Category: Mad Cow Disease

Researchers have provided new information about how communication among neurons may be prevented from deteriorating in conditions such as Alzheimer's disease (AD). The new results may lead to new therapies for the treatment of not only AD but also motor neuron diseases and prion diseases.

Most current research efforts to find a treatment for AD and similar conditions focuses on what happens to the main part — or body — of a neuron, but recent studies have examined how neuronal communication is impaired in human diseases such as AD. When a neuron interacts with another neuron, it uses an extension called an axon that releases chemicals, which diffuse across a tiny gap between the neurons called a synapse and crosses the other neuron.

Deterioration of synapses and axons can be delayed thanks to a protein created by a gene called the slow Wallerian degeneration (Wlds) gene. How this protein works is still a mystery, but it may lead to new therapies for the treatment of AD and other conditions.

Thomas H. Gillingwater and colleagues identified 16 proteins that are affected by the Wlds gene. Although details are still missing, Wlds probably prevents these proteins from deteriorating synapses and axons.

The scientists found that some of the proteins had previously been shown to deteriorate synapses and axons, but, unexpectedly, eight proteins regulate the function of mitochondria — cellular organelles that supply energy to cells. These results reveal for the first time that mitochondria are involved in the protection of neurons provided by the Wlds gene and suggest that targeting some of the proteins identified in this study may lead to novel therapies for the treatment of AD, motor neuron diseases, and prion diseases.

Article: "Differential proteomic analysis of synaptic proteins identifies potential cellular targets and protein mediators of synaptic neuroprotection conferred by the slow Wallerian degeneration (Wlds) gene," by Thomas M. Wishart, Janet M. Paterson, Duncan M. Short, Sara Meredith, Kevin A. Robertson, Calum Sutherland, Michael A. Cousin, Mayank B. Dutia, and Thomas H. Gillingwater

Scientists have discovered a new protein that may offer fresh insights into brain function in mad cow disease. "Our team has defined a second prion protein called 'Shadoo', that exists in addition to the well-known prion protein called 'PrP' " said Professor David Westaway, director of the Centre for Prions and Protein Folding Diseases at the University of Alberta.

"For decades we believed PrP was a unique nerve protein that folded into an abnormal shape and caused prion disease: end of story. This view is no longer accurate," Westaway adds.

The study was conducted jointly by the University of Toronto, University of Alberta, Case Western Reserve University (Ohio) and the McLaughlin Research Institute (Montana). The research is published today in the EMBO Journal and represents a culmination of work initiated at the University of Toronto in 1999, and then continued more recently at the University of Alberta.

This is the first discovery since 1985 of a new brain prion protein. "A second prion protein had been inferred by other research, based on indirect studies and the examination of DNA sequences," said lead author Joel Watts, a graduate student at the University of Toronto's Centre for Research in Neurodegenerative Diseases. "But we not only demonstrate that this theoretical protein really exists and shares several properties with healthy PrP; we have also defined an unexpected alteration in prion infections.

"As the PrP molecule alters shape and accumulates in a prion-affected brain, the Shadoo protein seems to disappear," Watts added. Since proteins in a living cell are the molecules "that do the work, this is likely to be significant," he said.

"Many facets of a prion disease like BSE are puzzling," Westaway said. "The puzzles include the cause of death of brain cells, the function of normal prion proteins, and the rules governing emergence and spread of prions from animal to animal. We believe the Shadoo protein can give us a fresh purchase on these important questions."

This research project was funded by the Canadian Institutes of Health Research (CIHR) and the Natural Sciences and Engineering Research Council (NSERC).

A role for prion proteins, the much debated agents of mad cow disease and vCJD, has been identified. It appears that the normal prions produced by the body help to prevent the plaques that build up in the brain to cause Alzheimer’s disease. The possible function for the mysterious proteins was discovered by a team of scientists led by Medical Research Council funded scientist Professor Nigel Hooper of the University of Leeds.

Alzheimer’s and diseases like variant Creutzfeldt-Jakob Disease follow similar patterns of disease progression and in some forms of prion disease share genetic features. These parallels prompted Professor Hooper’s team to look for a link between the different conditions. They found an apparent role for normal prion proteins in preventing Alzheimer’s disease.

"Our experiments have shown that the normal prion proteins found in brain cells reduce the formation of beta-amyloid, a protein that binds with others to build plaques in the brain that are found in Alzheimer’s disease," explains Professor Hooper.

He continues: "In vCJD, the normal version of prion protein, PrPc, found naturally in the brain is corrupted by infectious prions to cause disease. The normal function of PrPc has been unclear."

