The diseases are characterised by loss of motor control, dementia, paralysis wasting and eventually death, typically following pneumonia. Fatal Familial Insomnia presents with an untreatable insomnia and dysautonomia. Details of pathogenesis are largely unknown.

Visible end results at post-mortem are non-inflammatory lesions, vacuoles, amyloid protein deposits and astrogliosis.

GSS is distinct from CJD, it occurs typically in the 4th-5th decade, characterised by cerebellar ataxia and concomitant motor problems, dementia less common and disease course lasts several years to death. (Originally thought to be familial, but now known to occur sporadically as well).
CJD typically occurs a decade later has cerebral involvement so dementia is more common and patient seldom survives a year (originally thought to be sporadic, but now known to be familial as well).
FFI pathology is characterised by severe selective atrophy of the thalamus.
Alpers syndrome is the name given to prion diseases in infants.

Scrapie was the first example of this type of disease to be noticed and has been known about for many hundreds of years. There are two possible methods of transmission in sheep:

1. Infection of pasture with placental tissue carrying the agent followed by ingestion,or direct sheep-lamb transmission i.e. an acquired infection.
2. Parry showed considerable foresight by suggesting that it is not normally an infectious disease at all but a genetic disorder. He further suggested that selective breeding would get rid of the disease.

Humans might be infected by prions in 2 ways:

1. Acquired infection (diet and following medical procedures such as surgery, growth hormone injections, corneal transplants) i.e. infectious agent implicated.
2. Apparent hereditary mendelian transmission where it is an autosomal and dominant trait. This is not prima facie consistent with an infectious agent.

This is one of the features that single out prion diseases for particular attention. They are both infectious and hereditary diseases (?see below). They are also sporadic, in the sense that there are also cases in which there is no known risk factor although it seems likely that infection was acquired in one of the two ways listed above.

Kuru is the condition which first brought prion diseases to prominence in the 1950s. Found in geographically isolated tribes in the Fore highlands of New Guinea. Established that ingesting brain tissue of dead relatives for religious reasons was likely to be the route of transmission. They ground up the brain into a pale grey soup, heated it and ate it. Clinically, the disease resembles CJD. Other tribes in the vicinity with same religious habit did not develop the disease. It is speculated that at some point in the past a tribe member developed CJD, and as brain tissue is highly infectious this allowed the disease to spread. Afflicted tribes were encouraged not to ingest brain tissue and the incidence of disease rapidly declined and is now almost unknown.

The Trembling Mountain: A Personal Account of Kuru, Cannibals, and Mad Cow Disease
Robert Klitzman.
Working for Nobel Prize-winner Carleton Gajdusek, Klitzman lived amid the Fore, a previously cannibalistic tribe in some of the most remote parts of the country. Their community had been devastated by kuru, a deadly and heartbreaking neurological disease, spread by the ritual consumption of deceased relatives (including brain matter). Over the course of the narrative, the young Klitzman interviews stricken individuals, comes to grips with hugely divergent cultures and comes of age himself. (Amazon.co.UK)

Evidence suggests that a prion is a modified form of a normal cellular protein known as PrPc (for cellular), a normal host protein encoded by a single exon of a single copy gene. This protein is found predominantly on the surface of neurones attached by a glycoinositol phospholipid anchor, and is protease sensitive. Thought to be involved in synaptic function.
The modified form of PrPc which may cause disease i.e. the prion is known as PrPsc (for scrapie) which is relatively resistant to proteases and accumulates in cytoplasmic vesicles of diseased individuals.

It has been proposed that PrPsc when introduced into a normal cell causes the conversion of PrPc into PrPsc. The exact nature of the process is unknown but it could involve a chemical or conformational modification.

Fatal Protein: The Story of CJD, BSE and Other Prion Diseases
Rosalind Ridley, Harry Baker.
A clear and authoritative survey of the prion diseases, designed mainly for the non-specialist. It deals with the history of scrapie in sheep, the spread of kuru through cannibalism in Papua New Guinea, and the BSE epidemic in Britain - "mad cow disease" - which claimed over 170,000 cattle and threatened the rest of Europe. The authors present a detailed description of the human prion diseases, and examine the evidence that the new variant of the Creutzfeld-Jakob disease results from eating infected by BSE. At the heart of this book lies a discussion of the still controversial idea that the infectious agent in prion diseases is a normal brain protein which takes on abnormal, mutant shape, the so-called "prion theory". (Amazon.co.UK)

Several lines of evidence support a protein only model of infection:

1. Nucleic acid is not necessary for infectivity:

• Unusually small target size for ultraviolet and ionising radiation:
  
• The low ratio of nucleic acid to infectious material.
• Resistance of infectivity to agents which modify or damage nucleic acids but infectivity is susceptible to reagents which destroy proteins
• Failure to identify a scrapie specific nucleic acid either in prion preparations or infected brains using a variety of sophisticated techniques.

