Tuesday, September 1, 2015

Evidence for α-synuclein prions causing multiple system atrophy in humans with parkinsonism

 Evidence for α-synuclein prions causing multiple system atrophy in humans with parkinsonism

Stanley B. Prusinera,b,c,1, Amanda L. Woermana, Daniel A. Mordesd, Joel C. Wattsa,b,2, Ryan Rampersauda, David B. Berrya, Smita Patela, Abby Oehlere, Jennifer K. Lowef, Stephanie N. Kravitzf, Daniel H. Geschwindf,g, David V. Gliddenh, Glenda M. Hallidayi, Lefkos T. Middletonj, Steve M. Gentlemank, Lea T. Grinbergb,l, and Kurt Gilesa,b Author Affiliations

aInstitute for Neurodegenerative Diseases, University of California, San Francisco, CA 94143; bDepartment of Neurology, University of California, San Francisco, CA 94143; cDepartment of Biochemistry and Biophysics, University of California, San Francisco, CA 94143; dC. S. Kubik Laboratory for Neuropathology, Department of Pathology, Massachusetts General Hospital, Boston, MA 02114; eDepartment of Pathology, University of California, San Francisco, CA 94143; fCenter for Neurobehavioral Genetics, Center for Autism Research and Treatment, and Department of Neurology, University of California, Los Angeles, CA 90095; gDepartment of Human Genetics, University of California, Los Angeles, CA 90095; hDepartment of Epidemiology and Biostatistics, University of California, San Francisco, CA 94143; iSchool of Medical Science, Faculty of Medicine, University of New South Wales, and Neuroscience Research Australia, Randwick, NSW 2031, Australia; jAgeing Research Unit, School of Public Health, Imperial College London, London SW7 2AZ, United Kingdom; kCentre for Neuroinflammation and Neurodegeneration, Department of Medicine, Imperial College London, London SW7 2AZ, United Kingdom; lMemory and Aging Center, University of California, San Francisco, CA 94143 Contributed by Stanley B. Prusiner, July 22, 2015 (sent for review May 19, 2015)


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Significance Prions are proteins that assume alternate shapes that become self-propagating, and while some prions perform normal physiological functions, others cause disease. Prions were discovered while studying the cause of rare neurodegenerative diseases of animals and humans called scrapie and Creutzfeldt–Jakob disease, respectively. We report here the discovery of α-synuclein prions that cause a more common neurodegenerative disease in humans called multiple system atrophy (MSA). In contrast to MSA, brain extracts from Parkinson’s disease (PD) patients were not transmissible to genetically engineered cells or mice, although much evidence argues that PD is also caused by α-synuclein, suggesting that this strain (or variant) is different from those that cause MSA.



Prions are proteins that adopt alternative conformations that become self-propagating; the PrPSc prion causes the rare human disorder Creutzfeldt–Jakob disease (CJD). We report here that multiple system atrophy (MSA) is caused by a different human prion composed of the α-synuclein protein. MSA is a slowly evolving disorder characterized by progressive loss of autonomic nervous system function and often signs of parkinsonism; the neuropathological hallmark of MSA is glial cytoplasmic inclusions consisting of filaments of α-synuclein. To determine whether human α-synuclein forms prions, we examined 14 human brain homogenates for transmission to cultured human embryonic kidney (HEK) cells expressing full-length, mutant human α-synuclein fused to yellow fluorescent protein (α-syn140*A53T–YFP) and TgM83+/− mice expressing α-synuclein (A53T). The TgM83+/− mice that were hemizygous for the mutant transgene did not develop spontaneous illness; in contrast, the TgM83+/+ mice that were homozygous developed neurological dysfunction. Brain extracts from 14 MSA cases all transmitted neurodegeneration to TgM83+/− mice after incubation periods of ∼120 d, which was accompanied by deposition of α-synuclein within neuronal cell bodies and axons. All of the MSA extracts also induced aggregation of α-syn*A53T–YFP in cultured cells, whereas none of six Parkinson’s disease (PD) extracts or a control sample did so. Our findings argue that MSA is caused by a unique strain of α-synuclein prions, which is different from the putative prions causing PD and from those causing spontaneous neurodegeneration in TgM83+/+ mice. Remarkably, α-synuclein is the first new human prion to be identified, to our knowledge, since the discovery a half century ago that CJD was transmissible.