Using cells grown in the lab, the team looked at the effect of high and low levels of normal prion protein on the successful formation of beta amyloid, the source of Alzheimer’s plaques. They found that beta amyloid did not form in cells with higher than usual levels of PrPc. In comparison, when the level of PrPc was low or absent, beta amyloid formation was found to go back up again.

Mice genetically engineered to lack PrPc were also studied. Again, this revealed that in its absence, the harmful beta-amyloid proteins were able to form.

It appears that PrPc, the normal prion protein, exerts its beneficial effect by stopping an enzyme called beta-secretase from cutting up amyloid protein into the smaller beta-amyloid fragments needed to build plaques.

Further evidence for the protective role of normal prion proteins is provided by mutated versions that are linked to genetic forms of prion disease because beta-amyloid fragments are able to form when the normal prion protein is corrupted by genetic mutation.

Professor Hooper concludes: "Until now, the normal function of prion proteins has remained unclear, but our findings clearly identify a role for normal prion proteins in regulating the production of beta-amyloid and in doing so preventing formation of Alzheimer’s plaques. Whether this function is lost as a result of the normal ageing process, or if some people are more susceptible to it than others we don’t know yet."

"The next step for our research will be to look in more detail at how the prion protein controls beta amyloid, knowledge that could be used to design anti-Alzheimer’s drugs. Theoretically, if we can find a way of mimicking the prion’s function we should be able to halt the progress of Alzheimer’s. However, there’s still a lot of work to be done in looking at levels of prions in the human system and how these may alter as we age.

Sheep infected with scrapie and cows infected with BSE have elevated levels of manganese in their blood before clinical symptoms appear, according to new research.

The findings, published in the Journal of Animal Science, also show that scrapie-resistant sheep produce elevated levels of the metal when “challenged” with the disease.

This suggests that elevated manganese levels in the blood and central nervous system are caused by the animal’s initial response to the disease.

The findings raise the possibility of using manganese levels in the blood as a potential diagnostic marker for prion infection. At present, only post-mortem examination of the brain tissue gives a certain diagnosis.

Scrapie, Bovine Spongiform Encephalopathy (BSE) and Creutzfeldt-Jakob Disease (CJD) are neurodegenerative diseases that affect the brain and nervous system of sheep, cows and humans respectively.

They are transmitted by mis-formed prion proteins which cause tiny loss of brain cell in different regions of the brain, leading to impairment of brain function, including memory changes, personality changes and problems with movement that worsen over time.

“Definite diagnosis of prion disease is currently only possible post-mortem," said Professor David Brown from the University of Bath who led the study with colleagues from the universities of Hull and Edinburgh.

“These findings suggest that elevated blood manganese could be used as a robust diagnostic marker for prion infection, even before the onset of apparent clinical disease.

“In practice, however, it would be difficult implement a widespread screening programme, given that the mass spectrometry we use to measure levels is expensive and labour intensive.”

The research builds on the 2002 discovery that mice infected with scrapie have higher levels of manganese. This is the first time that tissue from farm animals infected with prion diseases have been studied in this way.

One of the most interesting findings from this study came from the analysis of blood samples from scrapie-resistant sheep.

When challenged with the disease, these sheep showed similar levels of manganese as non-resistant sheep challenged in the same way.

“Elevated levels of manganese in scrapie-resistant sheep imply that the change in blood manganese is a result of the scrapie challenge and not a consequence of scrapie pathology,” said Professor Brown, from the University of Bath’s Department of Biology & Biochemistry.

“Although these sheep are considered to be resistant to scrapie, they do show some indications that scrapie challenge results in similar metabolic changes as occur in non-resistant sheep.”

Another interesting finding was that although levels of manganese were elevated, there were differences in the blood levels of selenium and molybdenum in experimental and field cases of BSE in cows.

This suggests that the way a cow acquires the disease affects the metabolic processes involved.

“The origin of the increased manganese in the brains and blood of infected animals remains unknown,” said Professor Brown.

“The three possibilities are that there is decreased secretion of manganese from the body, release of manganese from other tissues or increased absorption of manganese from the environment.

“Currently there is insufficient evidence to favour any of these three theories.”

NewsPsychology (May 3, 2007) — An oral vaccine can prevent mice from developing a brain disease similar to mad cow disease, according to new research. Prion diseases, which include scrapie, mad cow disease, and chronic wasting disease, are fatal and there is no treatment or cure.

