Creutzfeldt-Jakob’s disease or Prion Disease or Mad Cow disease

It belongs to a group of neurodegenerative diseases called Transmissible Spongiform Encephalopathy (TSE).

The infectious agents responsible for TSE are Prions, PrPc converted into PrPsc. They are generally hard to eliminate, and are extremely resistant to:

HeatRadiationDisinfection Protein digestion (degradation)

Creutzfeldt-Jakob’s Disease (Mad cow disease or Prion disease)

Severe Brain Atrophy in CDJ’s patient

www.scienceclarified.com

http://www.biologie.uni-duesseldorf.de

KURU: laughing death -Papua New Guinea (1957)

KURU and prion’s disease: similarities in the symptoms and in cerebellar ataxia

Models for conformational conversion of PrPc into PrPsc

Tissues where PrPsc accumulates

!!! vCJD is of oral origin, and PrPsc could take years before converting PrPc Role of oral mucosas in first accumulating then spreading PrPsc

Symptoms in Creutzfeldt-Jakob’s Disease

-It could take years (decades) before a carrier of prion disease will become fully symptomatic.

-Symptoms are characterized by cognitive decline, which may be fulminant and progress to akinetic mutism within few weeks.

-Cerebellar signs are evident (balance and coordination dysfunction -ataxia, changes in gait, rigid posture, and seizures).

mood swings depression anxiety memory lapses social withdrawal clumsiness or lack of coordination insomnia

As the disease rapidly progresses, patients with all forms of CJD generally experience:

* visual deterioration and eventual blindness * dementia * involuntary muscle contractions * muscle paralysis * slurred speech * difficulty swallowing * incontinence * coma

Cow affected by Bovine Spongiform Encephalitis

www.jonbarron.org

Diagnosis:

-Electroencephalography

-MRI

“Probable” CJD is based on the clinical symptoms.

-Post-mortem immunohistochemistry of PrPsc aggregates.

-Biopsy of the tonsils and, in 30% of the cases, of skeletal muscles can confirm CDJ. Determination of protease-K resistant form PrPsc.

ControlCJD

Absence of Protease-K-digested PrP in CJD used for diagnosis

Prion protein

Spongiform (intracellular vacuolation) change in the cortical gray matter of the brain, characteristic of TSEs and prions

aggregates

Walker et al.,

*Neuronal death

*Neuronal apoptosis

*Astrogliosis (as a cause or a consequence of inflammation)

*Protein misfolding and aggregation

*Precipitation of aggregates (proteinaceous material) both at an intracellular and extracellular level (amyloid plaques)

Features of TSEs and CJD

Deposition of fibrillar proteinacious material in Creutzfeldt-Jakob’s disease (prion disease)

Prion disease: Alteration in the prion protein lead to both intracellular and extracellular accumulation of amyloid aggregates, plaques, similar to those characteristic of AD, and positive to prion protein staining. Probably, replication and accumulation of the protease insensitive PrPsc results in fibril formation and plaque deposition.

Alzheimer’s Creutzfeldt-Jakob’s

Aguzzi A, Haass C. Science. 2003 Oct 31;302(5646):814-8. Review.

Epidemiology of Creutzfeldt-Jakob’s disease (CJD)

CJD is, among the Transmissible Spongiform Encephalopathies, the most diffuse one. CJD can be classified as

Sporadic sCJD: etiology not known, caused by both exogenous and endogenous factors, represents 85% of all the cases of CJD. In the United States, there are approximately 200 sporadic CJD cases per year.

Familial fCJD: caused by mutations in the gene for PrP (prion protein). 15% of CJD cases are inherited.

Iatrogenic iCJD: caused by the spreading of the infectious agent due to contaminated surgical tools, to the transplantation of tissues, or to the administration of pituitary hormones from deceased patients affected by the disease. 1% of CJD cases. Variant vCJD: caused by the transmission of Bovine Spongiform Encephalopathy (BSE) prion to humans (aka Mad Cow disease).

