Scientists studying the brains of mice have discovered how the toxic protein that destroys the brain cells of Alzheimer's patients enters the brain. When the researchers gave mice a drug that blocked the process, flow of the protein into the brain was virtually halted and existing accumulations of it in the brain plummeted by more than 70 percent. The results of the research will be published in the July 1 issue of Nature Medicine.
The new findings center on amyloid beta, a tiny protein molecule that accumulates over time to form tell-tale plaques in the brain tissue of Alzheimer's patients. While various cells within the brain itself produce amyloid beta, that amount may be just the tip of the iceberg. Mounting evidence suggests that the bulk of amyloid beta is produced in cells throughout the body and gets circulated in the blood. The new study reveals for the first time how the protein gets from the blood into the brain, thwarting the brain's elaborate filtration mechanism that normally keeps away toxins. In mice that had been genetically engineered to develop Alzheimer's, the process ran wild, pouring amyloid beta into the brain at eight times the rate of healthy mice.
"For more than a decade we've known that this protein wreaks havoc in the brains of Alzheimer's patients, but we haven't known how it gets there or how to prevent it from getting there. This study answers both of those basic questions, and opens an entirely new avenue for the treatment of Alzheimer's disease," said lead author Berislav Zlokovic, M.D., Ph.D., of the University of Rochester Medical Center.
Since 1992, Zlokovic's research has focused on how amyloid beta protein in the blood is able to pass through the blood-brain barrier, a thin layer of cells that lines the inner walls of the brain's blood vessels. The blood-brain barrier blocks the passage of toxins while allowing the flow of oxygen, sugar, and other nutrients to brain cells. In the current study, Zlokovic and his colleagues found that amyloid beta protein molecules cannot flow through the blood-brain barrier unaided. Rather, they get through by riding piggyback on a much larger molecule, called RAGE, which is nontoxic and moves unfettered across the blood-brain barrier. Normally, RAGE is produced in small amounts by the cells that form the blood-brain barrier. But in mice that were genetically engineered to develop Alzheimer's disease, Zlokovic found that RAGE was produced in huge amounts – eight times normal – and ferried an avalanche of amyloid beta into the brain.
Zlokovic became interested in RAGE several years ago while studying amyloid beta proteins that had been culled from the brains of Alzheimer's patients who, years before their deaths, had agreed to donate their brains to the University's Alzheimer's research program. In the donated brain tissue, Zlokovic spotted something he hadn't seen before: Some of the amyloid beta proteins were attached to much larger molecules. The larger molecules turned out to be RAGE (short for receptor for advanced glycation end products), which had been observed in brain tissue by other researchers, but whose function was a mystery. Zlokovic's new study was conceived as a series of experiments designed to solve it.
In the first experiment, he injected mice with an agent, called anti-RAGE, that binds to RAGE molecules and disables them, making them unable to bind with amyloid beta proteins. Then he injected amyloid beta proteins into the mice and observed that, with the RAGE molecules disabled, transport of the amyloid beta proteins across the blood-brain barrier came to a sudden halt.
In another experiment, he administered a RAGE look-alike, called soluble RAGE. The soluble RAGE molecules attached themselves to amyloid beta proteins, but unlike RAGE, were unable to transport the proteins across the blood-brain barrier. With the amyloid beta proteins bound to the soluble RAGE molecules, however, the RAGE molecules naturally present in the mice were not able to bind to the proteins. In this experiment, like the first, the flow of amyloid beta across the blood-brain barrier also stopped abruptly.
With evidence that RAGE was involved in the transport of amyloid beta across the blood-brain barrier, the research team obtained mice in which the gene that produces the RAGE molecule was disabled or "knocked out" by genetic engineering. When the researchers injected amyloid beta protein into these mice, none of it crossed the blood-brain barrier, confirming RAGE's function.
Another experiment was conducted in mice that were genetically engineered to develop Alzheimer's disease. In half of the mice the team administered soluble RAGE once a day; the remaining mice were not given the agent. After three months, the researchers compared the amyloid beta levels in the brains of each group. In the group that had received soluble RAGE, both the amount of amyloid beta protein in their brains and the size of amyloid plaques were reduced by 70 percent compared to the mice that did not receive the agent.
The researchers also found that as RAGE transported increased levels of amyloid beta into the brain, blood flow within the brain was reduced by half. When soluble RAGE was administered to block the process, blood flow to the brain returned to normal. The new finding – that blood flow to the brain is sharply reduced in Alzheimer's mice – suggests that decreased cerebral blood flow may partially account for the confusion and dementia that plague Alzheimer's patients.
"The experiments in this study revealed a great deal of new information about Alzheimer's disease," said Zlokovic. "First, it is now very clear that the body regulates the movement of amyloid beta proteins across the blood-brain barrier. Second, we've shown that that we can use a drug to stop the flow of amyloid beta from the blood to the brain. Finally, we learned that when we block the flow of amyloid beta over time, the brain substantially rids itself of amyloid beta and the amyloid plaques shrink dramatically.
"For patients with Alzheimer's disease, these findings suggest that we can develop a new class of drugs that work by blocking the flow of the toxic Alzheimer's protein into the brain," said Zlokovic.
The researchers are looking ahead to clinical trials of new drug candidates. They are planning studies to determine whether soluble RAGE is likely to be safe when administered to people, and are working to identify other molecules that work in the same way as soluble RAGE.
Collaborators in the study, which was funded by a grant from the National Institutes of Health, included Mark Kindy, Ph.D., from the University of South Carolina and David Stern, Ph.D. and Shi Du Yan, Ph.D. from Columbia University.
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