Distinctive properties of plaque-type dura mater graft-associated
Creutzfeldt–Jakob disease in cell-protein misfolding cyclic amplification
Atsuko Takeuchi, Atsushi Kobayashi, Piero Parchi, Masahito Yamada, Masanori
Morita, Shusei Uno and Tetsuyuki Kitamoto
Abstract
There are two distinct subtypes of dura mater graft-associated
Creutzfeldt–Jakob disease (dCJD) with methionine homozygosity at codon 129 of
the PRNP gene. The majority of cases is represented by a non-plaque-type
(np-dCJD) resembling sporadic CJD (sCJD)-MM1 or -MV1, while the minority by a
plaque-type (p-dCJD). p-dCJD shows distinctive phenotypic features, namely
numerous kuru plaques and an abnormal isoform of prion protein (PrPSc)
intermediate in size between types 1 and 2. Transmission studies have shown that
the unusual phenotypic features of p-dCJD are linked to the V2 prion strain that
is associated with sCJD subtypes VV2 or -MV2. In this study, we applied protein
misfolding cyclic amplification (PMCA) using recombinant human prion protein as
a substrate and demonstrated that p-dCJD prions show amplification features that
are distinct from those of np-dCJD. Although no amplification of np-dCJD prions
was observed with either 129 M or 129 V substrate, p-dCJD prions were
drastically amplified with the 129 V substrates, despite the PRNP codon 129
incompatibility between seed and substrate. Moreover, by using a type 2
PrPSc-specific antibody not recognizing PrPSc in p-dCJD, we found that type 2
products are generated de novo from p-dCJD prions during PMCA with the 129 V
substrates. These findings suggest that our cell-PMCA is a useful tool for
easily and rapidly identifying acquired CJD associated with the transmission of
the V2 CJD strain to codon 129 methionine homozygotes, based on the preference
for the 129 V substrate and the type of the amplified products.
all iatrogenic cjd is, is sporadic cjd, until the iatrogenic event is
traced back, discovered, documented, put into the academic domain and finally
public domain. ...tss
These findings support the view of the co-occurrence of multiple PrPres
fragments within MM1 prions rather than the co-occurrence of multiple prion
strains within the same individual.
In conclusion, the present study, together with evidence from other
groups,11,12 suggests that the co-occurrence of multiple PrPres fragments within
a single sCJD patient is a universal phenomenon. These findings show that the
conventional typing of PrPres merely represents the predominant PrPres
subpopulation among multiple co-existing PrPres fragments. Besides the general
co-occurrence of multiple PrPres fragments, the condition of PK digestion easily
affects the size of PrPres.27,28 Indeed, insufficient PK digestion can generate
type 1 PrPres-specific antibody–reactive fragments in the sCJD patients
classified as type 2.29 Furthermore, it is possible that the conventional
Western blot analysis fails to detect type 2 PrPres in sCJD-MM1+2 cases showing
very focal perivacuolar PrP deposition in the brain.9 These confusing aspects of
PrPres typing question the validity of the conventional molecular typing system.
For a precise classification, it may be appropriate that the neuropathological
phenotyping [synaptic (SY), perivacuolar (PV), plaque (PL), or patchy plaque
(PP)] be combined with the molecular typing [eg, sCJD-MM1(+2)/SY+PV] the sCJD-MM
patient showing synaptic + perivacuolar PrP deposition, but not type 2 PrPres in
the conventional Western blot analysis.
Even though experiments in rodents performed some years ago demonstrated
the phenomenon of multiple PrPSc types within one animal [21], incongruity
regarding the frequency of co-occurrence of different PrPSc types in sCJD
patients still endures. Initial studies suggested that this is a relatively rare
event, occurring in less than 5% of patients [8]. This low incidence may, at
least partially, be attributed to the fact that these analyses are routinely
performed on a limited range of distinct brain regions per patient [8,13]. The
fact that region-specific presence of distinct PrPSc types may occur in sCJD was
highlighted by a publication investigating ten defined regions within the
central nervous system in 14 patients with sCJD [16]. The authors found more
than one PrPSc type in five individuals and hypothesized that the co-occurrence
of more than one PrPSc type could be the rule, rather than the exception, if the
entire central nervous system was investigated. In our analysis of nine distinct
central-nervous-system regions in 50 patients with sCJD using a standardized
protocol [22], we detected more than one PrPSc type in nine individuals. The
observation that most patients harboring two PrPSc types are codon-129
methionine/valine heterozygotes stresses the significance of this polymorphism
in the replicative cycle of PrPSc.
