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Although scientists understand what occurs during many of the 80
individual chemical reactions involved in blood clotting, many
questions about the dynamics of the entire reaction network remain.
Rustem Ismagilov, Associate Professor in Chemistry at the University
of Chicago, and graduate students Christian Kastrup, Matthew Runyon
and Feng Shen have now developed a technique that will enable
scientists to understand the rules governing complex biological
reaction networks. They will detail their technique in the online
early edition of the Oct. 16-20 issue of the Proceedings of the
National Academy of Sciences.
Life and death literally depend on a finely tuned blood-clotting
system. "Clotting has to occur at the right place at the right time,"
Ismagilov said. "A strong, rapid clotting response is essential to
stop bleeding at a wound, but such a clotting response at the wrong
spot can block blood vessels and can be life-threatening."
In the past, scientists have typically examined the blood-clotting
network using flasks containing homogenous mixtures-the test fluids
were the same throughout. But the contents of the circulatory system
are not homogeneous, said Kastrup, a Ph.D. student in chemistry and
the PNAS article's lead author. One of the great virtues of
microfluidics technology is its ability to control complex reactions
at critical times and locations.
"The blood-clotting system contains both fluids and surfaces in an
elaborate spatial environment, where localization of chemicals is very
important," he said. Microfluidic technology can address this issue
through its ability to control complex reactions at critical times and
locations.
In previous work, the Ismagilov group designed a simple laboratory
model to simulate blood clotting. In this model, Ph.D. student Runyon
and his associates devised three modules that correspond to the three
major stages of clotting: production of chemicals that activate
clotting, the inhibition of these activators, and formation of the
solid clot.
In this model, the scientists used only one chemical reaction in each
module instead of the 20-to-30 biochemical reactions that the modules
represent. Surprisingly, this simple model adequately reproduced many
features of blood clotting.
"There's a long history in chemistry of using simple models to
understand more complex behavior," Kastrup said. "Instead of looking
at hundreds of equations for blood clotting, we reduced it down to
three main equations. From these equations we were able to describe a
lot of the dynamics of clotting."
The ability of microfluidics to mimic the flow and geometry of human
blood vessels also proved critical.
"We had to use microfluidics to do all of this because that's how we
controlled where everything is," Ismagilov said of Runyon's previous
work. "It turned out that we got appropriate behavior only if we used
geometry similar to those observed in our vascular system. If we
changed the geometry to something that didn't look like a biological
system, the chemical system couldn't function. So geometry and flow
were very important."
In the latest advance, Kastrup used Runyon's model to see if he could
predict when clotting would occur in human blood. The team predicted
and verified that clotting occurs only at locations of vascular damage
larger than a critical size. "Surprisingly, this simple model made
correct, quantitative predictions about blood clotting," Kastrup said.
Furthermore, the model provided new details about the dynamics of
clotting. A big question in blood-clotting studies is the role of a
protein called tissue factor. Can tissue factor exist in blood without
the presence of clotting?
"From our experiments we see that it's not the overall concentration
of tissue factor that matters, but it's the localization of it that
makes a difference," Kastrup explained. That means a high
concentration of tissue factor at one location will result in clotting,
while the same number of molecules spread farther apart will not.
In the future, chemists might now be able to apply microfluidics to
the study of other complex reaction networks that control various
biological functions. And in the medical arena, the technique could
become a way to perform rapid and detailed diagnostic tests. "We'd
love to see that happen," Kastrup said.
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