The disease spreads when an animal eats the body parts of other animals contaminated with prions. The disease causes dementia and abnormal limb movements. Prion is a protein that is also an infectious agent. The proteins are so similar to proteins found normally that the immune system does not fight them off. To develop a vaccine that would stimulate the mice’s immune system, researchers attached prion proteins to a genetically modified strain of Salmonella.

For the study, the mice were orally vaccinated with a safe, attenuated Salmonella strain, which expressed the prion protein. Then they were divided into two groups — those who had high levels of antibodies in their blood and thus responded well to the vaccine and those with low levels of antibodies.

The mice with high levels of antibodies had no symptoms of the disease after 400 days. The mice with low levels of antibodies also had a significant delay in the onset of the disease. It normally takes 120 days for mice that have not been vaccinated to develop the disease.

“These are promising findings,” said study author Thomas Wisniewski, MD, of NYU School of Medicine in New York, and a member of the American Academy of Neurology. “We are now in the process of redesigning the vaccine so it can be used on deer and cattle.”

Wisniewski said much more work is needed before the vaccine could be considered for humans. “The human version of prion disease usually occurs spontaneously and only rarely because of eating contaminated meat,” he said. “But if, for example, a more significant outbreak of chronic wasting disease in deer and elk occurs and if it were transmissible to humans, then we would need a vaccine like this to protect people in hunting areas.”

He also noted that a vaccine that decreases the spread of prion disease in animals also reduces the possibility that the disease could infect humans. These findings were presented at the American Academy of Neurology’s 59th Annual Meeting in Boston, April 28 — May 5, 2007.

The study was supported by grants from the National Institutes of Health.

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The above story is reprinted (with editorial adaptations by newsPsychology staff) from materials provided by American Academy of Neurology.

Disclaimer: This article is not intended to provide medical advice, diagnosis or treatment. Views expressed here do not necessarily reflect those of NewsPsychology or its staff.

Burying prion-infected carcasses of cattle, deer and other animals in lime may actually enhance the spread of those infectious proteins through soil, a new study suggests. Placing quicklime on carcasses once was thought to be the best way to foster quick decay of bodies and to prevent the spread of disease.

The study is scheduled for the April 15 issue of ACS’ Environmental Science & Technology, a semi-monthly journal.

In the study, Joel A. Pedersen and colleagues from the University of Wisconsin cite the need for safe methods of disposing of prion-infected carcasses, noting that prions can resist harsh conditions such as strong disinfectants and dry-heat temperatures of 1,100°F that destroy other disease-causing agents and that prions can remain infectious in the soil for at least three years. Pedersen and colleagues investigated the effect of different conditions (pH, salinity) on the adsorption, or attachment, of prions to sand particles.

They found that prions become less firmly attached to sand particles, and thus potentially more mobile, under alkaline conditions. These conditions would be produced by lime, as well as in older landfills. In the natural environment, acidic conditions may keep prions near the soil surface, increasing the risk that animals will ingest prions and become infected, the report says. The team is conducting further research to determine whether these expectations are borne out.

For the first time, experimental results indicate that it is possible to use a resin filter to remove harmful prion proteins from the blood of an infected animal, a finding that has major implications for the removal of infectious prion proteins – the agents associated with variant Creutzfeldt-Jakob disease, mad cow disease, scrapie and other prion diseases in animals – during blood transfusions.

Dr. Ruben Carbonell, Frank Hawkins Kenan Professor of Chemical and Biomolecular Engineering and director of the Kenan Institute for Engineering, Technology and Science at North Carolina State University, and scientists from the University of Maryland at Baltimore’s VA Medical Center, the American Red Cross and ProMetic BioSciences, a biotechnology company, developed small resin beads with molecules that are able to bind to harmful prion proteins. The beads serve as an adsorption filter, capturing the bad proteins and allowing other blood components to be effectively cleansed of the prion-disease-causing agents.

A paper describing the research was published in the Dec. 23/30 version of The Lancet.

In prion diseases, which are called transmissible spongiform encephalopathies, prion proteins unfold and cause plaques in animal and human brains. Transmission of prion diseases has impacted the availability and cost of blood donations, especially in Europe.

In the Lancet study, the researchers took the blood of scrapie-infected hamsters and removed the white blood cells using a device called a leukofilter. The leukoreduced blood was then passed through another filter containing the new resin particles engineered to capture the prion proteins. A group of disease-free hamsters was inoculated with the blood that passed through the leukofilter only. A second group was inoculated with the blood that passed through both the leukofilter and the prion-capture filter.

