Many Paths, Few Destinations: How Stem Cells Decide What They'll Become

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Many Paths, Few Destinations: How Stem Cells Decide What They'll

Become

http://medicalnewscenter.com/out/out.cgi?

http://www.sciencedaily.com/releases/2008/05/080521131552.htm

How does a stem cell decide what specialized identity to adopt -- or

simply to remain a stem cell? A new study suggests that the

conventional view, which assumes that cells are " instructed " to

progress along prescribed signaling pathways, is too simplistic.

Instead, it supports the idea that cells differentiate through the

collective behavior of multiple genes in a network that ultimately

leads to just a few endpoints -- just as a marble on a hilltop can

travel a nearly infinite number of downward paths, only to arrive in

the same valley.

The findings, published in the May 22 issue of Nature, give a glimpse

into how that collective behavior works, and show that cell

populations maintain a built-in variability that nature can harness

for change under the right conditions. The findings also help explain

why the process of differentiating stem cells into specific lineages

in the laboratory has been highly inefficient.

Led by Sui Huang, MD, PhD, a Visiting Associate Professor in the

Children's Hospital Boston Vascular Biology Program (now also on the

faculty of the University of Calgary), and Hannah Chang, an MD/PhD

student in Children's Vascular Biology Program, the researchers

examined how blood stem cells " decide " to become white blood cell

progenitors or red blood cell progenitors.

They began by examining populations of seemingly identical blood stem

cells, and found that a cell marker of " stemness, " a protein called

Sca-1, was actually present in highly variable amounts from cell to

cell -- in fact, they found a 1,000-fold range. One might think that

low Sca-1 cells are simply those cells that have spontaneously

differentiated. However, when Huang and Chang divided the cells

expressing low, medium and high levels of Sca-1 and cultured them,

each descendent cell population recapitulated the same broad range of

Sca-1 levels over nine days or more, regardless of what levels they

started with.

" We then asked, are these cells also biologically different? " says

Huang, the paper's senior author. " And it turned out they were

dramatically different in differentiation. "

Blood stem cells with low levels of Sca-1 differentiated into red

blood cell progenitors seven times more often than cells high in Sca-

1 when exposed to erythropoietin, a growth factor that promotes red

blood cell production. Conversely, when stem cells were exposed to

granulocyte--macrophage colony-stimulating factor, which stimulates

white blood cell formation, those that were highest in Sca-1 were the

most likely to become white cells. Yet, in both experiments, all

three groups of cells retained characteristics of stem cells.

Huang and Chang then looked at the proteins GATA1 and PU.1,

transcription factors that normally favor differentiation into red

and white blood cells, respectively. Blood stem cells that were low

in Sca-1 (and most prone to become red blood cells) had much more

GATA1 than did the high- and medium-Sca-1 cells. Stem cells high in

Sca-1 (and least prone to become red blood cells) had the highest

levels of PU.1.

But most important, the differences in Sca-1, GATA1 and PU.1 levels

across the three cell groups became less pronounced over time, as did

the variability in the cells' propensity to differentiate, suggesting

that the differences are transient.

In a final step, Huang and Chang used microarrays to look at the

cells' entire genome. Again, they found tremendous variability within

the apparently uniform cell population: more than 3,900 genes were

differentially expressed (turned " on " or " off " ) between the low- and

high-Sca-1 cells. And again, this variability was dynamic: the

differences diminished over time, with gene activity in both the low-

and high-Sca-1 cells becoming more like that in the middle group.

Together, the findings make the case that a slow fluctuation or

cycling of gene activity tends to maintain cells in a stable state,

while also priming them to differentiate when conditions are right.

" Even if cells are officially genetically identical and belong to the

same clone, individual members of that population are quite different

at any given time, " says Huang. " This heterogeneity has usually been

seen as random 'measurement noise,' and, more recently, as 'gene

expression noise.' But it turns out to be very important, and is the

basis for stem cells' multipotency -- their ability to differentiate

into multiple lineages. "

" Nature has created an incredibly elegant and simple way of creating

variability, and maintaining it at a steady level, enabling cells to

respond to changes in their environment in a systematic, controlled

way, " adds Chang, first author on the paper.

Practically speaking, the work suggests that stem cell biologists may

need to change their approach to differentiating stem cells in the

laboratory for therapeutic applications.

" So far the process has been highly inefficient -- only 10 to 50

percent of cells respond to the hormone or whatever is given to make

them differentiate, " Huang says. " That is because of the cells'

inherent heterogeneity. People have been finding more and more

sophisticated stimulator cocktails, but we could make the process

more efficient by harnessing the heterogeneity and identifying cells

that are already highly poised to differentiate. "

Chang has already done follow-up experiments showing that stem cell

differentiation can be made dramatically more efficient by choosing

the right subpopulation of stem cells and stimulating them promptly,

while they are most apt to differentiate. " I'm not doing anything

complicated -- just using what nature already has, " she says.

But the findings also challenge biologists to change how they think

about biological processes. The work supports the idea of biological

systems moving toward a stable " attractor state, " a concept borrowed

from physics. In this case, blood stem cells tend to remain blood

stem cells, yet they experience inherent fluctuations in gene

activity and protein production that can sometimes be enough to tip

the balance and cause them to fall into other attractor states --

namely, red or white blood cell progenitors. Specific growth factors

can tip the balance, but these factors are part of an overall

landscape that guides cells toward different destinies. A marble

going downhill will eventually end up in a valley, but which valley

it falls into depends on the shape of the landscape.

" Growth or differentiation factors merely increases the probability

that a cell will grow or differentiate, " says Ingber, MD, PhD,

a co-author on the paper who, with Huang, served as Chang's mentor on

the project. " Cell differentiation is an ensemble property, a

collective behavior, inherent in the system's architecture and set of

regulatory interactions. "

A previous study by Huang established, for the first time, that a

given cell can exhibit a very different pattern of gene activity from

its neighbor, taking a very different path through the landscape, yet

end up in the same valley. He and his colleagues exposed precursor

cells to two completely different drugs (DMSO and retinoic acid) and

closely monitored the cells' gene expression. Both groups of cells

eventually differentiated to become neutrophils (a type of white

blood cell), but the molecular paths they took and their patterns of

gene expression were completely different until day seven, when they

finally converged.

The landscape analogy and collective " decision-making " are concepts

unfamiliar to biologists, who have tended to focus on single genes

acting in linear pathways. This made the work initially difficult to

publish, notes Huang. " It's hard for biologists to move from thinking

about single pathways to thinking about a landscape, which is the

mathematical manifestation of the entirety of all the possible

pathways, " he says. " A single pathway is not a good way to understand

a whole process. Our goal has been to understand the driving force

behind it. "

This study was funded by the Air Force Office of Scientific Research,

the National Institutes of Health, the Presidential Scholarship, the

Ashford Fellowship of Harvard University, and the Army Research

Office.