Category: Mad Cow Disease

Scientists at the National Institutes of Health (NIH) have discovered that an important cellular quality control mechanism may actually be toxic to some brain cells during prion infection. The research, published by Cell Press in the September 16th issue of the journal Developmental Cell, proposes a new general mechanism of cellular dysfunction that can contribute to the devastating and widespread neuronal death characteristic of slowly progressing neurodegenerative diseases.

Prions cause a number of untreatable and fatal neurodegenerative disorders, including bovine spongiform encephalopathy ("mad cow disease") in cattle and Creutzfeldt-Jakob disease in humans. "We know that abnormal metabolism of a normal prion protein (PrP) is at the root of these diseases. However, the pathways that lead to selective neuronal death are unknown," explains senior author Dr. Ramanujan S. Hegde from the National Institute of Child Health and Human Development in Bethesda, Maryland.

The endoplasmic reticulum (ER) is a membrane-bound subcompartment of the cell that helps fold newly-made proteins and route them to their final destinations within or outside the cell. When protein folding or trafficking is temporarily compromised, the ER experiences "stress" and compensates using a specific ER stress response.

Previous work demonstrated that part of the ER stress response is to re-route the trafficking of many proteins, including PrP. Instead of being transported into the ER, these proteins are sent to the cytosol to be destroyed. The phenomenon, termed pre-emptive quality control (pQC), protects cells in the short term by reducing the protein burden on the ER during times of compromised function. "Whether such re-routing of PrP for long time periods might contribute to neurodegenerative phenotypes in prion disease has been unclear," says Dr. Hegde.

Dr. Hegde and colleagues designed a series of experiments to investigate a potential pathway linking prion infection, ER stress, pQC and neurodegeneration. The researchers found that prion infection induced ER stress, and consequently reduced transport of PrP into the ER. They then engineered transgenic mice to express a form of PrP that isn't efficiently transported into the ER; this approach mimics what happens to PrP during ER stress. In fact, the re-routed PrP caused mild neurodegeneration, even in the absence of prion infection or ER stress.

The results establish a previously unappreciated link between ER stress, pQC and PrP-induced neuronal damage, showing that an ordinarily helpful quality control pathway can be detrimental over long periods of time. "We believe that one mechanism of prion-mediated neurodegeneration might involve an indirect and surprisingly subtle effect on PrP biosynthesis and metabolism," concludes Dr. Hegde. He is quick to note that the neurodegeneration caused by pQC of PrP may very well be the lesser of two evils. "The consequences of not re-routing PrP for degradation during ER stress might be even worse for neurons."

Hegde says that his lab is now investigating why PrP exposed to the cytosol via this pathway is harmful: "Our working hypothesis is that PrP in the wrong part of the cell makes inappropriate interactions with other proteins to compromise their function."

The researchers include Neena S. Rane, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD; Sang-Wook Kang, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD; Oishee Chakrabarti, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD; Lionel Feigenbaum, National Cancer Institute, Frederick, MD; and Ramanujan S. Hegde, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD.

 Blood transfusions are a valuable treatment mechanism in modern medicine, but can come with the risk of donor disease transmission. Researchers are continually studying the biology of blood products to understand how certain diseases are transmitted in an effort to reduce this risk during blood transfusions.

According to a study in sheep prepublished online in Blood, the official journal of the American Society of Hematology, the risk of transmitting bovine spongiform encephalopathy (BSE, commonly known as "mad cow disease") by blood transfusion is surprisingly high.

BSE is one of a group of rare neurodegenerative disorders called transmissible spongiform encephalopathies (TSEs), and there is no reliable non-invasive test for detecting infection before the onset of clinical disease. In addition to BSE, these diseases include scrapie, a closely related disease in sheep, and Creutzfeld-Jakob disease (CJD) in humans, which causes neurological symptoms such as unsteadiness and involuntary movements that develop as the illness progresses, rendering late-stage sufferers completely immobile at the time of death.