Coexistence of multiple PrPSc types in individuals with Creutzfeldt-Jakob
disease
Magdalini Polymenidou, Katharina Stoeck, Markus Glatzel, Martin Vey, Anne
Bellon, and Adriano Aguzzi
Summary
Background The molecular typing of sporadic Creutzfeldt-Jakob disease (CJD)
is based on the size and glycoform ratio of protease-resistant prion protein
(PrPSc), and on PRNP haplotype. On digestion with proteinase K, type 1 and type
2 PrPSc display unglycosylated core fragments of 21 kDa and 19 kDa, resulting
from cleavage around amino acids 82 and 97, respectively.
Methods We generated anti-PrP monoclonal antibodies to epitopes immediately
preceding the differential proteinase K cleavage sites. These antibodies, which
were designated POM2 and POM12, recognise type 1, but not type 2, PrPSc.
Findings We studied 114 brain samples from 70 patients with sporadic CJD
and three patients with variant CJD. Every patient classified as CJD type 2, and
all variant CJD patients, showed POM2/POM12 reactivity in the cerebellum and
other PrPSc-rich brain areas, with a typical PrPSc type 1 migration
pattern.
Interpretation The regular coexistence of multiple PrPSc types in patients
with CJD casts doubts on the validity of electrophoretic PrPSc mobilities as
surrogates for prion strains, and questions the rational basis of current CJD
classifications.
snip...
Discussion
The discovery of heritable polymorphic PK cleavage sites in PrPSc has been
used for the molecular classification of CJD cases.11,13,14,17 In concert with
the codon 129 PRNP haplotype, the different PrPSc types correlate with distinct
disease phenotypes. Most patients with the MM1 or MV1 subtype present with
so-called classic CJD, and show rapid progressive dementia, early myoclonus,
visual disturbances including cortical blindness, and a disease duration of
approximately 4 months. By contrast, patients with the MV2 or VV2 subtype show
an atypical disease course, with a longer disease duration (6–18 months), early
ataxia, predominant extrapyramidal symptoms, and late-onset dementia.18
Because of the limited resolution of conventional western blot systems, the
coexistence of PK-digested PrPSc bands at 19 kDa and 21 kDa is not recognisable
by conventional antibodies binding to both PrPSc types, including POM1. By
mixing brain homogenates that contain solely type 1 or type 2 PrPSc, we have
found that 3F4-based or POM1-based western blots consistently fail to detect
type 1 PrPSc whenever it represents less than 30–40% of total PrPSc. We
addressed these problems by developing a set of antibodies binding Nproximal
epitopes of PrP, which recognise PK-digested type 1 PrPSc (cleaved at amino acid
82 or upstream), but not type 2 PrPSc (cleaved at amino acids 97 or 86).
On investigation of 70 sporadic CJD cases, we determined that 50% of the
patients formerly classified as CJD type 2 had low but detectable amounts of
PrPSc type 1 in their cerebral cortex. Investigations of additional brain areas
revealed that significant amounts of PrPSc type 1 coexist with type 2 in at
least some areas of all patients classified as having type 2 disease.
Co-occurrence of CJD types in the same brain has been previously reported in
approximately 30% of the tested cases.24 Using unambiguous analytical tools, we
have established that type 2 PrPSc does not exist independently from type 1
PrPSc, at least not in the Swiss CJD patients included in our study.
Furthermore, using our type 1-specific antibodies, we found type 1 PrPSc
content in three cortical samples of vCJD patients.
