July 18, 2002
Researchers Produce Motor Neurons from Embryonic Stem Cells
Beginning with cultured mouse embryonic stem cells, researchers have
administered a precise mix of chemical signals to coax the cells to
differentiate into functioning motor neurons.
The achievement was made possible by a decade of work in deciphering
the signals that trigger differentiation of motor neurons, which are
responsible for controlling the movement of muscles. The experiments
represent an important step in applying that knowledge to grow
functioning neurons from stem cells — undifferentiated cells that have
the potential to become many different types of adult cells.
“This is just the starting point for trying to take a rational approach to studying the ability of ES-cell-derived motor neurons to restore function.”
Thomas M. Jessell
According to the researchers, the success of the experiments with
mouse cells suggests that the same type of approach might be used to
grow human motor neurons from stem cells. Such neurons could enable
regeneration of nerve tissue lost to disease or trauma.
The experiments by researchers led by Howard Hughes Medical
Institute investigator Thomas
Jessell at Columbia University were reported in an article that was
given immediate early publication status by the journal Cell and
published online on July 17, 2002. The research was funded in part by
Project A.L.S.
For more than 15 years, Jessell and his colleagues have been
attempting to untangle the delicate connections of nerve cells in the
developing spinal cord. Their studies have shown that the fledgling
vertebrate nervous system is crackling with activity — genes are being
turned on and off at a rapid pace, transforming immature cells into a
billions-strong network of specialized neural cells. Ultimately,
Jessell hopes that his research will provide a more thorough
understanding of how the central nervous system (CNS) is constructed —
this, he says, may suggest new ways to repair diseased or damaged
components of the mature CNS.
According to Jessell, the attempt to generate motor neurons from
stem cells relied on many years of research to identify the chemical
cues in the developing embryo that coax “naïve
neuroprogenitor” cells to differentiate into spinal cord motor
neurons. These chemical signals “direct” ES cells down a
developmental pathway in successive stages —first into neurons and
ultimately into ever more specialized spinal cord motor neurons. Two of
the key signals, said Jessell, are retinoic acid, which work with the
group of Thomas Edlund had shown converts mid-brain neural cells into
spinal cord progenitors, and Sonic hedgehog, a protein that converts
spinal cord progenitor cells into motor neurons.
“Until now, we have been trying to piece together these steps
as individual bites,” said Jessell. “But we hadn’t
shown that the normal signaling factors could be used conjointly to
take a naïve class of progenitors like ES cells, and by sequential
exposure to these factors, recapitulate this developmental
pathway.”
First author Hynek Wichterle, a postdoctoral fellow in
Jessell’s laboratory, began by using retinoic acid and other
chemical cues to induce ES cells to differentiate into mid-brain-type
neurons and then into spinal cord neurons. The scientists could follow
the steps of differentiation by looking for the expression of specific
transcription factors that define the identity of cells as spinal cord
progenitor cells.
“Retinoids will give you spinal cord identity, but that
doesn’t determine exactly which type of neuron emerges from these
spinal cord progenitor cells,” said Jessell. “And
that’s where Sonic hedgehog becomes important, because our work
and that of others had shown that you need Sonic hedgehog signaling at
exactly the right level of signal activation to generate motor
neurons.”
Thus, when the scientists exposed the cultured spinal cord neurons
to appropriate levels of the Sonic hedgehog protein, the cells
differentiated to become motor neurons. The dependence of this
differentiation on a narrow concentration range of the Sonic hedgehog
protein is significant, said Jessell, because in developing embryos the
amount of Sonic hedgehog governs what type of neuron will be
generated.
In additional experiments, the scientists used ES cells from
transgenic mice whose motor neurons were tagged with a fluorescent
marker. The fluorescent tagging enabled Jessell and his colleagues to
monitor, isolate and purify the specific motor neurons they had induced
— a technique that Jessell believes will be crucial to further
attempts to define the signaling pathways involved in neuronal
differentiation.
The researchers were also able to address an important question,
namely, whether the motor neurons they had developed in culture could
actually function in living animals. “We needed to demonstrate
how well these in vitro-generated motor neurons did when they
were put into a living embryo,” Jessell said. “So, Hynek
managed the very impressive technical feat of reintroducing these
ES-cell-derived motor neurons back into the spinal cords of chick
embryos at a stage when normal motor neurons are being
generated.” The scientists then tested in the chick embryos how
well the introduced neurons survived, integrated themselves into the
embryonic spinal cord and extended their long cable-like axons toward
their normal targets in muscle.
“I think our results documented that these ES-cell-derived
motor neurons do a pretty good job of mimicking their embryo-derived
counterparts in all of those three tests,” said Jessell.
“In general, I was pleasantly surprised by how well neuralized ES
cells recapitulate the developmental events that we have come to
associate with motor neuron progenitors and motor neurons.”
Jessell believes that these successes represent only the beginning
of a promising line of research. “This is just the starting point
for trying to take a rational approach to studying the ability of
ES-cell-derived motor neurons to restore function, not just in an
embryonic context, but in a more relevant adult context,” he
said.
Jessell and his colleagues hope to use ES-generated motor neurons in
experiments to identify all the genes that govern the pathways of motor
neuron differentiation. They are also developing collaborations with
neurologists to explore in mouse models whether their motor neurons can
regenerate spinal cords that have been damaged by trauma or
neurodegenerative diseases, such as amyotrophic lateral sclerosis.
The researchers also plan to explore whether the signaling pathways
of motor neuron differentiation mice resemble those in humans. “I
think one can be cautiously optimistic that such parallels will
exist,” said Jessell. “While many scientists have shown
that human ES cells can give rise to neurons, we don’t know
exactly which type of neurons they are. And, there’s a much
greater heterogeneity in the properties of human ES cells than in mouse
ES cells; so it may be necessary to sift through a number of the
available human ES cell lines before arriving at a cell which behaves
as its mouse counterpart. But, in principle there is no reason why this
type of approach might not be successful with human cells,” he
said.
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