A new variant of CJD (termed vCJD) was recognized in the United Kingdom in the mid-1990s, apparently as a result of the transmission of BSE to humans. Because the symptoms of this disease can take many years to appear, it was not known how many people might have been infected, and without a reliable test for identifying these individuals, clinicians were very concerned that the infection could be transmitted between people by blood transfusion or contaminated surgical and dental instruments.

As a result, costly control measures were introduced as a precautionary measure to reduce the risk of disease transmission, although at the time it was unclear whether there really was a significant risk or whether the control measures would be effective. This sheep study sought to better understand how readily TSEs could be transmitted by blood transfusion in order to help develop more targeted controls.

"It is vitally important that we better understand the mechanisms of disease transmission during blood transfusions so we can develop the most effective control measures and minimize human-to-human infections," said Dr. Fiona Houston, now a Faculty of Veterinary Medicine, University of Glasgow, UK, and lead author of the study.

The nine-year study conducted at the University of Edinburgh compared rates of disease transmission by examining blood transfusions from sheep infected with BSE or scrapie; the BSE donors were experimentally infected, while the scrapie donors had naturally acquired the disease. While scrapie is not thought to transmit to humans, it was included as an infection acquired under field conditions, which could possibly give different results than those obtained from experimentally infected animals. Because of the similarity in size of sheep and humans, the team was able to collect and transfuse volumes of blood equivalent to those taken from human blood donors.

The outcome of the experiment showed that both BSE and scrapie could be effectively transmitted between sheep by blood transfusion. Importantly, the team noted that transmission could occur when blood was collected from donors before they developed signs of disease, but was more likely when they were in the later stages of infection. Of the 22 sheep who received infected blood from the BSE donor group, five showed signs of TSEs and three others showed evidence of infection without clinical signs, yielding an overall transmission rate of 36 percent. Of the 21 infected scrapie recipients, nine developed clinical scrapie, yielding an overall transmission rate of 43 percent.

Investigators noted that the results were consistent with what is known about the four recorded cases of vCJD acquired by blood transfusion in humans. In addition to the stage of infection in the donor, factors such as genetic variation in disease susceptibility and the blood component transfused may influence the transmission rate by transfusion in both sheep and humans.

"The study shows that, for sheep infected with BSE or scrapie, transmission rates via blood transfusion can be high, particularly when donors are in the later stages of infection. This suggests that blood transfusion represents an efficient route of transmission for these diseases," said Dr. Houston. "Since the results are consistent with what we know about human transmission, the work helps justify the control measures put in place to safeguard human blood supplies. It also shows that blood from BSE- and scrapie-infected sheep could be used effectively in non-human experiments to answer important questions, such as which blood components are most heavily infected, and to develop much-needed diagnostic tests."

 Researchers from the University of Pennsylvania School of Medicine have shown, in unprecedented detail, how a small molecule is able to selectively take apart abnormally folded protein fibers connected to Alzheimer's disease and prion diseases. The findings appear online in the Proceedings of the National Academy of Sciences. Finding a way to dismantle misfolded proteins has implications for new treatments for a host of neurodegenerative diseases.

Abnormal accumulation of amyloid fibers and other misfolded forms in the brain cause neurodegenerative diseases. Similarly, build-up of abnormally folded prion proteins between neurons causes the human version of mad cow disease, Creutzfeldt-Jakob disease.

"Surprisingly, a small molecule called DAPH selectively targets the areas that hold fibers together, and converts fibers to a form that is unable to grow. Normally fibers grow from their ends, but the drug stops this activity," says senior author James Shorter, PhD, Assistant Professor of Biochemistry and Biophysics. "Our data suggest that it is possible to generate effective small molecules that can attack amyloid fibers, which are associated with so many devastating diseases."

