Giving Stroke Its Kick
Authors: Davenport R. John 
Published: August 2003

Like a hornet sting that turns a trusty steed into a bucking bronco, a small molecule incites a normally benign enzyme to cause mayhem in the brain, according to new research. The study suggests how nitric oxide (NO) contributes to damage from stroke or neurodegenerative disease and presents a possible new therapeutic target.
NO regulates a variety of normal brain processes, but it also appears to exacerbate damage associated with stroke or Alzheimer's disease. Mice that lack nitric oxide synthase (NOS), an NO-producing enzyme, suffer less destruction from stroke than normal animals do, for instance, although no one knows why. Previous work suggests that the molecule can alter the function of proteins by attaching to them in a process called S-nitrosylation. Furthermore, enzymes called matrix metalloproteinases (MMPs) contain an amino acid sequence that NO commonly decorates. MMPs chew up the protein network that entwines cells in tissues; when unchecked, the enzymes disrupt this foundation. Such destruction pushes cells to commit suicide.
In new research, Gu and colleagues establish the first connection between NO and MMPs in brain injury. To inflict stroke in mice, they tied off an artery that supplies blood to the rodents' brains. Then they looked for brain areas with active enzyme by using a compound that fluoresces after MMP clips it in two. Regions starved for blood contained more active MMPs than did regions with uninterrupted blood flow--but only in animals that carry NO synthase. The results suggest that NO is necessary for turbocharging MMP after stroke.
The researchers undertook additional studies to find out whether NO latches directly onto MMPs. In a test tube, NO attaches to a particular cysteine on MMPs, they found. Such modified enzymes more robustly snipped a test molecule than did bare MMPs, suggesting that NO activates MMPs. Further experiments extended this work to intact animals: MMPs from brains of rats that had suffered stroke weighed slightly more than expected--by exactly the mass of an oxidized derivative of NO. Moreover, the extra heft didn't appear in brains of animals treated with an NOS inhibitor. Together the results imply that NO activates MMPs by attaching to them. A final experiment suggests that NO activation contributes to cell death: When the scientists doused cultured neurons with NO-labeled MMPs, almost 25% of the cells showed molecular characteristics of cell suicide; when neurons also received a chemical that blocks MMP activity, only 5% exhibited such signs, similar to unexposed neurons.
"Nobody had ever pinned down the targets by which NO kills neurons and certainly not whether S-nitrosylation was involved," says neurologist Solomon Snyder of Johns Hopkins University in Baltimore. "MMPs look like a very reasonable candidate." Neuroscientist Michael Moskowitz of Harvard University adds that the research might reveal ways of minimizing the risk of bleeding that accompanies current stroke therapies. Designed to break up clots, these drugs also boost MMP activity, which could weaken blood vessels; reducing NO production or quelling MMP directly might prove more fruitful in keeping the reins on stroke damage.
--R. John Davenport

Z. Gu, M. Kaul, B. Yan, S. J. Kridel, J. Cui, A. Strongin, J. W. Smith, R. C. Liddington, S. A. Lipton, S-nitrosylation of matrix metalloproteinases: Signaling pathway to neuronal cell death.
Science
297
, 1186-1190 (2002).
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