Genetics: RNA Interference Cuts Hepatitis Down to Size Immunology: New Mouse Gives Glimpse into Complications of Diabetes Oncology: Zebrafish Lights Path of Leukemia   Researchers Report Quick, Inexpensive HIV Test Lawsuits for Medical Monitoring May Aid Public Health Pilot HIV Screening Finds Infection 43 Percent Higher than for Self-referrals   HSPH Names New Head of Population and International Health Spaulding Names New President New Appointments to Full and Named Professorships Farmer and Spengler Win Heinz Award MyCourses Training for Faculty Nominations Sought for Invitational Awards Nominations Sought for Dean's Award Recognizing Support of Women Staff Joslin Receives Funding to Train Pediatric Endocrinologists Honors and Advances News Brief In Memoriam: Donald O'Hara   Resident Mocks Gay Physician   How a Doctor Builds a Family   Front Page

IMMUNOLOGY New Mouse Gives Glimpse into Complications of Diabetes Model Works Toward Showing Complement-Diabetes Link In the vast realm of the immune system, there is no readier tribe than the 25 or so proteins that compose the complement system. Though some complement proteins wait for signals from different immune branches before launching their attack, others roam the bloodstream in a more roughshod manner, damaging not just intruders but also the host's own tissues. To protect themselves from this undisciplined response, host cells must cloak themselves with protective proteins. Below, mice lacking cd59 exhibit a host of anomalies. The molecule normally protects red blood cells from damage by the complement proteins that make up the membrane attack complex (MAC). Lacking such protection, the red blood cells of cd59-knockout mice undergo budding (upper left in this electron micrograph). Budding is the result of the MAC on red cells and also of the MAC-induced activation of platelets. (Image courtesy of H. Mulhern) Several years ago, José Halperin and his colleagues proposed that a lack of functional protective proteins might provide a key to understanding a range of human proliferative diseases, most notably the complications of type 2 diabetes. It was a controversial idea since the complement system was not thought to play a major role in this form of diabetes. To make matters more difficult, the researchers lacked a convincing animal model to prove their thesis. Halperin, Xuebin Qin, and their colleagues report in the February Immunity that they have created a mouse model that, with a little further tinkering, could demonstrate the complement-diabetes link. "With all this emerging evidence of the importance of complement not just in diabetes but in other diseases, the development of an animal model is key." --José Halperin. If their work pans out, the researchers will have filled a longstanding and costly gap in medical research. Diabetes is a major killer of Americans and an even larger cause of morbidity due to its complications, such as retinopathy, renal disease, and atherosclerosis. Yet researchers know little about what causes cells to proliferate in the far-flung corners of a diabetic's body--eyes, limbs, kidneys. Confounding their attempts is the lack of a research model. Mice and rabbits have been bred to develop the core mechanisms of diabetes, such as high blood sugar and low insulin levels, but no nonhuman animal so far has exhibited the complications of the disease in the same combination and intensity seen in humans. Building a Better Mouse The new theory builds on previous discoveries by Halperin, associate professor of medicine at HMS, and colleagues about how complement proteins work (see Focus, May 19, 2000). Unlike T cells, which attack infected host cells, complement proteins deal their death blows to intruders directly. They do this by inserting tiny pores into the intruder's membrane, allowing salt and water to rush in and burst the cell. For years, these pores, which are formed by a handful of complement proteins called the membrane attack complex (MAC), were thought to be the equivalent of one-way doors, allowing substances in but not out. But in the mid-1980s Halperin and colleagues showed that the MAC openings were capable of expelling growth factors and cytokines which, in turn, cause nearby cells to proliferate. "This will be a very important model for all the proliferative diseases--atherosclerosis, lupus, vasculitis, and nephritis--in which complement seems to play a role. It is a model that nobody had before," said José Halperin (front). His coauthors are (l to r) Xuebin Qin, Luciano Grubissich, and Martin Dobarro. (Photo by Graham Ramsay) Normally, cells are shielded against MAC attack by protective proteins, in particular cd59. According to Halperin and his colleagues, excess sugar in the blood of diabetics binds to the cd59 protein, essentially disabling it. Deprived of these protectors, cells become vulnerable to MAC attack and hence to complement-induced growth, leading to a host of proliferative conditions like retinopathy, atherosclerosis, and renal disease. The newly engineered mice, which lack the cd59 gene, give the researchers a real shot at proving their hypothesis. The researchers have not yet been able to determine whether the cd59- knockout mice spontaneously develop the complications of diabetes or if they will need to have high levels of blood sugar to do that. But the mice did develop many symptoms associated with the human disease paroxysmal nocturnal hemoglobinauria (PNH). This disease, which has been linked to a defect in the cd59 gene, is associated with the symptoms of hemolytic anemia, which the mice exhibit. Stress Tests To explore the diabetes link, the researchers are currently exposing the knockouts to high levels of glucose. In fact, they plan to subject their cd59-deprived animals to a variety of stressors like high cholesterol to demonstrate their thesis that complement is involved in atherosclerosis and other proliferative diseases. "With all this emerging evidence of the importance of complement not just in diabetes but in other diseases, the development of an animal model is key," said Halperin. What makes the model especially precious is the trouble it took to develop it. Researchers elsewhere had tried knocking out the cd59 gene in mice and met with failure. The animals exhibited mild versions of PNH symptoms, but they were essentially normal. The reason for their lack of symptoms became clear when Qin, a research associate in cell biology at HMS, and his colleagues identified and cloned a second cd59 mouse gene. Preliminary work suggested that the protein made by this second gene was much more active than that made by the first--a finding confirmed by knocking out the second gene. "The animals have this spectacular PNH phenotype. They have all the hematologic features of the human disease," Halperin said. They also exhibit a few unexpected characteristics, most notably a progressive loss of fertility in the males. Though initially surprising, the discovery makes perfect sense. Sperm face all kinds of obstacles once they enter the female genital tract, not least of which is attack by the female immune system, including the complement proteins. Normally, sperm blanket themselves in protective proteins. "Cd59 is one of the most conspicuously expressed proteins in the sperm," said Halperin. Lacking the protective coat, the knockout sperm were more vulnerable to attack, not just by female complement proteins but by the male's own complement system. As for why the decline is progressive, one possibility is that the complement system gets stronger, and hence sperm become more vulnerable, as an animal matures. Intriguingly, the researchers found evidence in old medical textbooks that infertility used to be a common complaint of diabetics. "Infertility was one of the major problems of diabetics before the availability of insulin," said Halperin. Insulin works by sweeping sugar out of the blood system, potentially preventing it from masking cd59. But if the researchers' theory is right, there might be other ways to rescue sperm in the diabetic patient, for example, by attaching cd59 to sperm cells in vitro. In fact, focusing on cd59 could lead to more general treatments for the complications of diabetes. One approach is to transfect the bone marrow cells of such patients with mutant versions of cd59 that cannot bind sugar. Another is to introduce into the diabetic patient's bloodstream small molecules that prevent sugar from binding to cd59. Such therapeutic approaches are, for the moment, futuristic, said Halperin. "First we need to prove the concept in our model," he said. Added Qin: "These animals will give us a hint of where we need to go with regard to understanding therapies and mechanisms. But it is all just starting." --Misia Landau