The researchers are now working on how DAPH acts as a wedge to stop the fibers from growing. "Presumably DAPH fits very neatly into the crevices between fiber subunits," explains Shorter. "When we grow yeast cells with the prion in the presence of DAPH, they begin to lose the prion. We also saw this in the test tube using pure fibers. The small molecule directly remodels fiber architecture. We've really been able to get at the mechanism by which DAPH, or any small molecule, works for the first time." DAPH was originally found in a screen of small molecules that reduce amyloid-beta toxicity in the lab of co-author Vernon Ingram, Shorter's collaborator at the Massachusetts Institute of Technology (MIT).

In a test tube, if a small amount of amyloid or prion fiber is added to the normal form of the protein, it converts it to the fiber form. But when DPAH is added to the mix, the yeast prion protein does not aggregate into fibers. "It's essentially stopping fiber formation in its tracks," says Huan Wang, first author and research specialist in Shorter's lab. "We were surprised to see two very different proteins, amyloid-beta and Sup35, sensitive to this same small molecule."

The next step is to identify more potent DAPH variants with greater selectivity for deleterious amyloids. Since some amyloids may turn out to be beneficial — for example, one form may be involved in long-term memory formation — it will be necessary to find a drug that does not hit all amyloids indiscriminately. "We'd need one that hits only problem amyloids, and DAPH gives us a hint that such selectivity is possible" says Shorter.

This work was initiated in Susan Lindquist's lab at MIT and completed at Penn. The study was funded by the National Institute of General Medical Sciences, the Alzheimer's Association, the Kurt and Johanna Immerwahr Fund for Alzheimer Research, a DuPont-MIT alliance, the American Heart Association, and pilot grants from the University of Pennsylvania Alzheimer's Disease Core Center and Institute on Aging.

For the first time, researchers have peered deeply at the atomic level into the protein that causes hereditary cerebral amyloid angiopathy (CAA) — a disease thought to cause stroke and dementia. The study pinpointed a tiny portion of the protein molecule that is key to the formation of plaques in blood vessels in the brain.

Researchers worldwide are working to understand how certain kinds of proteins, called prions, cause degenerative brain diseases such as CAA. More common prion diseases include bovine spongiform encephalopathy (mad cow disease), and Creutzfeldt-Jakob disease in humans. All are incurable and fatal.

Ohio State University chemist Christopher Jaroniec and his colleagues report their results in the online edition of the Proceedings of the National Academy of Sciences.

Jaroniec understands that any discovery related to prions could raise people’s hopes for a cure, but he emphasized that his study is only a first step towards understanding the structure of the prion for CAA. “This is a very basic study of the structure of the protein, and hopefully it will give other researchers the information they need to perform further studies, and improve our understanding of CAA,” he said.

His team partnered with biochemists from Case Western Reserve University, who took a fragment of the human prion protein for CAA and tagged it with chemical markers.

Jaroniec explained that, while the prion protein used in the study is associated with the development of hereditary CAA, it is not infectious.

After the researchers tagged the molecule, they created the right chemical conditions for it to fold into macromolecules called amyloid fibrils.

Researchers know that in the body, these fibrils form plaques that lodge in blood vessel walls in the brain. But nobody has been able to closely examine the molecular structure of CAA fibrils until now.

“These fibrils are very large and complex, and so traditional biochemical techniques won’t reveal their structure in detail,” Jaroniec said.

The assistant professor of chemistry at Ohio State is an expert in a technique that can reveal the structure of such large molecules: solid-state nuclear magnetic resonance (NMR) spectroscopy.

NMR works by tuning into the radio waves emitted by atoms within materials. Every atom emits radio waves at a particular frequency, depending on the types of atoms that surround it.

The NMR technique the chemists used, called “magic angle spinning,” involves spinning materials at a certain angle with respect to the NMR's magnetic field in order to remove radio interference among the atoms. It offers researchers a clear view of which atoms make up a particular molecule, and how those atoms are arranged.

So after the researchers let the prion proteins fold into amyloid fibrils, they used magic angle spinning NMR to study the fibrils’ structure.

They searched the NMR signals for the chemical tags that had been planted in the prions. In that way, they were able to determine what parts of the original prion protein were contained within the fibrils.