+ = inactivated; - = no change in infectivity

2. PrPsc is associated with scrapie infectivity:

• purification of scrapie infectivity results in preparations highly enriched for PrPsc
• purification of PrPsc results in enrichment of scrapie activity
• purification of PrPsc by SDS-PAGE also recovers infectivity
• PrPsc can be denatured and renatured without loss of infectivity

3. Susceptibility of a host to prion infection is co-determined by the prion inoculum and the PrP gene:

• disease incubation time for a single prion isolate various between mouse strains, this variation depends on the PrP gene, suggesting some forms of PrPc may be more easily converted to PrPsc than others.
• when prions are transmitted from one species to another disease develops only after a very long incubation period, if at all, but on serial passage in the new species the incubation time often decreases dramatically and then stabilises. This species barrier can be overcome by introducing a PrP transgene from the prion donor i.e. hamster PrPc but not murine PrPc is a suitable substrate for conversion to hamster PrPsc by hamster prions and vice versa:

4. Mutated gene can cause susceptibility to disease without apparent infection:

• Homozygosity at the polymorphic amino acid position 129 of PrP protein predisposes an individual to acquired and sporadic CJD
• 2 unrelated GSS families have the same double mutation i.e. 178 D-N; 200 E-K
• disease tightly linked to a P-L mutation at 102 in some familial GSS cases. 100 controls did not have this mutation nor did another 15 sporadic victims of CJD
• other familial cases have been shown to carry this or several other mutations e.g. 117-A-V; 198 F-S; N178; V129 = CJD; N178 M129 = FFI

5. Crucial experiment:

Mice carrying a murine transgene with the 102P-L GSS mutation spontaneously develop a lethal scrapie like disease. Brains also contain infectious prions because transmission to recipient animals has been demonstrated.
Mice lacking the PrPc gene develop normally, no evident physiological or behavioural problems. Suggests that loss of PrPc function is unlikely to be the cause of disease rather accumulation of PrPsc is responsible. When inoculated with prions they do not develop disease . There are obvious implications here for the livestock and pharmaceutical industries and treatment of familial cases. Animals expressing reduced levels of PrPc are also resistant to infection - perhaps more immediately relevant to human disease. Animals overexpressing PrPc are more likely to develop prion diseases.

The Pathological Protein: Mad Cow, Chronic Wasting, and Other Deadly Prion Diseases
Philip Yam.
200,000 cattle, but how many humans?
(Amazon.co.UK)

Evidence against the prion model:

Are prion diseases genetic disorders?

• Ala136Ala, Arg154Arg, Gln171Gln
• Val136Val, Arg154Arg, Gln171Gln

So how can a protein be infectious?

However, a more likely possibility is that the difference between PrPc and PrPsc is conformational. 3D structure of part of the murine PrPc expressed in E. coli has recently been determined. As expected from spectroscopy measurements PrPc is predominantly alpha helical and contains almost no beta sheet. The structure of PrPsc has not yet been determined but is predicted to be predominantly B- sheet. Proposed therefore that this protein can adopt 2 quite different stable conformations. The safe PrPc form is normally adopted but rarely it can switch to the PrPsc form. Mutations favour this switch. Propose that PrPsc is transdominant and converts PrPc to PrPsc in an exponential fashion. Precedents for this model do exist. There is a yeast mutant phenotype which doesn't correlate to any mutational difference in the gene structure but does correlate to a different protein structure. The Ure2p protein converts to an inactive conformation. Mutations in the tumour suppressor protein p53 which are associated with the onset of neoplastic disease have a different conformation to the normal protein. When normal protein is incubated with mutant protein its conformation is altered to the mutant form.

The first image below shows the structure of part of the hamster and mouse PrPc molecules superimposed. The close similarity in structures is obvious, as is the preponderance of alpha helical structure.The second shows the position of various mutations important for prion disease development in humans modelled on the hamster structure PrPc.Many of these mutations are positioned such that they could disrupt the secondary structure of the molecule.

Both images from Dr Thomas Pringle of the Sperling Foundation Official Mad Cow Disease Home Page

Strain Typing:

The existence of multiple strains of prion agents by definition means that the agents carry strain specific information. The virino hypothesis states that this is a small nucleic acid molecule. The prion hypothesis states that it is due to differences in the chemical or tertiary structure of the prion protein.
Strain discrimination relies on the use of mice of defined genotype and having different PrP gene sequences.
Measurement of parameters such as incubation period in these mice and the pathological changes occurring in the brain are made.

