Prion disease

Question 1

Define neurodegenerative disease

Answer 1

Group of related disorders with a variety of pathologies and aetiologies that are all characterised by progressive loss of function and eventual neuronal death.

Associated with ageing.

Question 2

2 common features of neurodegenerative diseases?

Answer 2

Progressive synapse loss
Build up of misfolded protein aggregates in the brain.

Question 3

What are prion diseases?

Answer 3

Also known as Transmissible spongiform encephalopathies (TSEs): rarer forms of neurodegeneration.

Features: accumulation of misfolded protein and progressive synapse loss as well as - spongiosis (holes in tissue) and astrocytosis (overproliferation of astrocytes).

Question 4

Evidence of glial change in prion disease?

Answer 4

GFAP: marker of astrocytes, showed increased number of astroctyes in infected.

Iba1: marker of microglia, showed increased number of microglia in infected.

Question 5

How does EEG of infected differ?

Answer 5

In late stages of disease: characteristic periodic sharp wave complexes (PSWC).

Aguglia et al., (1977): case studies of two women with CJD who developed PSWC but later lost PSWC shortly before death. Hypothesised that this could be due to increased GABA mediated inhibition that reduces amplitudes of evoked potentials before death in CJD.

Question 6

Why can’t we say that the spread of neurodegenerative diseases through protein misfolding is specific to prion disease?

Answer 6

Other neurodegenerative have prion-like propagation of misfolded protein.

Prion: PrP
AD/dementia: amyloidBeta and tau
Other tauopathies: tau
Parkinson’s: alpha-synuclein

Question 7

What is the worsening of neurodegenerative diseases often correlated to?

Answer 7

Increased protein accumulation

Question 8

Describe Scrapie

Answer 8

First described prion disease: found in sheep.

Infected: loss of coordination, uncontrollable urge to itch, ataxia, progressive paralysis and certain death.

Question 9

How was scrapie proved to be transmissible?

Answer 9

Cuillé and Chelle, 1936: performed intraocular inoculation (basically used brain homogenate) of brain tissue from an affected animal into two healthy sheep who subsequently developed.

Further demonstrated by accidental inoculation: vaccine targeting looping ill virus was created from lymphoid tissue of scrapie-infected animals and went on to infect hundreds of sheep that received vaccine.

Question 10

Describe kuru

Answer 10

Human prion disease that was devastating the Fore people of Papua New Guinea that pathologically resembles scrapie.
Confirmed to be infectious by transferring the disease to chimpanzees.

Transferred via ritualistic cannibalism: tribe ate dead to show love for them.
Eradicated after abolishment of practice.

Question 11

Describe BSE

Answer 11

Bovine Spongiform Encephalopathy: unsure whether crossed species barrier from sheep or occurred sporadically but the number of cases greatly increased when infected carcases were processed into cattle feed.

Infected cattle fodder was also fed to other animals and resulted in new TSEs (cats, greater kudu, cheetahs ostriches and more!)

Naturally, led to concerns of transmission to humans: new form of human prion disease CJD (Creutzfeldt-Jakob disease) was discovered and termed variant CJD (vCJD).

Question 12

Describe Creutzfeldt-Jacob disease

Answer 12

Most common human prion disease and is also known as subacute spongiform encephalopathy or neurocognitive disorder due to prion disease.

Always terminal: rapidly progressing dementia, development of movement disorder such as tremor, most have characteristic PSWC.
Death after 4 months.

Question 13

What are the different types of CJD disease?

Answer 13

1) sporadic: 90%, unknown stim results in formation of PrP Sc.
2) Genetic: 10%, genetic abnormality in Prnp gene.
3) Transmitted: rare
4) Iatrogenic: v rare - specific to surgery as very hard to clean medical. If had brain surgery after infected there is risk.

NB: overall there are 3 forms of prion disease with transmitted and iatrogenic combined into acquired category.

Question 14

Outline vCJD

Answer 14

Clinically and pathologically resembles CJD but has:
Longer duration (with dementia occurring much later)
Prolonged psychiatric symptoms
Only affected young people (mean onset 29 but patients as young as 16 developed it)
Frequently EEG changes were absent

Question 15

What is the molecular mechanism of prion disease?

Answer 15

Prnp gene encodes PrP protein of which there are 2 isoforms, same primary structure but altered secondary and tertiary.
1) PrP C: normal endogenous, secondary mainly alpha helices
2) PrP Sc: mutated, secondary mainly beta pleated sheets = insoluble and resistant to protease degradation.

PrP Sc is infective as initiates the conversion of PrP C into more PrP Sc. As brain becomes depleted of c it stimulates synthesis of more and means more substrate for pathological conversion.