They found, to their surprise, that some 80 percent of the original prion protein molecule was not present in the fibrils. The fibrils consisted exclusively of the remaining 20 percent — approximately 29 amino acids, of which two appear to be especially critical to the structure of the molecule.

Other studies have suggested that these two amino acids, numbered 138 and 139, were key to the formation of the CAA fibrils, Jaroniec said. But this study is the first to confirm their importance by studying them at the atomic level.

The researchers are continuing this work, and plan to examine the structure of the fibrils in more detail, as well as other strains of the CAA prion protein.

Jaroniec’s partners on this project included Jonathan Helmus and Philippe Naudaud, both doctoral students at Ohio State, and their collaborators at Case Western.

This research was funded by Ohio State University and the National Institutes of Health.

Prion infection of neurons increases the free cholesterol content in cell membranes. A new study suggests that disturbances in membrane cholesterol may be the mechanism by which prions cause neurodegeneration and could point to a role for cholesterol in other neurodegenerative diseases.

It is widely believed that prions (protein only infectious material) are the cause of rare progressive neurodegenerative disorders that affect both humans and animals. A prion is an infectious agent made solely of protein. However what is not known is how the prions damage brain cells (neurons).

Dr Clive Bate and colleagues from the Royal Veterinary College in the UK compared the amounts of protein and cholesterol in prion-infected neuronal cell lines and primary cortical neurons with uninfected controls. Protein levels were similar but the amount of total cholesterol (a mixture of free and esterified cholesterol) was significantly higher in the infected cell lines.

The cholesterol balance was also affected: the amount of free cholesterol increased but that of cholesterol esters reduced, suggesting that prion infection affects cholesterol regulation. The team attempted to reproduce the effects of prions on cholesterol levels, by stimulating cholesterol biosynthesis or by adding exogenous cholesterol. Both approaches resulted in increased amounts of cholesterol esters but not of free cholesterol.

The free cholesterol is thought to affect the function of the cell membranes and to lead to abnormal activation of phospholipase A2, an enzyme implicated in the depletion of neurons in prion and Alzheimer's disease.

Studies have recently shown that the controlling cholesterol levels within the brain is critical in limiting the development of neurodegenerative diseases such as Alzheimer's, Parkinson's and prion diseases, multiple sclerosis, and senile dementia. This study now gives far more specific insight into the kind of mechanisms at work. Dr Bate stated: "Our observations raise the possibility that disturbances in membrane cholesterol induced by prions are major triggering events in the neuropathogenesis of prion diseases".

Journal reference: Sequestration of free cholesterol in cell membranes by prions correlates with cytoplasmic phospholipase A2 activation. Clive Bate, Mourad Tayebi and Alun Williams. BMC Biology (in press).

Amyloid plaques, the hallmark of Alzheimer's disease, are clumps of fiber-like misfolded proteins which many experts think cause this devastating neurodegenerative disease.

While effective treatment remains an elusive goal, new research by University of Illinois at Chicago chemists suggests a possible new approach.

Yoshitaka Ishii, associate professor of chemistry, and his students managed to capture and characterize a crucial intermediate step in the formation of amyloid plaque fibers, or fibrils, showing tiny spheres averaging 20 nanometers in diameter assembling into sheet-like structures comparable to that seen in formation of fibrils.

Fibrils made of small proteins called amyloid-beta are toxic to nerve cells, but intermediate spheres, including those identified by Ishii's group, are more than 10 times as poisonous. That has made the spherical intermediates a new suspect for causing Alzheimer's disease.

"The problem with studying the structure of this intermediate form is that it's so unstable," said Ishii. His team's approach, he said, was to 'freeze-trap' the fleeting intermediate form, then use solid-state nuclear magnetic resonance to determine its structure and electron microscopes to study its morphology, or shape.