Measurement of Incubation period: Precise quantities of brain homogenates from clinically affected animals are injected intracerebrally into mice. Animals are then assessed regulary for the definite appearance of disease using a defined set of observations. This measurement is very precise with errors of less than 2% of the mean:

Pathological changes: Strains show very different and reproducible differences in the pattern of vacuolar degeneration of different inbred mice. These effects are scored in a variety of different brain sections. Up to 12 topologically specific sites of the brain are scored for vacuolation e.g. cortex, cerebellum, brain stem etc. This semiquantitative assessment or lesion profile is then plotted. Individual strains have a characteristic and highly reproducible lesion profile in a given mouse genotype e.g. Using this technique it is very clear that the recent outbreaks of new TSE diseases in various animals is caused by a prion with very similar characteristics ie BSE and quite distinct from previously recognized scrapie prion strains. Strain typing has also demonstrated beyond reasonable doubt in the last few months that nv-CJD or v-CJD are caused by a strain very much like the BSE agent. Some people now call this disease Human BSE:

BSE and vCJD

It is likely that a large proportion of the UK population was exposed to the infectious agent responsible for vCJD during the 1980-90s. The size of the epidemic resulting from this exposure will depend on what is the infectious dose exposure via the gastric route (which may be cumulative) and the size of the species barrier between animals and humans.

On the 18th January 2007 the UK HPA announced that there had been a fourth case of vCJD diagnosed in a patient nine years after they received a blood transfusion from a donor who later went on to develop vCJD. A recently published paper predicts that there will be 50 deaths from this source of vCJD infection by 2080. Professor Christopher Higgins, chair of the Spongiform Encephalopathy Advisory Committee (SEAC), said: "In the future, blood transfusions are going to be the most likely way of transmission":
Is there the potential for an epidemic of variant Creutzfeldt-Jakob disease via blood transfusion in the UK? J.R.Soc Interface Feb 7, 2007.

Deadly Feasts: The "Prion" Controversy and the Public's Health
Richard Rhodes.
You might think you're not interested in prion diseases, or maybe you're not worried because Mad Cow and its grim relatives will never cross the ocean to North America. Perhaps you even believe that as a vegetarian, these issues could never affect you. In "Deadly Feasts," Richard Rhodes shows in gruesome detail how very wrong you may be in those assumptions. (Amazon.co.UK)

Infection and Pathogenesis

• SCID mice are resistant to a prion challenge, confirming the importance of the lymphoid system.
• PrP null mice ie those in which both alleles have been disrupted, PrP ^o/o, cannot be infected.
• PrP^o/omice carrying a PrP ^+/+ brain graft can develop pathology following an intracerebral injection but only in the graft.
• PrP^o/o mice + graft + a reconstituted PrP^+/+ lymphoreticular system are resistant to a prion challenge.

This indicates another compartment in addition to the brain and the LRS must express PrP, if a peripheral prion challenge is to be successful. That compartment is probably a nerve. Mature B lymphocytes are also now known to be required for the development of the disease following infection from a peripheral route.

Tg PrP mice and knockout mice have been fantastically informative in prion research. In a further series of recent experiments PrP^o/o mice have had a hamster PrP transgene incorporated. This transgene has been put under the transcriptional control of either the glial fibrillary acidic protein promoter, or the neuron specific enolase promoter such that hamster PrP is only expressed in either the glial cells or neuronal cells. Following an intracerebral prion challenge both groups of animals can replicate the prion agent and develop disease pathology. So either glial cells or neural cells can propagate the disease independently. The fact that PrPsc intracerebral injection alone in PrP^o/o mice, does not cause pathology means that cells must be making PrP for a pathological result. It is known that cytokine levels are elevated in the later stages of a prion infection. Astrocytes and other glial cells produce these and presumably have a key role in pathogenesis. This conclusion is supported by the fact that murine retroviruses are known which infect glial cells and result in a spongiform degeneration of the brain.

Treatment:

MicroBiologyBytes: The Origin of BSE

Further Information:

• How Now Mad Cow: Online tutorial.
• Collinge J. (2001) Prion diseases of humans and animals: their causes and molecular basis. Ann Rev Neurosci. 24: 519-550
• Edskes HK. and Wickner RB. (2004) Transmissible spongiform encephalopathies: prion proof in progress. Nature 430: 977-979
• Soto C, Castilla J. (2004) The controversial protein-only hypothesis of prion propagation. Nat Med. 10: S63-67
• Wickner RB. et al. (2004) Prions: proteins as genes and infectious entities. Genes Dev. 18: 470-485

© MicrobiologyBytes 2007.