Question 16

Why is cross species infectivity uncommon?

Answer 16

Ability of PrP Sc to recruit and convert PrP C is reduced or abolished due to variances in amino acid sequence.

Question 17

PrP function?

Answer 17

Prnp encodes: Ubiquitously expressed gylcosylphosphatidylinositol (GPI) anchored glycoprotein that resides at the cell surface.

Role unknown but tightly developmentally regulated and highly conserved between mammalian species - suggests essential biological function.

Question 18

Prnp KO?

Answer 18

However, Prnp knockout mouse models have been generated and shown to be developmentally and phenotypically normal (Bueler et al.,1992; Manson et al.,1994a). This could be due to compensatory mechanisms.
Subtle alterations: impaired LTP and reduced afterhyperpolarisation potentials. Weakened GABA A mediated fast inhibition.
So overall could be role in modulation of neuronal excitability.

Even adult onset conditional knockout mice remain healthy. In these mice the Cre-LoxP system was used to excise the Prnp gene 9-10 weeks after birth (Mallucci et al.,2002).

KO mice cannot get prion disease which supports idea that PrP Sc is infectious agent

Question 19

Primary rat neurons cultured with PrP C?

Answer 19

Increased axon, dendrite and synapse formation and suggest role for PrP in maintenance of hippocampal networks

Question 20

How do you destroy prions?

Answer 20

It is very difficult to destroy prions and this poses risks not only for transmission of human prion as previously mentioned but also for animal cases i.e. BSE and how to dispose of carcasses etc. Known as zombie proteins as if not completely denatured they fold back into correct confirmation and re-nature.

Disposing in landfill and burning: not good option as prions leached into soil and groundwater.

Time: sheep reintroduced into a unihabited and supposedly decontaminated sheephouse after 16 yrs. They contracted scrapie which seems to be infectious after 16 yrs.

Protease degradation, radiation, fixation with formaldehyde: not possible due to size and structure of prion proteins.

NaOH: at high conc works as hydrolyses peptide bonds and destroys tertiary structure.

Question 21

Outline prion animal model

Answer 21

Rocky Mountain Laboratory (RML): mouse prion is inoculated into mouse brains. A true model of prion disease unlike other models (AD) where human mutant proteins expressed as prions naturally found in mouse species.

Characteristic spreading of prion that initiates in the hippocampus: leads to memory and learning deficits.
*) They do not burrow, this is due to motivational deficits.
*) Infected mice explore novel objects to same degree as familiar as cannot remember old object.

Astrocytosis is also present in the hippocampus.

Question 22

Outline prion animal model timescale of progression.

Answer 22

At 0 weeks: prion injection.
16 weeks: synapse loss.
18: protein synthesis decline.
20: neuronal loss (seen by spongiosis in brain)
24: terminally ill.

Question 23

Why are tg37 transgenic mouse models often used?

Answer 23

tg37 overexpresses PrP at approx 3 times wildtype levels.

They display the same characteristic features of synapse loss, neuronal loss, and behavioural deficits, in a stereotyped accelerated timescale. They typically become terminally sick by 12 weeks post inoculation.

Question 24

Why are repurposed drugs studied for prion disease?

Answer 24

Developing new drug = clinical trials etc therefore long process.
Repurposed useful as no need for above but could still be effective.

Trazodone: reduces spongiosis, increases time of survival and not toxic to pancreas.

Question 25

Prion proteins are found in both CNS and PNS but how are they transported?

Answer 25

Unknown:
Domino hypothesis - prion propagation occurs on neuronal cell
Streetcar hypothesis - nerve endings take up and transport PrP Sc.

Shim et al., 2022

Question 26

Chemical that shows therapeutic potential for treatment of prion

Answer 26

Lithium: neuroprotective role, reduces PrP Sc aggregation via autophagy (old cell parts).

Blocks GSK3 enzyme involved in prion based neurodegeneration and restores survival associated proteins (Wnt).

Administration of Li microemulsion = extended survival and reduced PrP Sc levels.

Question 27

Repurposed drug that shows therapeutic potential for treatment of prion disease.

Answer 27

Chlorpromazine: antipsychotic that crosses BBB. Inhibits PrP Sc formation by binding to human recombinant PrP.

Unfortunately failed to inhibit build-up in cell assay.

Question 28

Natural compound that shows therapeutic potential for treatment of prion disease.

Answer 28

Melatonin: hormone secreted by pineal gland that acts as antioxidant.

Induces autophagy and inhibits mitochondrial apoptosis.

Controversial as diff conc stimulate and inhibit prion proteins.

