Case # 68
Shan Kuang
Julian Lopez
Maria De Leon
Case Summary
Patient History:
 54 year old retired US Air Force Pilot
 Spent several years in Guam and Vietnam, and worked for 23
years as a pilot
 His eating habits were conventional, and he had no history of
alcohol use
 He has never been transfused, had no history of tick bites, and
had no known risk factors for HW
The patient’s problem:
 Chronic neurologic disorder
 Initial symptoms: forgetfulness, subtle behavioral changes,
headaches, and fatigue
 Later symptoms: disequilibrium, wide-based gait, double vision
 He died 6 months after the onset of symptoms
Key information pointing to diagnosis
Part 1
The disease may not have a bacterial cause:
 He was never febrile, indicating he had no
inflammatory immune response
 Treatment with antibiotics (penicillin and
tetracycline) against lyme disease and Rocky
Mountain Spotted Fever were ineffective
 Serology test for Borrelia burgdorferi and Rocky
Mountain Spotted Fever were negative
 Cultures were normal, and he was negative for
venereal disease
Key information pointing to diagnosis
Part 2
Clues to the disease may be found in the neurological tests:
 Computed tomogram scan revealed atrophy in the cerebrum and
cerebellum, which correlate with his behavioral, mental, and
physical changes
 He had no mass lesions, ruling out tumors and stroke
 Tests were done on his cerebrospinal fluid (CSF)
 Some viruses can cause neurological disorders
 HIV with dementia, Measles/rubeola with subacute sclerosing
panencephalitis, JC virus with progressive multifocal
leukoencephalopathy (myelin degradation)
 The results of the tests were normal, and he was HIV negative
 No biopsy was done, but an autopsy showed extensive
spongiform changes in the cerebrum, cerebellum, and basal
ganglia
Diagnosis
 Sporadic occurrence of classic Creutzfeldt-Jakob Disease due to
spontaneous transformation of normal prion proteins to abnormal
prions
Clinical and Pathologic Characteristics
Characteristic
Median age at death
Median duration of illness
Clinical signs and symptoms
Periodic sharp waves on
electroencephalogram
"Pulvinar sign" on MRI*
Presence of "florid plaques" on
neuropathology
Immunohitochemical analysis of brain
tissue
Presence of agent in lymphoid tissue
Source: CDC
Distinguishing Classic CJD from Variant CJD
Classic CJD
Variant CJD
68 years
28 years
4-5 months
13-14 months
Dementia; early neurologic signs Prominent psychiatric/behavioral symptoms;
painful dyesthesiasis; delayed neurologic signs
Often present
Often absent
Not reported
Rare or absent
Present in >75% of cases
Present in large numbers
Variable accumulation
Marked accumulation of protease-resistance
prion protein
Readily detected
Not readily detected
Increased glycoform ratio on immunoblot Not reported
analysis of protease-resistance prion
protein
Marked accumulation of protease-resistance
prion protein
Source: Adapted from Belay E., Schonberger L. Variant Creutzfeldt-Jakob Disease and Bovine Spongiform Encephalopathy. Clin Lab Med 2002;22:849-62.
*An abnormal signal in the posterior thalami on T2- and diffusion-weighted images and fluid-attenuated inversion recovery sequences on brain magnetic resonance imaging
(MRI); in the appropriate clinical context, this signal is highly specific for vCJD
Normal conformer
Rogue conformer
http://www.cmpharm.ucsf.edu/cohen/pages/welcome.html
Microbiology of Prions
 Prion diseases (transmissible spongiform encephalopathies) are infectious and




cause fatal neurodegenerative disorders in humans and animals
The disease is caused by proteinaceous infectious particles called prions
The abnormal prion form PrPsc causes the conformation change of normal
prions PrPc in the host to the abnormal form
The diseases can occur sporadically, through familial transmission, or
horizontal transmission
Prion diseases:
 Scrapie—sheep, goats
 Bovine spongiform encephalopathy—cattle
 Transmissable mink encephalopathy—mink
 Creutzfeldt—jakob disease—man, sporadic , familial, transmission;
 Gerstmann-Streussler disease—man, familial
 Fatal familial insomnia—man, familial
 Kuru—man, Fore people of New Guinea
 Chronic Wastings Disease—Elk, Deer
Pathogenesis of Prions Disease
 PrPC is a glycosylphosphatidylinositol-anchored glycoprotein present in
all cell types
 It has an unstructured N-terminal portion and a mostly α-helical folded C-
terminus
 It is expressed in all cell types, and is highly conserved among mammals
 Its function is not well known
 PrPsc consists of various strains with various incubation periods and
lesion profiles
 The C-terminal is folded into a tightly packed highly protease resistant
domain enriched in β-structure
 It is resistant to heat, ultra-violet light, and formaldehyde
 The secondary lymphoid organs play a major role in replication
 Normal PrPc upon exposure to its abnormal isoform PrPsc converts to the
abnormal form, which aggregates and accumulates in the brain causing
degeneration and amyloid buildup
 How PrPsc causes the conformation, invade the CNS, and causes
degeneration is not well understood
 Scrapie-infected transgenic mice with a soluble PrPc did not develop prion
disease
 Neither PrPc nor PrPsc are very immunogenic in the species normally