The researchers found that while leukoreduction itself removed a good deal of the bad proteins – approximately 72 percent – none of the nearly 100 hamsters inoculated with the leukoreduced, resin-filtered blood were infected with scrapie by the end of the 550-day test. Fifteen of 99 hamsters receiving leukoreduced blood not passed through the resin filter were infected with scrapie.

After the experiment was completed, the researchers analyzed the brains of hamsters still alive at the end of the testing period. No evidence of scrapie was discovered in brains of hamsters that were inoculated with the resin-filtered blood.

Aided by scientists in NC State’s Nonwovens Cooperative Research Center, located in the College of Textiles, Carbonell and his colleagues have now developed a new filter to remove prions from donated blood during transfusions. The device takes donated blood from a blood bag, passes it through several “sandwiches” of the prion-capture resin beads placed between nonwoven fabric membranes, and places the filtered blood in a separate blood bag prior to infusion into a patient or blood donation recipient.

The filter device, to be manufactured under the trade name P-Capt® Filter by MacoPharma, has received CE Mark regulatory approval in Europe.

Carbonell and his colleagues are now looking for ways of targeting other pathogens in blood such as Hepatitis A virus, B19 parvovirus and Hepatitis C virus. He says it should be possible to engineer new molecules that capture prion proteins and viruses and to place them on a single filter to further enhance the safety of blood transfusions.

The research was funded by Pathogen Removal and Diagnostic Technologies Inc. (PRDT), a joint venture of the American Red Cross and ProMetic BioSciences, owned in part by Carbonell and Dr. Robert Rohwer from the University of Maryland.

Using a combination of novel technologies, scientists at the Scripps Research Institute and the Whitehead Institute for Biomedical Research have revealed for the first time a dynamic molecular portrait of individual unfolded yeast prions that form the compound amyloid, a fibrous protein aggregate associated with neurodegenerative diseases such as Alzheimer's disease and variant Creutzfeldt-Jacob disease — the human version of mad cow disease.

The new findings, which are being published the week of February 12 in an online edition of the Proceedings of the National Academy of Sciences, offer significant insights into normal folding mechanisms as well as those that lead to abnormal amyloid fibril conversion. The new insights may lead to the discovery of novel therapeutic targets for neurodegenerative diseases.

Intriguingly, certain prions and amyloids can play beneficial roles. The subject of the new study, Sup35, enables protein-based inheritance in yeast. When this prion protein misfolds, it converts into self-perpetuating amyloid fibrils, thus altering its function in an inheritable manner. The research team used a combination of advanced biophysical methods to investigate these processes.

"By focusing on single unfolded prions, we were able to define the dynamics of two distinct regions or domains that determine conversion dynamics," said Ashok A. Deniz, a Scripps Research scientist who led the study. "Our research techniques can now be used to probe the structures of other amyloidogenic proteins. This could prove important in understanding the basic biology of amyloid formation, as well as in designing strategies against misfolding diseases."

Interestingly, the new study revealed that yeast prion protein Sup35 lacks a specific, static structure in its native collapsed state. Instead, the compact protein fluctuates among several different structures before forming intermediate shapes during the amyloid assembly process.

The intermediate stages of the process are critically important, Deniz noted: "No single native unfolded protein is capable of initiating the amyloid cascade because of this constant shape-shifting. To start the amyloid conversion process, it has to first convert to an intermediate species, consisting of multiple protein molecules. This insight may be important to finding potential new therapeutic targets for disease-causing amyloids."

To define the dynamic structural details of individual prions, Deniz and his colleagues employed several novel technologies including single-molecule fluorescence resonance energy transfer (SM-FRET) and fluorescence correlation spectroscopy (FCS).

Fluorescence resonance energy transfer is a highly sensitive tool used to measure molecular structure and dynamics such as in single proteins at the angstrom level, a measurement unit used to define molecular distances (a 10th of a millionth of a millimeter). Fluorescence correlation spectroscopy is a high resolution technique that measures time fluctuations in fluorescent emissions from tagged proteins, which provided information about changes in shape of Sup35 taking place on the nanosecond timescale (billionths of seconds).

A third technology, single molecule fluorescence coincidence, was used in an unusual way-to prove that the protein species under scrutiny were not oligomeric (consisting of multiple proteins in an aggregate). The technology, based on measuring fluorescence bursts from individual tagged proteins, enabled the scientists to determine that the proteins being studied were, in fact, single monomers and not aggregates.

Deniz said that future work with yeast prion mutants might resolve some of the questions that remain unanswered. "Our laboratory has spent a great deal of time in improving these techniques, and we have used them to uncover some very intriguing information about this particular monomer," he said. "This combination of techniques can now be used to study other amyloidogenic proteins, including prions, particularly small assemblies and intermediate stages of the aggregation process. These are currently considered the most toxic forms of amyloid-disease associated proteins."

