Finding suggests preventive treatment for killer disease
St. Louis, May 24, 2000 -- Up to 9 percent of people over age 65 are carrying a time bomb that one day could kill them in minutes: a weak area in the aorta, the main artery coursing from the heart. When the aorta ruptures like an overinflated inner tube, it spills blood into the abdomen, halting circulation. Now, researchers have identified a key enzyme that damages the aorta wall. They also have found that a drug called doxycycline, currently used as an antibiotic, keeps the enzyme in check and helps mice avoid abdominal aortic aneurysms.
"This might turn out to be the first feasible pharmacological therapy for preventing aneurysm expansion in patients," says Robert W. Thompson, M.D., an associate professor of surgery, radiology and cell biology and physiology at Washington University School of Medicine in St. Louis. Thompson and colleagues report their findings in the June 1 issue of The Journal of Clinical Investigation (JCI).
Abdominal aortic aneurysms kill at least 15,000 Americans each year, though the number may be higher because such deaths sometimes are attributed to heart attacks. No current treatment can prevent small aneurysms from enlarging, and if a large aneurysm is discovered before it ruptures, surgery is the only option.
The aneurysms develop when elastin, a structural protein, is broken down, allowing the wall to balloon out. Scientists have long suspected that enzymes called metalloproteinases (MMPs) are responsible for degrading elastin in the aortic wall. But they haven¹t known which member of this large family to blame.
To address this question, the researchers perfused the aortas of mice with a low concentration of the enzyme elastase, initiating aortic wall injury. The elastase disappeared within 24 hours, but 21 of 23 perfused mice had abdominal aortic aneurysms (defined as at least a doubling of aortic diameter) 14 days later. The enlarged area was infiltrated by inflammatory cells, particularly macrophages. These cells appeared to be secreting several different metalloproteinases, including one called MMP-9, antibody studies showed.
The mouse model of aortic aneurysms was developed in part by Robert Pyo, M.D., a former research fellow in vascular surgery, and Jason K. Lee, M.D., a current research fellow in Thompson¹s laboratory. Pyo and Lee are the first two authors of the JCI paper.
To determine whether metalloproteinases might be involved in aneurysm development, the researchers gave doxycycline, which acts as a nonspecific MMP inhibitor, to another group of mice for 14 days after elastase perfusion. Only 50 percent of these animals developed abdominal aortic aneurysms, suggesting the involvement of an MMP.
To find out whether MMP-9 or its cousin, MMP-12, is the key player, the researchers studied mice that lacked either enzyme or both enzymes. The genetically modified animals were produced by Robert M. Senior, M.D., J. Michael Shipley, Ph.D., and Steven D. Shaprio, M.D., all faculty members at the School of Medicine.
Fourteen days after elastase perfusion, 94 percent of unaltered mice and 100 percent of MMP-12-deficient mice developed abdominal aortic aneurysms. The aortas of these animals were 2.4 times wider than normal. But aneurysms developed in only 40 percent of MMP-9-deficient mice and in only 20 percent of mice that were deficient in both MMP-9 and MMP-12. Both types of mice that lacked MMP-9 had well-preserved elastin even though inflammatory cells had infiltrated their aortas.
When MMP-9-deficient mice were irradiated (to kill their bone marrow) and transplanted with bone marrow from normal mice, their aortas became significantly larger after elastase treatment. But bone marrow from MMP-9-deficient mice did not have this effect. "These results suggest that MMP-9 produced by inflammatory cells is one of the keys to the development of abdominal aortic aneurysms," Thompson says.
He hopes these findings and the results of two pilot clinical studies will lead to a multicenter trial of doxycycline for management of small abdominal aortic aneurysms. In 1999, his group gave a one-week course of the drug to eight patients who were about to have their aneurysms surgically repaired. Analysis of aneurysm tissue removed during surgery revealed that patients who had taken doxycycline had five times less MMP-9 production in the artery wall than those who had not taken the drug. This suggests that doxycycline might be able to inhibit aneurysm enlargement in humans as well as in mice. In a more recent pilot study, Thompson and others showed that doxycycline appears to be safe and well-tolerated by patients with abdominal aortic aneurysms. "Even though this drug has been safely used as an antibiotic for more than 40 years, we thought it was important to conduct a trial in this specific patient population," Thompson says.
Meanwhile, Thompson and other vascular surgeons at Washington University are helping patients with operable aneurysms by performing less invasive surgery than the traditional procedure, which opens up the abdomen to replace the weak segment of the aortic wall. The new endoluminal stent procedure uses a catheter to insert a graft through the femoral artery from the groin. When the graft reaches the area that needs to be repaired, it is dilated and held in place with stents. Thompson took part in several clinical trials of this procedure that were led by Gregorio A. Sicard, M.D., chief of vascular surgery at the School of Medicine. The FDA approved two of the endoluminal graft devices in October 1999, and the Washington University group now has treated more than 200 patients - more than any team in the nation. "The endoluminal stent procedure has revolutionized clinical care for abdominal aneurysms," Thompson says, "because it allows patients to go home within 24 hours - instead of five to seven days - and avoids the complications of open surgery."
In the future, Thompson hopes to be able to give patients a preventive drug rather than just repair enlarged aneurysms. "We expect the development of pharmacological treatments such as doxycycline will diminish the need for surgical repair," he says, pointing out that small aneurysms already can be detected with a high degree of accuracy by noninvasive tests such as abdominal ultrasound.
Pyo R, Lee JK, Shipley JM, Curci JA, Mao D, Ziporin SJ, Ennis TL, Shapiro SD, Senior RM, Thompson RW. Targeted gene disruption of matrix metalloproteinase-9 (gelatinase B) suppresses development of experimental abdominal aortic aneurysms. The Journal of Clinical Investigation 105, 1641-1649, June 1, 2000.
This research was supported by the National Heart, Lung, and Blood Institute (NHLBI), the Parker B. Francis Fellowship Program, the Alan A. and Edith L. Wolff Charitable Trust and the Pacific Vascular Research Foundation. In 1999, the NHLBI launched a research program on abdominal aortic aneurysms by awarding grants to nine institutions, including Washington University School of Medicine in St. Louis. As part of the school¹s aneurysm research center, two of these research grants are enabling Thompson and his colleagues to continue their studies of MMPs. A third project is directed by Jay W. Heinecke, M.D., professor of medicine.
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