neurodegeneration Parkinson's disease synucleinopathies strains



Transmission of multiple system atrophy prions to transgenic mice

Joel C. Wattsa,b, Kurt Gilesa,b, Abby Oehlerc, Lefkos Middletond, David T. Dextere, Steve M. Gentlemane, Stephen J. DeArmonda,c, and Stanley B. Prusinera,b,1 Author Affiliations

aInstitute for Neurodegenerative Diseases, and Departments of bNeurology and cPathology, University of California, San Francisco, CA 94143; and dAgeing Research Unit, School of Public Health and eCentre for Neuroinflammation and Neurodegeneration, Department of Medicine, Imperial College, London SW7 2AZ, United Kingdom Contributed by Stanley B. Prusiner, September 30, 2013 (sent for review August 21, 2013)


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Significance Multiple system atrophy (MSA) is a neurodegenerative disorder characterized by the accumulation of misfolded α-synuclein protein in glial cells within the brain. Transgenic mice expressing mutant α-synuclein that were inoculated with brain homogenate from MSA patients developed clinical, biochemical, and pathological signs of a neurodegenerative disease, indicating that MSA is transmissible under certain conditions. This transmissibility is reminiscent of the human prion disorders, such as Creutzfeldt–Jakob disease, and suggests that MSA is caused by the accumulation of toxic α-synuclein prions in the brain.

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Prions are proteins that adopt alternative conformations, which become self-propagating. Increasing evidence argues that prions feature in the synucleinopathies that include Parkinson’s disease, Lewy body dementia, and multiple system atrophy (MSA). Although TgM83+/+ mice homozygous for a mutant A53T α-synuclein transgene begin developing CNS dysfunction spontaneously at ∼10 mo of age, uninoculated TgM83+/− mice (hemizygous for the transgene) remain healthy. To determine whether MSA brains contain α-synuclein prions, we inoculated the TgM83+/− mice with brain homogenates from two pathologically confirmed MSA cases. Inoculated TgM83+/− mice developed progressive signs of neurologic disease with an incubation period of ∼100 d, whereas the same mice inoculated with brain homogenates from spontaneously ill TgM83+/+ mice developed neurologic dysfunction in ∼210 d. Brains of MSA-inoculated mice exhibited prominent astrocytic gliosis and microglial activation as well as widespread deposits of phosphorylated α-synuclein that were proteinase K sensitive, detergent insoluble, and formic acid extractable. Our results provide compelling evidence that α-synuclein aggregates formed in the brains of MSA patients are transmissible and, as such, are prions. The MSA prion represents a unique human pathogen that is lethal upon transmission to Tg mice and as such, is reminiscent of the prion causing kuru, which was transmitted to chimpanzees nearly 5 decades ago.

neurodegeneration bioluminescence imaging seeding proteinopathies



In the studies reported here, we demonstrate that a fatal synucleinopathy can be initiated in Tg(M83+/−:Gfap-luc) mice that do not spontaneously develop a neurologic illness, by intracerebral inoculation with brain homogenate from MSA patients. These results parallel recent reports describing the induction of α-synuclein deposits and dopaminergic neuron loss, but not overt clinical signs of neurologic dysfunction, in non-Tg mice following inoculation with recombinant α-synuclein fibrils (25, 26). Our study reveals that self-propagating, transmissible α-synuclein aggregates (i.e., α-synuclein prions) are formed not just in Tg mice that overexpress mutant α-synuclein, but also in the brains of individuals with a degenerative synucleinopathy such as MSA.

Despite the predilection for oligodendrocytic deposition of α-synuclein in MSA, we did not observe appreciable levels of phosphorylated α-synuclein deposition in oligodendrocytes within the brains of MSA-inoculated bigenic mice. This observation suggests that additional human brain-specific factors may be responsible for encoding the oligodendrocyte-specific tropism of α-synuclein aggregates in MSA. However, a more simple explanation is that the heterologous Prnp promoter that drives mutant α-synuclein expression in TgM83 mice does not engender a native spatial pattern of α-synuclein expression. This difference may preclude deposition in mature oligodendrocytes, which do not express α-synuclein mRNA (31). Inoculation of Tg mice expressing A53T mutant human α-synuclein under the control of the SNCA promoter or even non-Tg mice with MSA brain homogenate may help to resolve this issue.