While mammalian prion proteins are different from those of yeast in their amino acid sequence, they do share some basic features, including their ability to catalyze the conversion to amyloid fibers. Some studies suggest that prions may also play key roles in certain critical processes such as long-term memory. Other authors of the study, A Natively Unfolded Yeast Prion Monomer Adopts An Ensemble of Collapsed and Rapidly Fluctuating Structures, are Samrat Mukhopadhyay and Edward A. Lemke of The Scripps Research Institute; and Susan Lindquist and Rajaraman Krishnan of the Whitehead Institute for Biomedical Research.

The study was supported by the National Institutes of Health, The DuPont-MIT Alliance, and the Alexander von Humboldt Foundation.

 Studies in mice have indicated that the effects of prion disease could be reversed if caught early enough. The researchers said that their findings support developing early treatments that aim to reduce levels of prion protein in the brains of people with prion disease. Also, they said that their findings suggest testing the efficacy of treatments in a new way: by analyzing their cognitive effects in prion-infected mice.

The researchers, Giovanna Mallucci and colleagues, reported their findings in the February 1, 2007 issue of the journal Neuron, published by Cell Press.

Prion disease–such as the version of Creutzfeldt-Jakob disease believed to be contracted from cattle with "mad cow disease"–is caused by aberrant, infective proteins. It has been thought that the disease is untreatable.

However, in previous studies with prion-infected mice, Mallucci and colleagues found that early brain degeneration can be reversed if prions are depleted in neurons.

In the new studies published in Neuron, they established that cognitive and behavioral impairments–which appear early in humans with prion disease–can be reversed if prion depletion is done early. What's more, they found that the neurological pathology of the disease is reversed along with the cognitive and behavioral deficits.

In their studies, the researchers measured the effects of prion disease on the animals' ability to discriminate novel objects in their cage and on normal burrowing behavior. In both cases, deficits in those abilities appeared early in the disease. Also, studies of the animals' brain tissue revealed a parallel impairment of signaling among brain cells.

However, when the researchers manipulated the animals to deplete their brains of the prion protein, their memory ability and burrowing behavior recovered. Importantly, found the researchers, the signaling among brain cells also recovered.

"Overall, we conclude that the dramatic benefits to neuronal function and survival in prion-infected mice we have shown here support targeting neuronal [prion protein] directly as a therapeutic approach," wrote Mallucci and colleagues.

"Our findings of early reversible neurophysiological and cognitive deficits occurring prior to neuronal loss open new avenues in the prion field," they wrote. "To date, prion infection in mice has conventionally been diagnosed when motor deficits reflect advanced neurodegeneration. Now the identification of earlier dysfunction helps direct the study of mechanisms of neurotoxicity and therapies to earlier stages of disease, when rescue is still possible.

"Eventually it may also enable preclinical testing of therapeutic strategies through cognitive endpoints. These data now lead to the hope that early intervention in human prion disease will not only halt clinical progression but allow reversal of early behavioral and cognitive abnormalities," wrote the scientists.

Do genes affect bovine spongiform encephalopathy–also known as BSE, or "mad cow" disease? Are some cattle more susceptible than others?

To address these and other questions, Agricultural Research Service (ARS) scientists at the U.S. Meat Animal Research Center in Clay Center, Neb., have sequenced the bovine prion gene (PRNP) in 192 cattle that represent 16 beef and five dairy breeds common in the United States.

This work, partially funded by a grant from the U.S. Department of Agriculture's Cooperative State Research, Education and Extension Service, is expanding the understanding of how the disease works.

BSE is a fatal neurological disorder characterized by prions–proteins that occur naturally in mammals–that fold irregularly. Molecular biologist Mike Clawson and his Clay Center colleagues are examining PRNP variation in order to learn if and how prions correlate with BSE susceptibility.

From the 192 PRNP sequences, Clawson and his colleagues have identified 388 variations, or polymorphisms, 287 of which were previously unknown. Some of these polymorphisms may influence BSE susceptibility in cattle.

Comparing PRNP sequences from infected and healthy cattle may enable researchers to identify genetic markers in the prion gene that predict BSE susceptibility. In addition to PRNP, the team is currently sequencing several closely related genes, which will also be tested for their association with BSE.

The prevalence of BSE in the United States is extremely low, but this research could improve understanding of the disease and prepare the cattle industry to respond if another prion disease should arise in the future.

ARS is the USDA's chief scientific research agency.