Glycotyping of blots incubated with 3F4/POM1 versus POM2/POM12 showed no
significant differences in glycoform ratios, neither for the same groups
incubated with different antibodies, nor for the two PrPSc groups, in accordance
with previously published data.33 The homogeneity of glycoforms suggests that
our collective is comparable to those investigated in other countries, despite
the peculiarities of Swiss CJD epidemiology.34
The results presented here do not question the validity of the established
correlations between PrPSc types and clinical findings. The coincident presence
of PrPSc type 1 may not alter the expected clinical outcome of patients with
predominant PrPSc type 2 deposition. However, the existence of heritable PrPSc
types implies that strainspecific characteristics of prions are enshrined in the
conformation of PrPSc, and that the different core fragment sizes of PK-digested
PrPSc may be used as surrogates of such conformational variations. In the light
of the data presented here, the strength of these arguments becomes somewhat
questionable.
Why did the co-existence of type 1 and type 2 PrPSc go often undetected in
the past, despite its ubiquitous presence? Maybe the 21 kDa band characteristic
of type 1 PrPSc is easily obfuscated by the simultaneous occurrence of the
neighbouring 19 kDa type 2 band and of the monoglycosylated PrPSc band. We
tested this hypothesis by ascertaining the detectability threshold of type 1
PrPSc in mixtures of pure type 1 and type 2 samples. Indeed, we found that type
1 PrPSc becomes undiscernible as soon as 40% or more type 2 PrPSc is present.
This observation provides a plausible explanation for the failure to appreciate
the invariable coexistence of PrPSc types in previous reports.
To determine whether a similar phenomenon occurs in patients classified as
type 1 would be interesting. However, this issue is impossible to address at
present, as there is no CJD type-2-specific antibody. The fact that the type 1
coexistence is also apparent in cortical samples from CJD patients, indicates
that the currently described phenomenon might be a more general one. The above
results set the existing CJD classifications into debate and introduce
interesting questions about human CJD types. For example, do human prion types
exist in a dynamic equilibrium in the brains of affected individuals? Do they
coexist in most or even all CJD cases? Is the biochemically identified PrPSc
type simply the dominant type, and not the only PrPSc species?
Evidence for human transmission of amyloid-β pathology and cerebral amyloid
angiopathy
07 02:27 AM
Terry S. Singeltary Sr. said:
re-Evidence for human transmission of amyloid-β pathology and cerebral
amyloid angiopathy
2015-12-07 02:27 AM
Terry S. Singeltary Sr. said: re-Evidence for human transmission of
amyloid-β pathology and cerebral amyloid angiopathy
Nature 525, 247?250 (10 September 2015) doi:10.1038/nature15369 Received 26
April 2015 Accepted 14 August 2015 Published online 09 September 2015 Updated
online 11 September 2015 Erratum (October, 2015)
I would kindly like to comment on the Nature Paper, the Lancet reply, and
the newspaper articles.
First, I applaud Nature, the Scientist and Authors of the Nature paper, for
bringing this important finding to the attention of the public domain, and the
media for printing said findings.
Secondly, it seems once again, politics is getting in the way possibly of
more important Transmissible Spongiform Encephalopathy TSE Prion scientific
findings. findings that could have great implications for human health, and
great implications for the medical surgical arena. but apparently, the
government peer review process, of the peer review science, tries to intervene
again to water down said disturbing findings.
where have we all heard this before? it’s been well documented via the BSE
Inquiry. have they not learned a lesson from the last time?
we have seen this time and time again in England (and other Country’s) with
the BSE mad cow TSE Prion debacle.
snip...see full Singeltary Nature comment here;
see Singeltary comments to Plos ;
Subject: 1992 IN CONFIDENCE TRANSMISSION OF ALZHEIMER TYPE PLAQUES TO
PRIMATES POSSIBILITY ON A TRANSMISSIBLE PRION REMAINS OPEN
BSE101/1 0136
IN CONFIDENCE
CMO
From: . Dr J S Metiers DCMO
4 November 1992
TRANSMISSION OF ALZHEIMER TYPE PLAQUES TO PRIMATES
1. Thank you for showing me Diana Dunstan's letter. I am glad that MRC have
recognised the public sensitivity of these findings and intend to report them in
their proper context. 'This hopefully will avoid misunderstanding and possible
distortion by the media to portray the results as having more greater
significance than the findings so far justify.