Ishii and his coworkers confirmed that the intermediate spherical stage of amyloid is more toxic than the final-form fibrils. Their findings are the first to pinpoint sheet formation at the toxic intermediate stage in the misfolding of the Alzheimer's amyloid protein and support the notion that the process of forming the layered sheet structure might be what triggers toxicity and kills nerve cells.

"Our method characterized the detailed molecular structure of this unstable, intermediate species," Ishii said. "To the best of our knowledge, this is the first characterization of detailed molecular structures for toxic amyloid intermediates. We found that the structure was very similar to the final (fibril) form, which wasn't expected at all."

Ishii said a complete determination of the intermediate structure remains to be done, but he is confident his lab will be able to do that. Once completed, the findings may provide pharmaceutical manufacturers with the information they need to create drugs that will prevent interaction between the toxic molecules and nerve cells.

Ishii said the method can also be applied to structural studies of proteins associate with other neurodegenerative diseases, including Parkinson's, and prion diseases, such as Creutzfeldt-Jakob.

"We're also interested in applying our technique in the nanoscience field to examine the formation process of peptide-based nano-assemblies," he said.

The findings were reported online December 2 in Nature Structural & Molecular Biology.

UIC students co-authoring the paper include former doctoral student Sandra Chimon, candidates Medhat Shaibat, Christopher Jones and Buzulagu Aizezi, and former undergraduate Diana Calero.

An effective and sensitive new method for detecting and characterizing prions, the infectious compounds behind diseases like mad cow disease, is now being launched by researchers at Linköping University in Sweden, among other institutions.

Mad cow disease (BSE), which has caused the death of more than 200,000 cattle and 165 people in the U.K., has now abated. But other prion disorders are on the rise, and there is concern that new strains will infect humans. Prions are not readily transmittable from species to species, but once they have broken through the species barrier they can rapidly adapt and become contagious within the species. Intensive work is now underway to find new, more sensitive methods for detecting these potentially deadly protein structures and distinguish between various strains.

The method now being presented in the journal Nature Methods is based on a fluorescent molecule, a so-called conjugated polymer, which was developed at Linköping University.

The research team infected genetically identical laboratory mice with BSE, scrapie (which afflicts sheep), and CWD (chronic wasting disease or “mad elk disease,” which is epidemic in the central U.S.) for several generations in a row. Gradually new strains of prions emerge, making the diseases more fatal to the mice. Tissue samples from mice were examined using the fluorescent molecule, which seeks out and binds with prions. This is signaled by a shift in color. By tweaking the molecule, the team has been able to get it to show different colors depending on the structure of the prion­each prion strain emits its own optical fingerprint.

This is an important difference compared with other techniques used to find prions, such as antibodies and the well-known stain Congo red.

The technique has also proven to work well on tissue sections from dead animals, such as cows infected with BSE. Now the scientists want to move on and look for alternative sample-taking methods for diagnosing and tracking prion diseases in humans in early stages.

The method would then be useful for screening blood products, since there is a risk that people can be carriers of prions without having any symptoms of disease. In the U.K. it was discovered that 66 people had received blood from blood donors who were infected with the human form of BSE (a variant of Creutzfeldt-Jakob’s disease, vCJD), and among them, four individuals have been shown to be infected (source: Health Protection Agency, Jan. 2007).

“Using our methods, we can directly see the structure of the prions and thereby deduce the disease,” says Peter Nilsson, one of the lead authors of the article. Nilsson developed the technique as a doctoral student at Linköping University and now, as a post-doctoral fellow with Professor Adrian Aguzzi’s research team in Zürich, has been applying the technology to prion diseases. After New Year’s he will assume a post-doc position at Linköping.

“For us researchers it is truly exciting to use this technique to understand more about both prions and other defectively folded proteins that give rise to similar disorders, such as Alzheimer’s,” says Peter Hammarström, co-author and research director of the prion laboratory at Linköping.

Another co-author is Kurt Wüthrich, the 2002 Nobel laureate in chemistry.