used to prepare antibodies
Primary Research Article Contributing
to the Understanding of this Disease
 Reinald Pamlona, Alba Naudi, et. al, 2008, Increased oxidation,




glycoxidation, and lipoxidation of brain proteins in prion disease,
Free Radical Biology & Medicine, 45: 1159-1166
PrPsc may be directly involved in the pathogenesis of because it is
temporally and anatomically correlated with the neuropathology
 However, PrPsc may be involved in more complex effects
involving other molecules
Neurodegeneration may result as a consequence of loss of
function of PrPc or a toxic gain of function from PrPsc
The study aims to see whether oxidative stress is a mechanism
involved with neurodegeneration in prions disease
Experimental Setup:
 Brains of 6 patients with sporadic CJD, age matched
 Brains of syrian hamsters inoculated with 263K scrapie prions
 Gas chromatography-mass spectrometry was used to analyze protein
oxidation products and fatty acids
 SDS-PAGE was used to determine ERK1/2 and p38 activation
Results
Fig 1. Protein oxidation products
are higher in brains affected by
prion disease in humans. Similar
results were shown in scrapieaffected Syrian hamsters. There is a
positive correlation with GSA and
MDAL and CML, and MDAL and
CML (figure not shown).
Fig 3. Modest yet significant
changes in fatty acid composition is
associated with prion disease in
humans. Lower intensity changes
were found in scrapie-infected
hamsters
Fig 5. There is significant
negative correlation between
protein oxidation product
GSA and poly unsaturated
fatty acids in the human
brain. Similar correlations
were found in PUFA and
MDAL and PUFA and CML.
Fig 6. There is increased phosphorylation of ERK1/2 and p38 in scrapie-affected hamster
brains.
Conclusion
 The amount of specific chemically defined oxidation, lipoxidtion, and glycoxidation
products are higher in brains from both CJD affected humans and scrapie-infected
hamsters
 The slight changes in fatty acids may be due to a homeostatic response to reduce
oxidative damage, a loss of function in homeostatic systems, or a higher rate of lipid
damage that cannot be fixed
 The correlation between different oxidation products suggest an elevated
underlying oxidative stress that acts on the different possible oxidative routes and
different available oxidizable substrates
 The negative correlation of oxidation products and PUFA suggests a wider
association of fatty acid composition and protein oxidative damage
 Some studies suggest that oxidative modifications lead to loss of protein function,
while other studies suggest that oxidative stress induces a cell signaling cascade
that increases reactive oxygen species (ROS) and causes over activation of routes
that lead to neurodegeneration
 The increased phosphorylation of ERK1/2 and P38 suggests the latter, but more
needs to be examined
 The results suggest that patients with compromised antioxidant defenses may have
accelerated clinical progression of prions disease
Diagnostic Tests for Prion Disease
 Currently there is no single diagnostic test for CJD
 Neurological examinations, standard diagnostic tests, electroencephalogram,
and magnetic resonance imaging are used to rule out treatable forms of
dementia
 A biopsy or autopsy is used to confirm diagnosis of CJD
 Biopsy poses some risk to the surgeon and patient, and the results are
dependent on the tissue obtained being affected
 Laboratory testing:
 Bioassays and cell assays: the subject is injected with the test sample
 Conformation-dependent immunoassay: selective precipitation of PrPsc
followed by immunodetection
 Protein-misfolding cyclic amplification (PCMA): works similarly like PCR, it
allows conversion of PrPc to PrPsc
 Western blotting: antibodies are added after treatment with protease
 Immunohistochemistry: look for characteristic deposition patterns
 Paraffin embedded tissue (PET): technique that combines WB and IHC
 New techniques are being developed
 Amyloid seeding assay (ASA): prions are capable to seed amyloid
formation
 Monoclonal antibodies 1.5D7 and 1.6F4 have higher affinity to PrPsc and
lower affinity to PrPc than current antibodies being used
Prevention of disease
 85% cases of classical CJD is sporadic, 5-10% is genetic
 Prevention in a hospital setting:
 Wash hands and exposed skin, and cover cuts and wounds
 Wear surgical gloves and facial protection
 Soak instruments in undiluted chlorine bleach, use autoclave to sterilize
 Be careful of transmission modes
 Vertical transmission
 Surgery: dura mater grafts, exposure to brain tissue and CSF, and transplanted
corneas, implantation of unsterilized electrodes in brain
 Contaminated human growth hormones and blood (people who reside in a
country that is known for BSE for more than 3 months cannot donate blood)
 Prevention of BSE and vCJD
 The FDA has prohibited the slaughter of downer-cows (non-ambulatory)
 Beef is recalled from cattle slaughtered in plants confirmed with BSE positive
cows
 In 2009 the FAD and CIFA (Canada) have enhanced a feed-ban on animal feeds,
pet foods, and fertilizers
Epidemiology and Threats
•Approximately 1 in 1 million
people are affected with CJD
worldwide; recently there are fewer
than 300 cases of CJD per year in
the U.S.