2. Using a highly unusual route of transmission (intra-cerebral injection)
the researchers have demonstrated the transmission of a pathological process
from two cases one of severe Alzheimer's disease the other of
Gerstmann-Straussler disease to marmosets. However they have not demonstrated
the transmission of either clinical condition as the "animals were behaving
normally when killed". As the report emphasises the unanswered question is
whether the disease condition would have revealed itself if the marmosets had
lived longer. They are planning further research to see if the conditions, as
opposed to the partial pathological process, is transmissible.
what are the implications for public health?
3. The route 'of transmission is very specific and in the natural state of
things highly unusual. However it could be argued that the results reveal a
potential risk, in that brain tissue from these two patients has been shown to
transmit a pathological process. Should therefore brain tissue from such cases
be regarded as potentially infective? Pathologists, morticians, neuro surgeons
and those assisting at neuro surgical procedures and others coming into contact
with "raw" human brain tissue could in theory be at risk. However, on a priori
grounds given the highly specific route of transmission in these experiments
that risk must be negligible if the usual precautions for handling brain tissue
are observed.
1
92/11.4/1.1
BSE101/1 0137
4. The other dimension to consider is the public reaction. To some extent
the GSS case demonstrates little more than the transmission of BSE to a pig by
intra-cerebral injection. If other prion diseases can be transmitted in this way
it is little surprise that some pathological findings observed in GSS were also
transmissible to a marmoset. But the transmission of features of Alzheimer's
pathology is a different matter, given the much greater frequency of this
disease and raises the unanswered question whether some cases are the result of
a transmissible prion. The only tenable public line will be that "more research
is required’’ before that hypothesis could be evaluated. The possibility on a
transmissible prion remains open. In the meantime MRC needs carefully to
consider the range and sequence of studies needed to follow through from the
preliminary observations in these two cases. Not a particularly comfortable
message, but until we know more about the causation of Alzheimer's disease the
total reassurance is not practical.
J S METTERS Room 509 Richmond House Pager No: 081-884 3344 Callsign: DOH
832 llllYc!eS 2 92/11.4/1.2
>>> The only tenable public line will be that "more research is
required’’ <<<
>>> possibility on a transmissible prion remains
open<<<
O.K., so it’s about 23 years later, so somebody please tell me, when is
"more research is required’’ enough time for evaluation ?
Self-Propagative Replication of Ab Oligomers Suggests Potential
Transmissibility in Alzheimer Disease
*** Singeltary comment PLoS ***
Alzheimer’s disease and Transmissible Spongiform Encephalopathy prion
disease, Iatrogenic, what if ?
Posted by flounder on 05 Nov 2014 at 21:27 GMT
Sunday, November 22, 2015
*** Effect of heating on the stability of amyloid A (AA) fibrils and the
intra- and cross-species transmission of AA amyloidosis Abstract
Amyloid A (AA) amyloidosis is a protein misfolding disease characterized by
extracellular deposition of AA fibrils. AA fibrils are found in several tissues
from food animals with AA amyloidosis. For hygienic purposes, heating is widely
used to inactivate microbes in food, but it is uncertain whether heating is
sufficient to inactivate AA fibrils and prevent intra- or cross-species
transmission. We examined the effect of heating (at 60 °C or 100 °C) and
autoclaving (at 121 °C or 135 °C) on murine and bovine AA fibrils using Western
blot analysis, transmission electron microscopy (TEM), and mouse model
transmission experiments. TEM revealed that a mixture of AA fibrils and
amorphous aggregates appeared after heating at 100 °C, whereas autoclaving at
135 °C produced large amorphous aggregates. AA fibrils retained antigen
specificity in Western blot analysis when heated at 100 °C or autoclaved at 121
°C, but not when autoclaved at 135 °C. Transmissible pathogenicity of murine and
bovine AA fibrils subjected to heating (at 60 °C or 100 °C) was significantly
stimulated and resulted in amyloid deposition in mice. Autoclaving of murine AA
fibrils at 121 °C or 135 °C significantly decreased amyloid deposition.