The article “Prion strain discrimination using luminescent conjugated polymers” by Christina J Sigurdson, K Peter R Nilsson, Simone Hornemann, Guiseppe Manco, Magdalini Polymenidou, Petra Schwartz, Mario Leclerc, Per Hammarström, Kurt Wüthrich, and Adriano Aguzzi was published in Nature Methods online on November 18 and will appear in the December issue of the printed journal.

Diseases that cause neurons to break-down, such as Alzheimer's, Multiple Sclerosis and Creutzfeldt-Jakob disease (Mad Cow Disease), continue to be elusive to scientists and resistant to treatments.

A new finding from University of Michigan researchers demonstrates an unpredicted link between a virtually unknown signaling molecule and neuron health.

In a study in PNAS, graduate student, Yanling Zhang, postdoctoral fellow Sergey Zolov and Life Sciences Institute professor Lois Weisman connect the loss of this molecule to massive neurodegeneration in the brain.

The molecule PI(3,5)P2 is a lipid found in all cells at very low levels. Lipids are a group of small organic compounds. While the best studied lipids are fats, waxes and oils, PI3,5P2 is a member of a unique class of lipids that signal the cell to perform special tasks.

Weisman said it was surprising to find that PI(3,5)P2 plays a key role in the survival of nervous system cells.

"In mice, lowered levels of PI(3,5)P2 leads to profound neurodegeneration," said Weisman. "It suggests that we have a good place to look to find treatments for neurodegenerative diseases such as Alzheimer's."

Weisman, who is also professor of Cell & Developmental Biology at the U-M Medical School and her colleagues, began from clues that were hidden in a conserved genetic pathway in yeast (a pathway that has remained the same in yeast, plants and humans over evolutionary time). Studies in yeast showed that the enzyme that manufactures the lipid is governed by the FIG4 and VAC14 genes, which exist in yeast, mice and humans.

Working with two independently derived mouse models, Weisman's team and collaborators including graduate student Clement Chow and Professor Miriam Meisler of the Department of Human Genetics at the U-M Medical School, reached the same conclusions in a pair of important papers for neuroscience research.

Building on research from Meisler, a mouse geneticist, and Weisman, a yeast geneticist, the collaborators published a paper in Nature, July 5, 2007, showing that in mice, the FIG4 gene is required to maintain normal levels of the signaling lipid and to maintain a normal nervous system. Importantly, they found that human patients with a very minor defect in their FIG4 genes had serious neurological problems.

The signaling lipid PI(3,5)P2 (short for phosphatidylinositol 3,5-bisphosphate) is part of a communication cascade that senses changes outside the cell and promotes actions inside the cell to accommodate to the changes.

Weisman's team found that mice missing the VAC14 gene, which encodes a regulator of PI(3,5)P2 levels, suffer massive neurodegeneration that looks nearly identical to the neurodegeneration seen in the FIG4 mutant mice. In both cases the levels of PI(3,5)P2 are one half of the normal levels. The fact that both mice have half the normal levels of the lipid and also have the same neurodegenerative problems provides evidence that there is a direct link between the lipid and neuronal health.

The new findings indicate that when Vac14 is removed, the cell bodies of many of the neurons appear to be empty spaces and the brain takes on a spongiform appearance.

The paper appearing in the online version of the Proceedings of the National Academy of Sciences October 22, 2007 is "Loss of Vac14, a regulator of the signaling lipid phosphatidylinositol 3,5-bisphosphate, results in neurodegeneration in mice," by a team of collaborators from U-M: Yanling Zhang, Sergey N. Zolov, Clement Y. Chow, Shalom G. Slutsky, Randal J. Westrick, Sean J. Morrison, Miriam H. Meisler, and Lois S. Weisman and the University of Iowa: Simon C. Richardson, Robert C. Piper, Baoli Yang, and Johnathan J. Nau.

The previous paper "Mutation of FIG4 causes neurodegeneration in the pale tremor mouse and patients with CMT4J," was published in Nature on-line, June 17, 2007.