•Most of classical CJD cases are
sporadic due to spontaneous
mutation of the prion protein
•Some cases were acquired through
transmission (fewer than 1%)
Source: CDC
•The first case of BSE was in the 1970s in the UK
•Cattle was fed meat-and-bone meal contaminated with BSE-infected products
•In 2008 there were more than 484,500 cases of BSE in the UK
•As of 2010 there were 21 cases of BSE in North America
•vCJD is hypothesized to occur due to ingestion of BSE prion infected meat
•The first case occurred in the UK in 1996
•In 2006 there were 200 cases worldwide, 164 in the UK and 3 in the U.S.
•There are no treatments for prion disease, only drugs to alleviate symptoms
•Opiates, clonazepam and sodium valproate to relieve myoclonus
Take home message
 Sporadic CJD involves a chronic neurologic disorder with initial






symptoms of forgetfulness, subtle behavioral changes, headaches, and
fatigue. Later symptoms include disequilibrium, wide-based gait, and
double vision. It is a terminal disease.
The main theory is that classic CJD is caused by abnormal prions that
convert the normal host prions to its abnormal isotype which
accumulates in the brain and causes neurodegeneration.
The main diagnostic tests for prions are PCMA, IHC, and WB. But these
techniques are limited because the antibodies used are not specific
for PrPsc.
There is a lack of treatment in this disease.
Banning contaminated food prevents vCJD and careful hospital
procedures prevents iatrogenic transmission. However most of the
disease is acquired genetically or sporadically.
There are limitations in testing and knowing the structure of PrPsc
because they are protease resistant and similar to PrPc. Hence it is
hard to know the pathology of prions disease.
Various strains of prions cause different morphology in the brain.
References
 http://health.nih.gov/topic/CreutzfeldtJakobDisease
 http://www.cdc.gov/ncidod/dvrd/vcjd/index.htm
 http://web.uct.ac.za/depts/mmi/jmoodie/neurol2.html#Retrovirus Disease
 Cordes, H., Bergstrom, A.L., Ohm, J., Laursen, H., and Heegaard, P.M. (2008)
Characterisation of new monoclonal antibodies reacting with prions from both
human and animal brain tissues. Journal of Immunology Methods, 337, 106-120
 Maurizio Polano, Alpan Bek, Federico Benetti, Marco Lazzarino, and Giuseppe
Legname. (2009) Structural Insights into Alternate Aggregated Prion Protein
Forms. J. Mol. Biol., 393, 1033-1042
 Nuvolone, M., Aguzzi A., and Heikenwalder, M. (2009) Cells and prions: A license
to replicate. FEBS Letters, 583, 2674-2684
 Pamplona, R., Naudi A., Gavin,R., Pastrana, M.A., Sajnani, G., Ilieva, E.V., Antonio del
Rio, J., Portero-Otin,M., Ferrer, I., and Requena, J.R. (2008) Increased oxidation,
glycoxidation, and lipoxidation of brain proteins in prion disease. Free Radical
Biology & Medicine, 45, 1159-1166
 Willey, Joanne et al. 2008. Prescott, Harley, & Klein’s Microbiology, 7th ed.
McGraw Hill, NY