Incidence of BSE reported worldwide

Incidence of vCJD reported worldwide

The prion protein: functional domains and mutations causing inherited prion’s diseases

OR: not required for PrPc function, it might influence the change of conformation in PrPsc, as OR KO mice do not propagate the disease. Protects from apoptosis.

CC1: probably involved in protein internalization/trafficking. CC1 KO mice are viable and could develop the disease.

CC2: might work in concert with HC region, as partial deletion of either or the other domain, or ablation of one domain and partial deletion of the other accelerate the pathology in mice.

C-terminal: gene KO on H2, H3 or both domains leads to ataxia and neuron disease, BUT FAIL TO REPLICATE PRIONS. No transmission of disease from H2 KO and H3 KO to other animals. H2 and H3 might stabilize the conformation of the protein. C-terminal deletion prevents GPI anchoring of the protein, no development of the disease.

Functional domains of the prion protein

Prion Protein: domains and -helix structures

www.chemsoc.org

-PrPc contains 208 aminoacid residues and is abundantly expressed in neurons and glial cells-Signal peptide sequence-Octarepeats followed by a short Hydrophobic/toxic structure-The C-terminal portion of the protein is a globular structure that contains 3 -helix domain and 2 -helical domains. This domain folds rapidly and is extremely stable

Amyloid plaques in TSE

GSS disease Gerstmann-Straussler-Sheinker

disease

Kuru disease

Kuru disease

Aguzzi et al., Nat Rev Mol Cell Biol. 2007 Jul;8(7):552-61

Caughey and Byron, 2006 Nature 443-19

Physiologic role of PrPc

Antiapoptotic function:

PrPc KO mice are more susceptible to apoptosis.

Following ischemic injury, PrPc KO mice have increased infarct volume and increased caspase 3 activation.

Infarcts are larger in PrPc KO mice after ischemic injury

Levels of activated caspase 3 are increased in PrPc KO mice after ischemic injury

PrPc protects against oxidative stress

PrPc Ko mice are more susceptible to damage by H2O2

PrPc KO mice have reduced SOD activity

Brain of PrPc KO mice has increased levels of oxidated proteins, lipids, DNA.

PrPc also involved in maintaining mitochondrial integrity

Control

Prion’s infected

Mitochondrial structure is disrupted in Prion’s infected hamsters

PrPc maintains synaptic architecture and function

PrPc localizes mainly at the synaptic terminal

PrPc KO mice have impaired Glutamatergic and GABAergic transmission, as well as decreased LTP.

Synaptic loss is an early pathologic change in prion’s disease.

PrP stains as a flocculate/amorphous form at a synaptic level

PrPc physiological functions

How does the prion form? How does it propagate?

Sakaguchi

Mechanism of replication of PrPSc from PrPC

Nature of the prion

The prion is the minimum required infectious agent able to convert normal cellular prion protein PrpC into the scrapie PrPsc.

Is the prion a virus?NO, the prion is not a virus, as RNA and DNA material are totally absent in its composition, and the minimal molecular weight necessary for infectivity is ~2x105Da, so small to exclude the size of a virus.

Is the prion proteinaceous material?YES

The “unfortunate” goal of CDJ disease is to convert normal cellular prion protein PrPC into the scrapie form PrPsc.

This will result in

1-reduction of the form with -sheet conformation 2-accumulation of the form with -sheet conformation, which will form aggregates and deposit both at an intracellular and extracellular level.

Models for conformational conversion of PrPc into PrPsc

Sakaguchi

Different biochemical and structural properties between PrPC and PrPSc

The “protein-only” hypothesis

-In vitro data suggest that prion infectivity is achieved also de novo in the test tube.

-Propagation of conformationally changed yeast prions has been achieved.

-In vivo, prion protein null mice (PrP-/- or PrP KO) are resistant to prion infection, do not propagate prion infectivity after exposure to PrPSc, suggesting that PrPC is necessary to propagate the infectivity caused by PrPSc.