Although some investigators prefer to use alternate terms to describe the recently recognized “prion” proteins involved in PD, Alzheimer’s disease, and the tauopathies, the shared features of these protein-mediated degenerative diseases are becoming progressively more apparent. Some terms suggested to distinguish self-propagating Aβ, tau, and α-synuclein aggregates from those composed of PrP include “prion-like protein aggregates,” “transmissible proteins,” “templated proteins,” “prionoids,” “proteopathic seeds,” “misfolded proteins,” and “protein pathogens” (32, 33). However, we believe that α-synuclein aggregates fulfill all of the criteria for being labeled a prion. First, brain homogenates prepared from MSA patients or spontaneously ill TgM83+/+ mice containing abundant α-synuclein deposits induce the deposition of insoluble α-synuclein in the brains of recipient Tg(M83+/−:Gfap-luc) mice following intracerebral inoculation [Figs. 3 and 4 (23, 24)], demonstrating that α-synuclein aggregates, like PrPSc, are self-propagating. Second, intracerebral inoculation with samples containing pathological α-synuclein aggregates causes not only seeding of protein aggregation in the brain, but also the induction of clinical signs of neurologic dysfunction, indicative of true disease transmission [Table 1 (23, 24)]. Third, disease can be initiated by recombinant α-synuclein that had been polymerized into fibrils (24), indicating that α-synuclein aggregates, like PrP (34), are sufficient to induce disease. Fourth, transmission of a degenerative synucleinopathy can occur in animals that do not develop spontaneous illness within their normal lifespan [Figs. 1 and 2 (25, 26)], ruling out disease acceleration as a mechanism of transmission. Together, these data mount a compelling case for α-synuclein aggregates in the brains of MSA patients as being prions.

Although α-synuclein aggregates are clearly capable of behaving like prions at the molecular level, there is currently no evidence to suggest that MSA or the other human synucleinopathies are transmissible between humans, in contrast to CJD, which can be transmitted through the use of PrPSc-infected dura mater grafts or growth hormone preparations (35) as well as the reuse of PrPSc-contaminated neurosurgical instruments (36). It is currently unknown whether α-synuclein prions can attach to surgical instruments and to what extent they may persist following sterilization. Although attempts to transmit PD to monkeys by intracerebral inoculation were unsuccessful (37), our transmission data suggest that caution should be exercised when reusing neurosurgical instruments that have been previously used on suspected cases of MSA or PD to minimize any risk for iatrogenic transmission of the disease. Although deep brain stimulation is not commonly used to treat MSA patients, its increasingly wide use to control dyskinesias often found in many patients with advanced PD requires surgical implantation (38) and, as such, may represent a potential risk for human-to-human transmission of α-synuclein prions.

The rapid transmission of the MSA inocula, compared with TgM83+/+ samples, in Tg(M83+/−:Gfap-luc) mice was surprising for two reasons. First, the MSA samples do not harbor α-synuclein with the A53T mutation, which is present in the TgM83 line. For PrPSc prions, even a single amino acid mismatch between PrPSc in the inoculum and PrPC in the host can dramatically prolong the disease incubation period (39). Thus, there does not appear to be a substantial “transmission barrier” between the WT α-synuclein aggregates present in the MSA inocula and the A53T mutant α-synuclein present in the mice. Second, the levels of insoluble phosphorylated α-synuclein were much lower in the MSA brains than in the brains of the spontaneously ill TgM83+/+ mice used as inocula. This observation could suggest that the most infectious α-synuclein species may consist of smaller, more soluble assemblies, as has been observed for PrPSc and Aβ prions (40, 41). However, a more likely explanation for the rapid transmission of MSA prions is that these α-synuclein aggregates constitute a distinct “strain” of prion from the aggregates found in spontaneously ill TgM83+/+ mice. In prion disease, distinct strains are believed to result from conformational differences in PrPSc (42, 43). Indeed, conformationally distinct “strains” of recombinant α-synuclein aggregates that possess varying ability to initiate tau aggregation have recently been identified (44). Thus, the α-synuclein aggregates found within oligodendrocytes in the brains of MSA patients may be conformationally distinct from those found in the brains of TgM83+/+ mice, engendering distinct transmission properties. The rapid transmissibility of the MSA strain of α-synuclein prions may reflect the fact that the α-synuclein aggregates are not sequestered in Lewy bodies, which may constitute a protective mechanism to limit the spread of a distinct group of strains of α-synuclein prions in PD and DLB.

