July 22, 2004
Researchers Buzzing Over New Wound-Healing Model
The image shows epidermal cell nuclei (green) in fruit fly larvae. The dark hole in the center of the image is a wound that is healing.
Howard Hughes Medical Institute researchers have made progress in
understanding the genetic program that is deployed to help wounds heal.
Their insights come via a somewhat surprising route - hours spent
making tiny punctures in fruit fly larvae, then observing and analyzing
how the cells recover.
The painstaking analysis reveals that two biochemical pathways are
involved in wound healing. And because the work was done in fruit
flies, which are amenable to a wide range of genetic tools, there is
hope that the new model will guide researchers to points in the
wound-healing process that can ultimately be controlled by clinicians.
With time and better understanding of how wounds are repaired, doctors
may be able to intervene more successfully in cases where wounds fail
to heal properly.
“Our hope is that understanding the molecular pathways in the fly, to the extent that they are conserved in humans, might allow the design of mechanistic and pharmacological approaches to promote or alter wound-healing to greatly improve outcomes.”
Mark A. Krasnow
The researchers, postdoctoral fellow Michael J. Galko and Howard
Hughes Medical Institute investigator Mark A. Krasnow, both at the
Stanford University School of Medicine, published their findings in the
August 2004 issue of Public Library of Science Biology.
Krasnow, whose lab is known for studies of genetic control of lung
development, is branching out because he realizes that a simplified
model of wound healing is badly needed. “Because of its clinical
importance, there has been a huge amount of work in mammalian
wound-healing. The challenge has been that the process is so
complicated in mammals,” said Krasnow. “And it's not
possible to conduct the necessary interventional experiments on humans
in a clinical setting.
“There have been attempts to study a simplified process in
cell culture, but that approach removes the process from its natural
context,” he said. “What Michael has done is develop a
genetically tractable wound-healing model that allows genetic
dissection of this complex process.”
The fruit fly Drosophila has a long history of genetic study,
so it made sense to Galko and Krasnow to consider using it as the basis
for a simplified model system. Wound-healing is likely quite similar in
flies and humans, said Krasnow, since the basic process is probably
evolutionarily ancient, with its major components in place since before
insects and mammals diverged.
To create a wound, Galko made tiny punctures through the outer
cuticle, or exoskeleton, and into the epidermis of fly larvae with a
needle about the diameter of a human eyelash. Galko also perfected a
technique to create larger wounds that do not form a scab. Without an
opaque scab obstructing their view, researchers can see changes in the
epidermal cells directly. To produce these scabless wounds, Galko
pinched the transparent larval cuticle to damage the epidermal cells
beneath the cuticle surface.
Once the researchers created the wounds, they could analyze the
genetic changes in the surrounding cells during healing. Any cellular
changes could be seen using a microscope. The researchers observed that
while puncturing the epidermal cells caused immediate bleeding, a plug
that became the basis for scab formation rapidly sealed the wound.
“We saw that the epidermal cells around the wound rapidly
began to fuse with each other and orient themselves toward the wound,
extending long, fine processes along the plug,” said Galko.
“And by eight hours after wounding, the processes from opposite
ends of the wound contacted one another and reestablished continuity of
the epidermis.”
To study the genetic control of the process, the researchers
analyzed the activation of a signaling pathway involving the enzyme Jun
N-terminal kinase (JNK), which is activated by cellular stress, and has
been associated with the wound-healing process. The researchers found
that the pathway was dramatically activated around the fly wounds
during healing. Inactivating the pathway inhibited epidermal spreading
and epithelial restoration, but did not affect scab formation,
indicating that the two processes are separately controlled.
In contrast, when the researchers inactivated another gene, called
lozenge, they blocked scab formation but the epidermal cells
still tried to spread and close the hole in the epidermis. To their
surprise, the researchers found that without a scab, the JNK pathway
became hyperactive. This showed that scab formation and epithelial
reestablishment, although under separate genetic control, can influence
each other.
“Wound healing isn't like a bucket brigade putting out a
fire,” said Krasnow. “It's more like a modern fire
department with different firefighters responding to and containing
different parts of the damage, each calling out to let the others know
how their task is progressing and if they need help.”
According to Krasnow, the distinct but interdependent genetic
responses are evidence of coordination by multiple signals emanating
from the wound site. Their findings set the stage for the
identification and characterization of those signals and the cellular
processes that they control. Further studies, Krasnow said, will
involve searching for mutations in fly genes that affect wound healing
and tracing the genes involved.
Krasnow said that the new insights into wound healing could have
important clinical implications. “Many diabetics suffer such
problems as foot ulcers, in which wounds fail to heal. And in some
cases of trauma, exaggerated wound-healing responses can create
disfiguring scars. Thus, it's extremely important to learn to speed up
wound-healing or change it to a scarless process. However, attempting
to develop such treatments in the absence of a basic cellular and
mechanistic understanding of the process is a huge gamble, and it's
difficult to imagine that it could be successful.
“Our hope is that understanding the molecular pathways in the
fly, to the extent that they are conserved in humans, might allow the
design of mechanistic and pharmacological approaches to promote or
alter wound-healing to greatly improve outcomes,” said
Krasnow.
Image: Krasnow Lab/HHMI at Stanford University School of Medicine
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