August 26, 2005
Researchers Devise New Technique for Creating Human Stem Cells
The image shows human embryonic stem cells stained for a characteristic marker protein (Tra1-60, red dots). When adult skin cells are fused with embryonic stem cells, the hybrid cells (not shown) re-express this marker, suggesting that the cells have reverted to the embryonic state.
Researchers have developed a new technique for creating human
embryonic stem cells by fusing adult somatic cells with embryonic stem
cells. The fusion causes the adult cells to undergo genetic
reprogramming, which results in cells that have the developmental
characteristics of human embryonic stem cells. The new technique may
permit scientists to derive new human embryonic stem cell lines without
the need to use human embryos.
“The long term goal for this experiment was to do cell fusion in a way that would allow the elimination of the embryonic stem cell nucleus to create an embryonic stem cell from the somatic cell.”
Douglas A. Melton
This approach could become an alternative to somatic cell nuclear
transfer (SCNT), a method that is currently used to produce human stem
cells. SCNT involves transferring the nuclei of adult cells, called
somatic cells, into oocytes in which scientists have removed the
nuclei.
The researchers said that — while the technique might one day be
used along with SCNT, which involves the use of unfertilized human eggs
— technical hurdles must be cleared before the new technique sees
widespread use. It is more likely that the new technique will see
immediate use in helping to accelerate understanding of how embryonic
cells “reprogram” somatic cells to an embryonic state.
The researchers published their findings in the August 26, 2005,
issue of the journal
Science
. Senior author Kevin Eggan and
Howard Hughes Medical Institute investigator Douglas A. Melton, both at
Harvard University, led the research team, which also included Harvard
colleagues Chad Cowan and Jocelyn Atienza.
In theory, researchers can induce embryonic stem cells to mature
into a variety of specialized cells. For that reason, many researchers
believe stem cells offer promise for creating populations of
specialized cells that can be used to rejuvenate organs, such as the
pancreas or heart, that are damaged by disease or trauma. Stem cells
also provide a model system in which researchers can study the causes
of genetic disease and the basis of embryonic development.
Eggan, Melton and their colleagues decided to pursue their
alternative route after other researchers had shown that genetic
reprogramming can occur when mouse somatic cells are fused to mouse
embryonic stem cells. The scientists knew that if their studies were
successful, it would provide the research community with a new option
for producing reprogrammed cells using embryonic stem cells, which are
more plentiful and easier to obtain than unfertilized human eggs.
In the studies published in
Science
, the researchers combined
human fibroblast cells with human embryonic stem cells in the presence
of a detergent-like substance that caused the two cell types to fuse.
The researchers demonstrated that they had achieved fusion of the two
cell types by searching the fused cells for two distinctive genetic
markers present in the somatic fibroblast and stem cells. The
researchers were also able to further confirm that fusion occurred by
studying the chromosomal makeup of the fused cells. Their analyses
showed that the hybrid cells were “tetraploid” - meaning
they contained the combined chromosomes of both the somatic cells and
the embryonic stem cells.
One of the key findings from the study was that the fusion cells
have the characteristics of human embryonic stem cells. “Our
assays showed that the hybrid cells, unlike adult cells, showed the
development potential of embryonic stem cells,” said Eggan.
“We found they could be induced to mature into nerve cells, hair
follicles, muscle cells and gut endoderm cells. And, since these cell
types are derived from three different parts of the embryo, this really
demonstrated the ability of these cells to give rise to a variety of
different cell types.”
Furthermore, Eggan noted that genetic analyses of the fused cells
revealed that the somatic cell genes characteristic of adult cells had
all been switched off, while those characteristic of embryonic cells
had been switched on. “With the exception of a few genes one way
or the other — which is perhaps because these cells are now tetraploid
— the hybrid cells are indistinguishable from human embryonic stem
cells,” he said.
“The long term goal for this experiment was to do cell fusion
in a way that would allow the elimination of the embryonic stem cell
nucleus to create an embryonic stem cell from the somatic cell,”
said Melton. “This paper reports only the first step toward that
goal, because we end up with a tetraploid cell. So, while this does not
obviate the need for human oocytes, it demonstrates that this general
approach of cell fusion is an interesting one that should be further
explored.”
The researchers also performed fusion experiments using pelvic bone
cells as the somatic cells and a different human embryonic cell line,
to demonstrate that their technique was not restricted to one adult
cell type or embryonic cell line.
In both cases, the researchers observed extensive reprogramming of
the somatic cells. “We were surprised at how complete the
reprogramming was,” said Eggan. “I think we were expecting
that there would be more 'memory' of the adult state than the embryonic
in the hybrid cells. It was quite clear that when we looked at these
hybrid cells, they had completely reverted to an embryonic
state.”
Melton said that the remaining technical hurdle is figuring out a
way to eliminate the embryonic stem cell nucleus in the hybrid cell,
causing it to have a normal number of chromosomes. One problem, said
Melton, is that the nucleus in stem cells is large, occupying nearly
the entire cell. Thus, it is not practical to physically extract the
nucleus, as is currently done with oocytes, which have a relatively
small nucleus. An alternative approach of destroying the embryonic stem
cell nucleus with chemicals or radiation would induce the cell's
suicide program, called apoptosis, he said.
Melton emphasized that “at this stage in our understanding,
the hard fact is that the only way to create an embryonic stem cell
from a somatic cell is by nuclear transfer into oocytes. Taking
advantage of this current capability — such as colleagues in South
Korea and other countries are doing — is critical if we are to
maintain the progress necessary to realize the extraordinary clinical
potential of this technology.”
Eggan added that the most realistic current promise of the fusion
technique is in studying the machinery of genetic reprogramming of
somatic cells by embryonic cells. “It is extremely difficult to
study the reprogramming process using eggs, because in the case of
humans it is very difficult to obtain eggs in any quantity and
difficult or impossible to genetically manipulate them,” he said.
“But embryonic stem cells can be grown in large quantities. We
can isolate the components of the reprogramming machinery, and we can
genetically manipulate the cells to analyze the reprogramming
process.”
Image: Chad A. Cowan, HHMI at Harvard University, for Science.
|
Jennifer Michalowski