— In work originating from the Bavarian Research Cooperation Prions (FORPRION), which ended in 2007, a team led by the scientist Prof. Dr. Christine Leib-Mösch has been able to show that prion proteins may activate endogenous retroviruses in infected brain cells.

In the Institute of Molecular Virology of the GSF – National Research Center for Environment and Health in Neuherberg/Munich (Helmholtz Association of German Research Centres) the group is continuing to search for cellular components whose make-up is changed as a result of a prion infection. In collaboration with colleagues from the Technical University of Munich and the University of Heidelberg,  the group used micro-array technologies – micro-arrays are chips with thousands or tens of thousands of DNA or protein probes – and could demonstrate that the expression of endogenous retroviruses is influenced by infectious prion proteins in tests with mouse cells.

Prions – an abbreviation for proteinaceous infectious particles – work as a trigger to a set of diseases of the brain and nervous system, the so-called spongiform encephalopathies. These include BSE in cattle, scrapie in sheep and Creutzfeldt Jakob’s Disease in humans. Prions are structural variants of a normal protein found in healthy tissues – especially in the brain.

The devastating effect of infectious prions is that, once they have entered the organism, they can modify the normal "healthy" prion proteins to create more infectious prions, and thus cause the illness to progress. However, as yet, little is known about the molecular mechanisms of pathogenesis, the role of co-factors and the interaction of prion proteins with cellular components.

Retroviruses insert their genetic information into the genome of host cells. In the case of endogenous retroviruses, this involves retroviral infections from long ago, which were transmitted through many generations by means of the germ line. Nearly ten percent of the genome of mice and humans consists of endogenous retroviral sequences that have accumulated during the course of evolution. Indeed, most structural genes of endogenous retroviruses are inactive, but many regulatory elements, such as binding sites for transcription factors, often remain active and can influence neighbouring cellular genes.

The GSF scientists infected mouse neural cells kept in culture with infectious prion proteins and subsequently analysed the expression patterns of endogenous retroviruses. The results showed that the expression of a set of endogenous retroviral sequences is influenced by the prion infection: in comparison with uninfected cells, the expression partly increased but also partly decreased – depending on the cell line and the type of endogenous retroviruses. These effects could be suppressed by pentosan-polysulphate, an anti-prion drug, which means that the influence of the expression can be attributed to the prions and not to some secondary effects.

These observations suggest that prion proteins may stimulate the production of retroviral particles by activation of endogenous retroviruses. Subsequently, these retrovirus-like particles could transport prion proteins from cell to cell, and thus spread the infection.

These studies were carried out within the scope of the “Bavarian Research Cooperation Prions” (FORPRION) in the Association of Bavarian Research Cooperations. FORPRION was founded in 2001 following the appearance of the first BSE cases in Bavaria and was financed equally from the budgets of the Bavarian State Ministry for Science, Research and Art, and the Bavarian State Ministry of Health Food and Consumer Affairs. 

Through basic and applied research the consortium aims to make progress in the diagnosis and therapy of human and animal prion diseases, as well as in the field of preventive consumer protection. FORPRION linked up 25 projects, based at five Bavarian universities and in institutes of the Max Planck Society. The financial support of the Bavarian Research Cooperation Prions FORPRION ended in June, 2007.

Reference: A. Stengel, C. Bach, I. Vorberg, O. Frank, S. Gilch, G. Lutzny, W. Seifarth, V. Erfle, E. Maas, H. Schätzl, C. Leib-Mösch, A. D. Greenwood: Prion infection influences murine endogenous in neuronal cells. Biochemical and Biophysical Research Communications 343 (2006) 825–831

Working in close collaboration with an international group of researchers, scientists at The Scripps Research Institute have shown for the first time that small clumps of abnormal prion proteins called oligomers cause the widespread death of neurons. In contrast, much larger prion aggregates known as fibrils proved to be far less toxic.

The findings suggest that small protein aggregates play a central role in prion diseases; similar mechanisms have been proposed for the so-called "amyloid" neurodegenerative diseases, including Alzheimer's. The work may provide novel therapeutic approaches for treating people with these conditions. 

