EurekAlert
Public release date: 27-Mar-2003
Contact: Bob Kuska
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301-594-7560
NIH/National Institute of Dental and Craniofacial Research
Scientists report important data in stem cell debate

Developmental biologists have long maintained that adult stem cells
cannot be reprogrammed. Once a stem cell commits to a specific
tissue, such as the brain, it can't turn back its biological clock
and become blood, bone, or any other type of adult stem cell. But,
about four years ago, this fundamental idea received a serious jolt
when scientists reported inducing a small percentage of adult stem
cells from the brain to switch into cells from the blood. This led to
a series of confirmatory findings that seemed to have settled the
question, when two teams of researchers last year dropped the small
bombshell that, in their laboratory studies, some stem cells
physically fused with other cells. This finding suggested that maybe
the brain-to-blood and other stem cell switches, a process that
scientists call transdifferentiation, weren't what they seemed. Maybe
the stem cells had fused with cells from other tissues all along.
Now, in the current issue of The Lancet, a team of scientists at the
National Institutes of Health offers a key piece of new evidence to
advance the debate. In a study of five women who had received bone
marrow transplants from their brothers several years earlier, the
team reports finding cheek cells that contained the male Y
chromosome, a sign that some transplanted stem cells had
differentiated into cheek cells. Moreover, the group found almost no
evidence of fusion among the cells in the cheek.

Because the researchers analyzed cells extracted directly from
patients, which previous studies did not for various technical
reasons, these data offer strong evidence that transdifferentiation
does occur. The study also shows the power of using oral tissues to
pursue complex biological questions, an idea that is gaining wider
favor among biologists.

"Being so accessible, the mouth is one of the best 'laboratories' in
the body to study many issues in human biology that go beyond dental
research," said Dr. Bruce Baum, a scientist at the National Institute
of Dental and Craniofacial Research (NIDCR) and an author on the
study. "This is clearly an excellent case in point."

While the pros and cons of transdifferentiation may sound of interest
to academicians only, the issue potentially has profound public
health implications. If transdifferentiation is a biological reality,
scientists would have a potential inroad to therapeutically
manipulate adult stem cells, the long-lived "progenitor" cells that
produce the myriad specialized cells in our tissues.

In theory, scientists could gather the most easily obtainable adult
stem cells, such as those from the blood, switch them into another
type of adult stem cell, then prompt them to produce large amounts of
tissue-specific cells. This harvest of specialized cells could be
transplanted to heal wounds more efficiently or even possibly to
construct replacement tissues, such as a salivary gland or a tooth.

As appealing as this scenario sounds, scientists continue to grapple
with the question: How does one conclusively prove that
transdifferentiation is possible in adult stem cells? After all, the
current data suggest that the phenomenon occurs in only a small
percentage of the cells, raising the additional question of whether
transdifferentation would be robust enough for therapeutic purposes.

Leaders in the field recently established scientific criteria of
proof to guide subsequent experiments. Yet, even with these
established criteria as their guide, scientists have struggled with
several technical difficulties. For one, much of the work, pro and
con, has occurred in cell culture. That is, cells have been extracted
from the body and placed into an artificial medium, where they behave
differently than in their natural environment, a little like trying
to study a fish out of water.

Secondly, the other studies involved cells from tissue sections,
which literally provide scientists with only part of the picture.
"What happens with other tissues, such as the liver or muscle, is you
must slice the tissue into sections to obtain samples," said Dr.
Simon Tran, an NIDCR scientist and the lead author on the study.
"This means you must overlay parts of the cell to reconstruct them or
just analyze a portion of the cell."

Tran said he and his colleagues at NIDCR, in particular Dr. Stanley
Pillemer, reasoned that the mucosal cells of the cheek, which help to
form the moist tissue that lines the inside of the mouth, might solve
these technical problems. He said the cells replicate frequently,
meaning a ready supply exists, and they can be collected non
invasively from patients. Just as importantly, because the cheek
cells haven't been sliced in two during processing, scientists can
analyze the entire cell under a microscope.

To test this idea, Tran said he found, through his collaborators at
NIH's National Heart, Lung, and Blood Institute, five women who had
received bone-marrow transplants several years earlier from their
brothers. Tran gathered the cheek cells from each of the women, then
returned to his laboratory to confront two basic questions: Could he
identify cheek cells that contained both an X and Y chromosome, an
indication that the transplanted bone-marrow stem cells had
differentiated into cheek cells? If so, could he also show that these
cells were indeed functioning as cheek cells?

Tran said that's where he ran into technical difficulties of his own.
He needed to create a two-in-one assay that could detect both the Y
chromosome and the structural protein cytokeratin, a standard
identifier of mucosal cells, such as those from the cheek. "It's been
done before," said Tran, "but never on cheek cells."

Tran said he turned for help first to Dr. Eva Mezey, who studies
adult stem cells at NIH's National Institute of Neurological
Disorders and Stroke, and later to Dr. Amalia Dutra, a researcher at
NIH's National Human Genome Research Institute, and Dr. Michael
Brownstein, a scientist at NIH's National Institute of Mental Health.
"I would have never been able to do this study if I had been in
another type of setting," said Tran, highlighting the unique
multidisciplinary environment at NIH that allows more complex studies
to be undertaken.

Once the assay was up and running, the scientists discovered that all
five women had cheek cells that contained both X and Y chromosomes.
The range was from 0.8 percent in one woman to 12.7 percent in
another. Because some of the women had sons, the scientists performed
an additional DNA analysis that ruled out the possibility of the
cells originating from their male offspring. The cells also tested
positive for cytokeratin.

Interestingly, of the 9,700 cells that were examined in the study,
only two showed signs of possible fusion. In the previous reports of
cell fusion from celll culture studies, the rate was also extremely
low, ranging from one every 100,000 to one million adult stem cells.

While Tran said this paper does not provide the definitive answer to
the transdifferentiation debate, it does offer a higher level of
evidence. "I hadn't studied stem cells previously," said Tran, who is
employing tissue engineering techniques to develop an artificial
salivary gland. "But I saw an opportunity to help forward the field
with a model that perhaps some had overlooked. I think these data
will be helpful to many."

###

SOURCE: EurekAlert
http://www.eurekalert.org/pub_releases/2003-03/niod-sri032403.php

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