The successful transmission of MSA prions to Tg(M83+/−:Gfap-luc) mice described herein represents a unique human neurodegenerative disease that demonstrates lethality upon transmission to animals and is reminiscent of the transmission of kuru, CJD, and related diseases to nonhuman primates (45, 46). Although Aβ and tau prions derived from the brains of Alzheimer’s disease or tauopathy patients, respectively, stimulate prion formation as detected by protein aggregation and deposition upon inoculation into susceptible Tg mice, neither induces overt signs of neurologic disease nor lethality in the recipient animals (12, 47). Importantly, MSA-inoculated bigenic mice may comprise a reliable system for assessing the therapeutic efficacy of drugs designed to target the formation of α-synuclein prions.


Multiple System Atrophy (MSA)

What is Multiple System Atrophy (MSA)?

Multiple system atrophy (MSA) is a rare neurodegenerative disease marked by a combination of symptoms affecting movement, blood pressure, and other body functions; hence the label "multiple system" atrophy. According to the American Autonomic Society, Multiple System Atrophy (MSA) is a sporadic, progressive, adult-onset disorder characterized by autonomic dysfunction, parkinsonism and ataxia (a failure of muscular coordination) in any combination.

Symptoms of MSA include:

Orthostatic hypotension,or a significant fall in blood pressure when standing, causing dizziness, lightheadedness, fainting, or blurred vision urinary difficulties or constipation motor control symptoms, including tremor, rigidity, and loss of muscle coordination, loss of balance male impotence (inability to achieve or maintain an erection) speech or swallowing difficulties Who gets MSA?

MSA affects both men and women primarily in their 50s.

What causes MSA?

Multiple system atrophy is associated with deterioration and shrinkage (atrophy) of portions of the brain (cerebellum, basal ganglia and brainstem) that regulate internal body functions, digestion and motor control.

There is no known cause for brain changes in MSA.

How is MSA diagnosed?

Diagnosis of MSA can be challenging because there is no test that can make or confirm the diagnosis in a living patient. Certain signs and symptoms of MSA also occur with other disorders, such as Parkinson's disease, making the diagnosis more difficult.

If your doctor suspects multiple system atrophy, he or she will obtain a medical history and perform a physical examination. You may receive a referral to a neurologist or other specialist for specific evaluations that can help in making the diagnosis.

Tests that may be helpful in making a diagnosis include:

Tilt table test - In this procedure, your blood pressure is monitored while you are on a special table that will tilt you to an almost upright position. This allows the physician to record blood pressure irregularities, and information about whether they occur with a change in physical position. Blood tests A sweat test to evaluate perspiration Tests to assess your bladder and bowel function Electrocardiogram to track the electrical signals of your heart Brain-imaging tests, particularly a magnetic resonance imaging (MRI) scan, to determine if another condition might be triggering symptoms Pharmacological challenge tests (administering certain medications and observing the patient’s body’s reaction to them, in controlled clinical settings) For patients with sleep irregularities, particularly if they involve interrupted breathing or snoring, physicians may recommend an evaluation in a sleep laboratory to determine if there is an underlying and treatable sleep disorder, such as sleep apnea.

What is the treatment for MSA?

There is no known cure for MSA, so management of the disease focuses on treating the more disabling symptoms listed above.

A clinical trial of the drug Rifampicin is being conducted by the Autonomic Disorders Consortium.


