Prion Theory of Mad Cow Disease Called Full of Holes

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frenchie

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Prion Theory of Mad Cow Disease Called Full of Holes


By Neil Osterweil, Senior Associate Editor, MedPage Today
Reviewed by Robert Jasmer, MD; Assistant Professor of Medicine, University of California, San Francisco
March 30, 2006


MedPage Today Action Points
• Explain to interested patients that the authors of this study suggest that abnormal prion proteins implicated in the development of transmissible spongiform encephalopathies, such as variant Creutzfeldt-Jakob disease, may be markers for disease rather than the infectious agents.
• Explain further that this conclusion comes from animal research, and that there is a great deal of uncertainty about the origins of variant Creutzfeldt-Jakob disease and transmission risk in humans.
Review
PENICUIK, Scotland, March 30 - Just when the fog was beginning to clear on how variant Creutzfeldt-Jakob) disease (vCJD infects humans, more fog has rolled in.
Although precious little is known about the transmission of vCJD, it is widely assumed that the disease is caused by eating beef from cattle with bovine spongiform encephalopathy (BSE), the so-called mad-cow disease.
The prion protein infection from transmissible BSE is then thought to travel to the brain via peripheral nerves, perhaps with assistance from the lymphoreticular system.
Now, research by investigators here and in Norway into the transmission route of the brain-wasting infection scrapie in sheep, a related prion-associated disease, has cast doubt on whether abnormal prion proteins are truly the infectious agents for vCJD disease infection in humans after all.
Their research questions whether prion protein is really infectious, and it suggests that prion protein is merely a secondary marker of the presence of the scrapie agent. If that is so, as their findings indicate, it might also be so for ostensibly infectious nature of prions in vCJD. That, however, leaves open the issue of what the disease's infectious agent might be.
Although people with a specific genetic mutation are thought to be less susceptible than others to infection with vCJD because their digestive systems degrade the infectious prion protein, experiments in sheep indicate that prion protein is taken up equally in the gut by animals with ostensibly protective, neutral, or susceptible mutations in the same gene.
So reported Martin Jeffrey, Ph.D., the Veterinary Laboratories Agency-Lasswade here, and colleagues, in the April issue of the Journal of Pathology.
"This clearly shows that resistance is not achieved by blocking uptake of abnormal proteins from the gut," said Dr. Jeffrey." It must be achieved by some other mechanism."
A specific polymorphism, or variant, in the prion protein gene in humans has been shown to be a major determinant of susceptibility to prion diseases. The polymorphism, which occurs at codon 129 of the gene, determines whether the amino acids methionine or valine is present at that location.
About 40% of Caucasians are homozygous for methionine, an additional 50% of Caucasians heterozygous (i.e., have one valine and one methionine allele), and another 10% are homozygous for valine.
Some researchers suggest that resistance to BSE/vCJD infection in humans is conferred by decreased intestinal absorption of prions in people who carry the valine-valine polymorphism in the prion-protein gene, and that people with other mutations more readily absorb the proteins. They would thus be more susceptible to developing the brain-eating disorder.
But in their studies in sheep, at least, Dr. Jeffrey and colleagues found that both the abnormal and normal forms of the protein are absorbed equally in the gut across all genotypes, and that amplification of the damaged protein that occurs in sheep that go on to develop scrapie occurs else in the gastrointestinal tract.
They found this by conducting experiments in 50 sheep with varying degrees of genetic resistance to scrapie. They inoculated surgically isolated gut-loops with a scrapie brain pool homogenate, normal brain, or sucrose solution.
After surgery, the animals were sacrificed anywhere from 15 minutes to one month, or when they reached clinical disease.
Using immuohistochemistry, the investigators found evidence of the injected prion protein in villous lacteals and in submucosal lymphatics from 15 minutes to 3.5 hours post-challenge. They also found the inoculum-associated protein in dendritic-like cells in the draining lymph nodes at up to 24 hours post-challenge.
The protein was not, however, taken up by Peyer's nodules, where replication of infection is known to occur. Replication in the nodules was demonstrated by the accumulation of disease-associated forms of prion protein in the nodules at 30 days and 18 to 22 month in sheep that developed clinical signs of scrapie.
"These results indicate discrepancies between the routes of transportation of prion-protein from the inoculum and sites of de novo-generated disease-associated prion-protein subsequent to scrapie agent replication," the authors wrote.
"The fact the prion protein isn't taken up by the Peyer's nodules questions whether prion protein is really infectious, or whether prion protein is really just a secondary marker of the presence of the scrapie agent," Dr. Jeffrey said.
Further doubts about the gut transmission came from the fact that even after a pre-digested mixture containing high levels of disease-specific prion protein and alimentary fluids was injected into the gut, only faint traces of the prion protein could be detected on high sensitivity Western blotting.
"Think about it," Dr. Jeffrey said. "A sheep grazing in a field is not naturally exposed to highly infected brain and could only pick up a tiny amount of prion protein from other tissues. This will then be exposed to 48 hours or more digestion before it arrives in the gut, and our experiments show that after this, the chance of there being more than an immeasurably small amount of prion protein left to absorb is very small."
"As sheep can become infected, the theoretical probability of this being due to an invisible subfraction of digestion-resistant prion protein molecules is unlikely," he continued. "The possibility of there being infectious molecules other than prion protein must therefore be seriously considered."
In an accompanying commentary, Nicole Sales, Ph.D., of the department of infectology at the Scripps Research Institute in Jupiter, Fla., suggested that the study by Dr. Martin and colleagues argues for a stronger role of oral transmission of spongiform encephalopathy in humans, particularly among younger people.
"As Jeffrey et al's study shows that the inoculum is substantially digested in the distal part of the digestive tract, it raises the possibility that the oral cavity plays a role in the infection process," she wrote. "In this site, where there is a 'stronger' nondigested inoculum, the loss of the first dentition teeth or tonsillectomies may constitute aggravating factors as suggested previously/ Along the same lines, infection by the BSE agent has been performed via the intratonsillar route in non-human primates and has proved to be very efficient."
Dr. Jeffery did not buy this argument, however.
"Were infection to be acquired from the mouth, then the first tissues to accumulate infectivity would be lymph nodes in the throat or the tonsils. But we don't tend to see this in animals, and have no reason to believe it would be different in humans," he countered.