-Prion protein null mice do not propagate prion infection also when infected with infectious brain tissue. This implies that the protein PrPC ALONE (and not in combination with other cellular or putative viral factors) is sufficient to propagate the prion infectivity.

-Brain homogenates spiked with PrPSc and subjected to sonication and recovery amplify the infectious species which maintains infectivity when “transferred” to new tissue.

Prion Profile: Far left, infectious prion particles extracted from yeast cells. At right is an example of what yeast prions can do when mixed with an isolated yeast protein. These fibrils are essentially long series of prions all linked together after replicating many times in the presence of the protein.

Prion Particles Prion Particles+yeast protein

Fibrils of prion protein: self propagating mechanism?

Nat Med. 2004 Jul;10 Suppl:S63-7. Review.Soto and Castilla

PMCA Protein Misfolding Cyclic Amplification

PMCA: protein misfolding cyclic amplification: a technique to amplify prions in vitro

PMCA to generate synthetic strains of prions

Inoculation of synthetic strains of prions to generate animal models for prion’s disease

The “seeding:nucleation” model in Prion’s replication

PrPc localizes at the plasma membrane and undergoes internalization

Upon infection, PrP relocalizes from the plasma membrane to endocytic compartments

Where does the conversion from PrPc to PrPsc occur?

Transport of PrPsc?

Cell biology of PrP in scrapie infected cells

Prion protein scrapie accumulation and propagation from normal cellular Prion protein (PrPC to PrPsc)

Caughey and Byron, 2006 Nature 443-19

Dendrites of a clinical diseased intracerebrally inoculated with a synthetic PrP source.

Axon terminal from a 22CH scrapie-infected hamster

Dendrite of natural sheep scrapie brain

Membrane modifications are associated with PrP propagation

Post-translational modifications on Prion protein: Glycosylation and binding to metals

Caughey and Byron, 2006 Nature 443-19

GPI anchor as a target for the treatment of CJD?

Inhibition of GPI-anchor could reduce the amount of PrPC at the plasma membrane, reducing propagation of the prion

Caughey and Byron, 2006 Nature 443-19

Can cytosolic aggregates seed for propagation?

PrP aggregates can be found at an intracellular level

Cytosolic PrPC can form intracellular aggregates in presence of extracellular PrP fibrils

Induced intracellular aggregates can be transmitted to progeny

Formation of aggregates can also be induced in neighboring adjacent cells

Direct transfer of prion cytosolic aggregates from donor to recipient cells

PrPsc is toxic both when extracellular and intracellular

MECHANISM?

Loss of physiologic function or gain of toxic function?

PrPc KO mice are viable, develop normally and have no severe pathologies observed later in life.

PrPc KO do not develop the disease as they do not propagate the formation of PrPsc

Gain of toxic function rather than loss of physiological function

A role for Autophagy?

Can autophagy target the degradation of PrPsc aggregates and be used in the therapy of prion’s disease?

Accumulation of degenerating lysosomes and other organelles at the dark synapse in human vCJD brain

Main steps in macroautophagy

Autophagic vacuoles form at the synapse of human vCJD brain

Autophagic vacuoles formed in scrapie-infected hamster brain

Autophagy-inducing drugs can regulate autophagy in scrapie infected cells

Autophagy-inducing drugs can reduce the PK resistant load of PrPsc aggregates

Autophagy as an approach to reduce prion infection

Small molecules used to regulate autophagy in neurodegenerative diseases

Approaches for the treatment of CJD

-Use of autophagy-inducing molecules

Use of molecules that associate with PrPC and interfere with the capacity of PrPSc to bind and to propagate the infectious principle.

-GPI inhibitors

-PrPSc antibodies: problems in achieving the right level of immunization(prion protein is a self antigen, and immunesystem is involved in the replication of the prion agent and its ultimate access to the CNS).

Active immunization with recombinant protein has relatively modest therapeutic effects in mice.

Passive immunization effective within a month after exposure to the contaminant. However, too expensive and not effective if performed in advanced stages of the disease, when the animal is symptomatic.