim will experimentally
infect zebrafish with bacterial strains from cystic fibrosis patients in
an effort to better understand why they are so pathogenic. The number of
bacterial strains and the many CFTR gene mutations (more than 900,
according to the National Human Genome Research Institute) make the
microbiology portion of the project statistically strong. It is basic
science that will be arduous in its compilation and analysis, but
valuable. The bacteria preferentially affects cystic fibrosis patients
with chronic infection that causes chronic inflammation.
"We're hoping to determine some of the
key factors of the innate immune response that contribute to the
detrimental inflammatory response seen in cystic fibrosis patients," Kim
says. "If we can establish a way to control inflammation, cystic
fibrosis patients will have a better outcome."
By Margaret Nagle
for more stories from this issue of UMaine Today Magazine.