Moreover, amyloid deposition in mice injected with murine AA fibrils was more
severe than that in mice injected with bovine AA fibrils. Bovine AA fibrils
autoclaved at 121 °C or 135 °C did not induce amyloid deposition in mice. These
results suggest that AA fibrils are relatively heat stable and that similar to
prions, autoclaving at 135 °C is required to destroy the pathogenicity of AA
fibrils. These findings may contribute to the prevention of AA fibril
transmission through food materials to different animals and especially to
humans.
Purchase options Price * Issue Purchase USD 511.00 Article Purchase USD
54.00
*** Transmission of Creutzfeldt-Jakob disease to a chimpanzee by electrodes
contaminated during neurosurgery ***
Gibbs CJ Jr, Asher DM, Kobrine A, Amyx HL, Sulima MP, Gajdusek DC.
Laboratory of Central Nervous System Studies, National Institute of Neurological
Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892.
Stereotactic multicontact electrodes used to probe the cerebral cortex of a
middle aged woman with progressive dementia were previously implicated in the
accidental transmission of Creutzfeldt-Jakob disease (CJD) to two younger
patients. The diagnoses of CJD have been confirmed for all three cases. More
than two years after their last use in humans, after three cleanings and
repeated sterilisation in ethanol and formaldehyde vapour, the electrodes were
implanted in the cortex of a chimpanzee. Eighteen months later the animal became
ill with CJD. This finding serves to re-emphasise the potential danger posed by
reuse of instruments contaminated with the agents of spongiform
encephalopathies, even after scrupulous attempts to clean them.
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&list_uids=8006664&dopt=Abstract
the warning shots fired over the bow of the boat that were never heard ;
PITUITARY EXTRACT
This was used to help cows super ovulate. This tissue was considered to be
of greatest risk of containing BSE and consequently transmitting the disease...
NON-LICENSED HUMAN TISSUE DEVICES WERE NOT COMMERCIALLY AVAILABLE
snip...
I was quite prepared to believe in unofficial pituitary hormones, also in
the 1970's, whether as described by Dr. Little, or in other circumstances, for
animal use.
snip...
The fact that there were jars of pituitaries (or extract) around on shelves
is attested by the still potent 1943 pituitaries, described in Stockell Hartree
et al. (J/RF/17/291) which had come from the lab. at Mill Hill. Having taken the
trouble to collect them, they were not lightly thrown out...
3. The extraction is from a pool of pituitary glands collected from
abbatoirs and the process used is unlikely to have any effect on the BSE agent.
Hormones extracted from human pituitary glands have been responsible for a small
number of Creutzfeldt Jacob disease in man.
SEE LOOPHOLE ;
SEE LOOPHOLE SHOULD BE CLOSED ;
snip...see at bottom ;
Friday, January 10, 2014
vpspr, sgss, sffi, TSE, an iatrogenic by-product of gss, ffi, familial type
prion disease, what it ???
Tuesday, August 4, 2015
*** FDA U.S. Measures to Protect Against BSE ***
*** now, from all the consumption and exposure above, now think iatrogenic
cjd tse prion at a hospital near you, what if?
Thursday, August 13, 2015
Iatrogenic CJD due to pituitary-derived growth hormone with genetically
determined incubation times of up to 40 years
Monday, August 17, 2015
*** FDA Says Endoscope Makers Failed to Report Superbug Problems OLYMPUS
*** I told Olympus 15 years ago about these risk factors from endoscopy
equipment, disinfection, even spoke with the Doctor at Olympus, this was back in
1999. I tried to tell them that they were exposing patients to dangerous
pathogens such as the CJD TSE prion, because they could not properly clean them.
even presented my concern to a peer review journal GUT, that was going to
publish, but then it was pulled by Professor Michael Farthing et al... see ;
Thursday, September 10, 2015
25th Meeting of the Transmissible Spongiform Encephalopathies Advisory
Committee Food and Drug Administration Silver Spring, Maryland June 1, 2015
Sunday, January 17, 2016
Of Grave Concern Heidenhain Variant Creutzfeldt Jakob Disease
Saturday, February 13, 2016
The Risk of Prion Infection through Bovine Grafting Materials in dentistry
kind regards, terry