[Perhaps the focus should be on what a stem cell can do and not from
where it comes.]
Embryonic stem cell strategy advanced with UCSF finding
UCSF scientists are reporting what they say is a significant
improvement in the technique for genetically reprogramming mouse
cells to their embryonic state, a process that transforms the cells,
in essence, into embryonic stem cells.
The finding, published on-line as an immediate early publication
in "Cell Stem Cell" (Sept. 6, 2007), builds on the strategic
breakthrough reported by Shinya Yamanaka, MD, PhD, in 2006, and
confirmed in the spring of 2007 both by Yamanaka's team and, in
independent studies, by scientists at MIT, Harvard and UCLA.
The advance by the UCSF team should accelerate research aimed at
improving the original strategy, the team says, and increase its
potential use for studying disease development and creating patient-
specific stem-cell based therapies.
The work is the result of a collaboration between the labs of Miguel
Ramalho-Santos, PhD, and Robert Blelloch, MD, PhD, of the UCSF
Institute for Regeneration Medicine.
"The new technique removes a major technical hurdle that has likely
discouraged many labs around the world from carrying out studies on
the strategy," says senior author Ramalho-Santos, a UCSF Fellow and a
member of the Diabetes Center. For separate reasons, he says, removal
of the hurdle increases the technique's potential use in developing
patient-specific cellular therapies.
"Now, laboratories will be able to use the approach to study a broad
range of normal and diseased cells of interest," says the first
author of the study, Blelloch, an assistant professor of
urology. "There will be a much greater ability to precisely dissect
the mechanisms of reprogramming and to identify the genes that will
be most effective in transforming adult cells."
Yamanaka's strategy -- over-expressing certain genes in mouse skin
cells to initiate reprogramming – relied on the insertion of a
foreign "drug resistance" gene into the mouse skin cells. This gene
would "switch on" in those cells that successfully converted to
embryonic stem cells, thus providing a means of detecting them. The
drawbacks of this technique were that it was technically difficult to
carry out and, because it involves a foreign gene, would raise safety
concerns that would hinder its use in cell-based therapies.
In the current study, the UCSF scientists developed an alternative to
this genetically engineered "switch" technique. They developed serum-
free conditions in the cell culture dish that both promoted more
successful reprogramming and generated embryonic stem cells that
could be detected based on their form and structure, alone.
Scientists are interested in reprogramming because of its potential
for developing human embryonic stem cells that contain the genetic
makeup of individual patients. In theory, any patient's cell, say, a
skin cell, could be reprogrammed. If the resulting embryonic stem
cell could then be prompted in the culture dish to specialize into
one of the various cell types of the body, such as of the heart, lung
and brain, the resulting cells could provide the starting point for a
host of clinical-research strategies.
Researchers could create dopamine-producing cells from Parkinson's
disease patients and study them in the culture dish to learn the
earliest steps of disease development. They could also test
experimental drugs on such cells in the culture dish.
Alternatively, they could generate healthy specialized cells from
patients who had donated their genetic material, and transplant them
into tissues -- without the risk of prompting immune rejection -- to
treat failing hearts, neurological diseases such as Parkinson's
disease and amyotrophic lateral sclerosis, spinal cord injury and
diabetes.
The reprogramming strategy pioneered by Yamanaka -- who in August
began his transition from Kyoto University to the UCSF-affiliated
Gladstone Institute of Cardiovascular Disease and UCSF -- involved
over-activating four genes in mouse skin cells in the culture dish.
His team showed that over-expressing these genes – oct4, sox2, klf4
and c-myc – can cause the full complement of genes in mouse cells to
lose their adult functions and begin functioning as they would have
as embryonic stem cells. Yamanaka named these cells "induced
pluripotent (iPS) cells."
But because only a very low percentage of cells complete reversion to
the embryonic stem cell state with this technique, and because the
cells are situated among millions of cells in the culture dish that
do not complete the transformation, the scientists had a difficult
time identifying the fully reprogrammed cells. Thus, they developed
the technique of inserting the foreign "drug-resistance" gene into
the mouse skin cells. This gene was designed to only "switch on" in
cells that completed the reversion to the embryonic stem cell state.
With addition of the drug to the culture dish, the vast majority of
cells, those that had not reverted to embryonic stem cells, died.
Only those that had reverted survived and could then be expanded.
With the alternative technique developed by the UCSF team, the
efficiency of embryonic stem cell production remained low. However,
the mouse skin cells that did start to revert to embryonic stem cells
could readily be identified by their form and structure in the
absence of any drug. The researchers went on to show that these cells
indeed behaved like embryonic stem cells and could give rise to all
cell types of the body.
Separately, the team demonstrated that reprogramming could be
achieved when one of the four genes over-expressed to initiate
reprogramming -- c-myc -- was replaced with a related gene, known as
n-myc. These genes are involved in the formation of different tumors,
so by beginning to replace genes in this method the researchers may
find combinations of reprogramming genes that are safer, says
Blelloch.
"Studies should address the relative efficacy of n-myc versus c-myc
in reprogramming and whether n-myc reactivation, like c-myc, results
in tumor formation," he says.
An ongoing limitation of the Yamanaka method, notes Blelloch, is that
it requires viral-mediated integration of four foreign genes – so-
called transgenes. The goal would be to add the genes only
temporarily, or to use chemical compounds that could mimic the effect
of the genes in the cells. This will be a key focus of ongoing
studies, he says.
The biggest hurdle, of course, says Ramalho-Santos, will be
translating the methods from the mouse to human cells, a process that
could take years. Researchers around the world, including Ramalho-
Santos, Blelloch and Yamanaka, are working intently on this challenge.
"It's a very exciting time in stem cell biology, as exemplified in
the studies of reprogramming,
enough that an embryonic stem cell can give rise to all cell types of
the body. But that's what embryonic stem cells do. They grow and in
the end give rise to the whole organism.
"But taking back a differentiated cell to the embryonic stem cell
state – that's truly mesmerizing. It goes against the flow of
development -- and yet we can do it. And we're getting easier
technical ways to do it."
###
Other co-authors of the study were Monica Venere, PhD, a postdoctoral
research fellow in of the Blelloch lab and Jonathan Yen, an
undergraduate student at Johns Hopkins University and a summer
research volunteer in the UCSF Diabetes Center and Institute for
Regeneration Medicine.
The study was funded by the National Institutes of Health, the
Sandler Foundation, the Juvenile Diabetes Research Foundation, the
Pew Charitable Trust, the UCSF Department of Urology and the UCSF
Institute for Regeneration Medicine.
UCSF is a leading university that advances health worldwide by
conducting advanced biomedical research, educating graduate students
in the life sciences and health professions, and providing complex
patient care.
Related links:
UCSF Institute for Regeneration Medicine http://irm.ucsf.
Blelloch lab http://irm.ucsf.
Ramalho-Santos lab http://irm.ucsf.
Public release date: 10-Sep-2007
Contact: Jennifer O'Brien
jobrien@pubaff.
415-476-2557
University of California - San Francisco
http://www.eurekale
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StemCells subscribers may also be interested in these sites:
Children's Neurobiological Solutions
http://www.CNSfoundation.org/
Cord Blood Registry
http://www.CordBlood.com/at.cgi?a=150123
The CNS Healing Group
http://groups.yahoo.com/group/CNS_Healing
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