Primary source: Journal of Pathology
Source reference:
Jeffrey, M et. al. "Transportation of prion protein across the intestinal mucosa of scrapie-susceptible and scrapie-resistant sheep." J Pathol 2006; 209: 4-14
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What can we learn from the oral intake of prions by sheep?
N Sales, PhD *
Department of Infectology, The Scripps Research Institute, 5353 Parkside Drive, Jupiter, Florida, USA

email: N Sales ([email protected])

*Correspondence to N Sales, Department of Infectology, The Scripps Research Institute, 5353 Parkside Drive, Jupiter, Florida, USA.

Keywords
prion • scrapie • sheep • ingestion • vCJD • BSE • TSE


Abstract
The central nervous system is the ultimate target of prions, the agents responsible for fatal neurodegenerative diseases known as transmissible spongiform encephalopathies (TSEs). The neuro-invasive phase and its related clinical signs take place after a long incubation period. During this asymptomatic phase, however, active transport and replication of the infectious agent take place in peripheral sites. The oral infection route has been extensively studied because of its implication in the recent epidemic of bovine spongiform encephalopathy (BSE) in cattle and of the resulting human cases of variant Creutzfeldt-Jakob disease (vCJD). Rodent models have been useful in studying some aspects of this pathogenesis. Now, new data on the initial steps of oral infection have been obtained in sheep. This species is naturally infected with scrapie by horizontal transmission and there is strong evidence implicating the oral route. Furthermore, the existence of resistant and susceptible genotypes offers the possibility of comparative studies. The data were obtained using surgical and biochemical procedures to modulate the efficiency of oral infection and show that, in sheep, the abnormal prion protein (PrP) associated with the infectious agent crosses the intact intestinal barrier at the level of the enterocytes and then passes rapidly into lymph. These steps are identical in susceptible and resistant sheep. Thereafter, replication takes place in lymphoid structures. Other results in the same study indicate that alimentary fluids almost completely degrade the PrP of the inoculum. Though not directly transposable to human diseases, in which it is not possible to study these early stages, these data allow the elaboration of a simplified concept for the pathogenesis of TSEs. They also suggest that human contamination at the level of the oral cavity might be more important than previously suspected. Copyright © 2006 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.



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Received: 27 January 2006; Accepted: 27 January 2006

http://www3.interscience.wiley.com/cgi- ... 4/ABSTRACT


Original Paper
Transportation of prion protein across the intestinal mucosa of scrapie-susceptible and scrapie-resistant sheep
M Jeffrey 1 *, L González 1, A Espenes 2, CMcL Press 2, S Martin 1, M Chaplin 3, L Davis 3, T Landsverk 2, C MacAldowie 4, S Eaton 4, G McGovern 1
1Veterinary Laboratories Agency (VLA)-Lasswade, Pentlands Science Park, Bush Loan, Penicuik, Midlothian EH26 0PZ, UK
2Department of Basic Sciences and Aquatic Medicine, Norwegian School of Veterinary Science, N-0033 Oslo, Norway
3VLA-Weybridge, New Haw, Addlestone, Surrey KT15 3NB, UK
4Moredun Research Institute, Pentlands Science Park, Bush Loan, Penicuik, Midlothian EH26 0PZ, UK

email: M Jeffrey ([email protected])

*Correspondence to M Jeffrey, Veterinary Laboratories Agency (VLA)-Lasswade, Pentlands Science Park, Bush Loan, Penicuik, Midlothian EH26 0PZ, UK.

Funded by:
UK DEFRA; Grant Number: SE1951, SE1955
EU; Grant Number: QLK5-CT-2001-02332

Keywords
intestine • alimentary • pathogenesis • scrapie • prion


Abstract
To determine the mechanisms of intestinal transport of infection, and early pathogenesis, of sheep scrapie, isolated gut-loops were inoculated to ensure that significant concentrations of scrapie agent would come into direct contact with the relevant ileal structures (epithelial, lymphoreticular, and nervous). Gut loops were inoculated with a scrapie brain pool homogenate or normal brain or sucrose solution. After surgery, animals were necropsied at time points ranging from 15 min to 1 month and at clinical end point. Inoculum-associated prion protein (PrP) was detected by immunohistochemistry in villous lacteals and in sub-mucosal lymphatics from 15 min to 3.5 h post-challenge. It was also detected in association with dendritic-like cells in the draining lymph nodes at up to 24 h post-challenge. Replication of infection, as demonstrated by the accumulation of disease-associated forms of PrP in Peyer's patches, was detected at 30 days and sheep developed clinical signs of scrapie at 18-22 months post-challenge. These results indicate discrepancies between the routes of transportation of PrP from the inoculum and sites of de novo-generated disease-associated PrP subsequent to scrapie agent replication. When samples of homogenized inoculum were incubated with alimentary tract fluids in vitro, only trace amounts of protease-resistant PrP could be detected by western blotting, suggesting that the majority of both normal and abnormal PrP within the inoculum is readily digested by alimentary fluids. Copyright © 2006 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.



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Received: 12 October 2005; Revised: 13 December 2005; Accepted: 23 December 2005
Digital Object Identifier (DO




http://www3.interscience.wiley.com/cgi- ... 5/ABSTRACT


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