"Our new study clearly establishes these misfolded prion protein oligomers as the major neurotoxic agent in both in vitro and in vivo experiments," said Professor Corinne Lasm⁄zas, a Scripps Research scientist in the Florida campus's Department of Infectology who led the study. "This new discovery reveals the most likely culprit responsible for the death of neurons associated with spongiform encephalopathies and probably other neurodegenerative diseases."

The researchers posit that prion oligomers damage neurons by disturbing neuronal membranes and hence cell signaling, as well as by building up excessively within cells, eventually triggering apoptotic or programmed cell death.

The brain on prions

Infectious prions (short for proteinaceous infectious particles) are unique pathogens associated with some 20 different diseases in humans and other animals, including mad cow disease and a rare human form, Creutzfeldt-Jacob disease. Prions, thought to be composed solely of protein, have the ability to reproduce, despite the fact that no nucleic acid genome has yet been found.

Mammalian cells normally produce what is known as cellular prion protein; during infection with a prion disease, though, the abnormal protein converts the normal host prion protein into a toxic form.

Oligomers are an intermediate aggregation state of the proteins, which start out as individual molecules or monomers. The fibril end-stage consists of much larger clumps or sheets (polymers) of proteins.

"When we look at the brain of an individual or an animal affected by a prion disease," Lasm⁄zas noted, "we don't find neurons dying in the same region as large fibril deposits (also called plaques). One theory suggests that these large fibril deposits may actually be the brain's way of containing the toxicity of the intermediate-stage oligomers."

Toxic proteins are instead likely to accumulate in intracellular degradation pathways like the proteasome—the part of the cell designed to eliminate damaged or unwanted proteins. Old age brings with it an increased tendency for proteasome dysfunction and protein damage or misfolding, which could explain the increase in amyloid diseases in later life.

Preventing neuron death

In the new study, the scientists first analyzed the neurotoxic properties of prion protein oligomers in neuronal cultures. Exposure of these neurons to prion protein oligomers resulted in a loss of nearly 50 percent of the neurons when compared to the untreated control cells, a level considered highly toxic.

The study's results also showed that exposure of a specific surface region of the prion protein oligomer was required to initiate this common neurotoxic mechanism; antibodies that recognized this region were able to inhibit cellular toxicity and prevent neuronal death.

"In our in vitro studies, there was a dramatic antibody effect—if you block this region of the oligomers, you completely inhibit neuronal toxicity," she said. "This region represents a very good therapeutic target, but there may be other target regions as well."

The scientists' in vivo findings with mouse models supported the picture that small prion aggregates, rather than plaque-type prion deposits, were responsible for neuronal dysfunction and death.

"Our new work demonstrates that the prion-induced neurodegeneration mechanism we uncovered in prion diseases is similar to that of other diseases such as Alzheimer's, Huntington's, and Parkinson's," Lasm⁄zas said. "The degree of this commonality is remarkable, and our findings open new avenues for the development of neuroprotective strategies that directly target toxic prion oligomers."

Other authors of the study, "In Vitro and In Vivo Neurotoxicity of Prion Protein Oligomers," include Steve Simoneau, Jean-Guy Fournier and Julien Comte of the Commissariat a` l'Energie Atomique, France; Human Rezaei and Jeanne Grosclaude of the Institut National de la Recherche Agronomique, France; Gunnar Kaiser-Schulz, Franziska Wopfner and Hermann Sch…tzl of the Institute of Virology, Germany; Nicole Sales of The Scripps Research Institute; Maxime Lefebvre-Roque of the Commissariat √ l'Energie Atomique and The Scripps Research Institute; and Catherine Vidal of the Institut Pasteur, France.

The study was published in the August 31, 2007, online edition of the journal PloS Pathogens. (doi=10.1371/journal.ppat.0030125) 

The study was supported by the European Union, the NeuroPrion Network of Excellence, and the German Science Foundation.