Question 29

How can immunotherapy be used to treat prion disease?

Answer 29

1) Active: vaccine
2) Passive: antibody

Due to structual diff between PrP C and PrP Sc the immune system differentiates between them. This phenomenon seen in terminal stage of mice.

Vaccine that seemed to work in deer did not work in elks.

Question 30

What do all prion treatments have in common?

Answer 30

None of them prevent disease being fatal

Question 31

Evidence that PrP Sc is infectious agent

Answer 31

1) Agent is too small to be bacteria or virus
2) Treatments that destroy nucleic acids doesn’t remove infectious nature.
3) Infectivity is proportional to conc of PrP Sc.
4) PrP Sc propagates in vitro by inducing misfolding of PrP C.

Soto and Satani 2011

Question 32

How does PrP Sc result in neurodegeneration?

Answer 32

Essentially this is not known

• Loss of biological function of PrP c?
• Toxicity of misfolded protein: evidence as other NDD are the same and where protein aggregates = brain most damaged and in vitro show misfolded are toxic to neurons in culture.

NB correlation is not causation = rare cases where disease has appeared with absence of detectable PrP Sc or converse where there is abundant deposition without neurodegeneration.

Soto and Satani, 2011

Question 32

What is thought to cause brain dysfunction in prion disease?

Answer 32

Brain dysfunction seems to begin at the synapse: synaptic abnormalities detected around time issues with burrowing observed. Abnormalities correlated with protein aggregation.

Soto and Satani 2011

Question 33

Mallucci lab findings

Answer 33

Brain dysfunction seems to begin at the synapse: synaptic abnormalities detected around time issues with burrowing observed.

Mallucci lab: deleted PrP C from neurons only and discorvered that early neuropathological and behavioural abnormalities did not occur. Despite extraneuronal accumulation of PrP Sc.

Concluded that propagation of non-neuronal PrP Sc is not pathogenic.

Soto and Satani 2011

Question 34

Microglia activation in prion disease

Answer 34

triggered by PrP Sc deposits and occurs before neuronal loss however cannot say whether this further causes degeneration or plays a role in slowing down degen by removing PrP Sc deposits.

Soto and Satani 2011

Question 35

How does PrP Sc cause cell death?

Answer 35

Not all mechanisms known.

One way: causes endoplasmic reticulum stress which triggers apoptosis

Soto and Satani 2011

Question 36

Link to AD, PD, HD

Answer 36

Same mechanisms and similar symptoms: some evidence that these diseases can be transmissible by infection.
* spreading pathology: dysfunction starts in discrete brain region and seems to spread in predictable manner.
IN vitro: mutant fibrils can transmit conformation of wild-type protein that drives it to become mutant.

Question 37

How long is the incubation period for prion diseases?

Answer 37

Incubation period: time from acquiring infective agent until symptoms show.

Kuru up to 50 years - Showing up in elderly members of the tribe born before 1959. As did not have mutation to cause sporadic prion disease. 20 years ago they were showing up with symptoms despite elimination of ritualistic cannibalism.

Wadsworth et al., 2008

Question 38

How do genes change susceptibility to prion disease?

Answer 38

Prnp has common polymorphism with either methionine or valine at residue 129.

Most sporadic cases occur in homozygous individuals.
All clinical cases reviewd have been homozygous for methionine.
Most elderly survivors of kuru are heterozygous.

Wadsworth et al., 2008

Question 39

Kuru symptoms vs CJD symptoms?

Answer 39

The central clinical feature of kuru is progressive cerebellar ataxia, and, in sharp contrast to sporadic CJD, dementia is a late and less prominent feature.

Wadsworth et al., 2008

Question 40

Outline evidence that Kuru originated from an individual with sporadic CJD.

Answer 40

Wadsworth et al., 2008

• Same transmission properties (PrP Sc only detectable in vCJD in lymphoreticular tissues).
• Same PrP Sc types as classical (2 and 3, there are 4 different types of PrP Sc with 1-2 seen in classical and 4 seen only in variant)
• Same molecular and neuropathological phenotype (in mice spongiform change restricted to thalamus and hypothalamus)

Question 41

PD evidence of transmissible

Answer 41

Frost and Diamond, 2010

PD (controversial)= patients underwent fetal transplant surgery and engrafted mesencephalic da neurons developed alpha-syn+ Lewy bodies.

• Indistinguishable from lesions in host (44-46).

Question 42

Tauopathies: evidence transmissible

Answer 42

Human tau induces misfolding of tau in mice. This doesn’t occur when tau is not present in infected material.

Frost and Diamond, 2010.