Tuesday, November 26, 2013

Transmission of multiple system atrophy prions to transgenic mice


Self-Propagative Replication of Ab Oligomers Suggests Potential Transmissibility in Alzheimer Disease

Received July 24, 2014; Accepted September 16, 2014; Published November 3, 2014


Singeltary comment ;


Saturday, March 21, 2015

*** Canada and United States Creutzfeldt Jakob TSE Prion Disease Incidence Rates Increasing



Sunday, November 23, 2014

*** Confirmed Variant Creutzfeldt-Jakob Disease (variant CJD) Case in Texas in June 2014 confirmed as USA case NOT European ***

the patient had resided in Kuwait, Russia and Lebanon. The completed investigation did not support the patient's having had extended travel to European countries, including the United Kingdom, or travel to Saudi Arabia. The specific overseas country where this patient’s infection occurred is less clear largely because the investigation did not definitely link him to a country where other known vCJD cases likely had been infected.


Sunday, December 14, 2014

*** ALERT new variant Creutzfeldt Jakob Disease nvCJD or vCJD, sporadic CJD strains, TSE prion aka Mad Cow Disease United States of America Update December 14, 2014 Report ***


Thursday, July 30, 2015

Professor Lacey believes sporadic CJD itself originates from a cattle infection number of cattle farmers falling victim to Creutzfeld-Jakob Disease is much too high to be mere chance


Tuesday, August 4, 2015

FDA U.S. Measures to Protect Against BSE








Zoonotic Potential of CWD Prions


Liuting Qing1, Ignazio Cali1,2, Jue Yuan1, Shenghai Huang3, Diane Kofskey1, Pierluigi Gambetti1, Wenquan Zou1, Qingzhong Kong1 1Case Western Reserve University, Cleveland, Ohio, USA, 2Second University of Naples, Naples, Italy, 3Encore Health Resources, Houston, Texas, USA




***These results indicate that the CWD prion has the potential to infect human CNS and peripheral lymphoid tissues and that there might be asymptomatic human carriers of CWD infection.***




P.105: RT-QuIC models trans-species prion transmission


Kristen Davenport, Davin Henderson, Candace Mathiason, and Edward Hoover Prion Research Center; Colorado State University; Fort Collins, CO USA




***This insinuates that, at the level of protein:protein interactions, the barrier preventing transmission of CWD to humans is less robust than previously estimated.***





From: Terry S. Singeltary Sr.


Sent: Saturday, November 15, 2014 9:29 PM


To: Terry S. Singeltary Sr.










*** The association between venison eating and risk of CJD shows similar pattern, with regular venison eating associated with a 9 FOLD INCREASE IN RISK OF CJD (p = 0.04). (SEE LINK IN REPORT HERE...TSS) PLUS, THE CDC DID NOT PUT THIS WARNING OUT FOR THE WELL BEING OF THE DEER AND ELK ;





*** These results would seem to suggest that CWD does indeed have zoonotic potential, at least as judged by the compatibility of CWD prions and their human PrPC target. Furthermore, extrapolation from this simple in vitro assay suggests that if zoonotic CWD occurred, it would most likely effect those of the PRNP codon 129-MM genotype and that the PrPres type would be similar to that found in the most common subtype of sCJD (MM1).***



*** The potential impact of prion diseases on human health was greatly magnified by the recognition that interspecies transfer of BSE to humans by beef ingestion resulted in vCJD. While changes in animal feed constituents and slaughter practices appear to have curtailed vCJD, there is concern that CWD of free-ranging deer and elk in the U.S. might also cross the species barrier. Thus, consuming venison could be a source of human prion disease. Whether BSE and CWD represent interspecies scrapie transfer or are newly arisen prion diseases is unknown. Therefore, the possibility of transmission of prion disease through other food animals cannot be ruled out. There is evidence that vCJD can be transmitted through blood transfusion. There is likely a pool of unknown size of asymptomatic individuals infected with vCJD, and there may be asymptomatic individuals infected with the CWD equivalent. These circumstances represent a potential threat to blood, blood products, and plasma supplies.



Tuesday, May 26, 2015


*** Minimise transmission risk of CJD and vCJD in healthcare settings ***


Last updated 15 May 2015



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 ;



 *** 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



Saturday, December 13, 2014


*** Terry S. Singeltary Sr. Publications TSE prion disease Peer Review ***


Diagnosis and Reporting of Creutzfeldt-Jakob Disease


Singeltary, Sr et al. JAMA.2001; 285: 733-734. Vol. 285 No. 6, February 14, 2001 JAMA





Terry S. Singeltary Sr.
























iatrogenic, what if ???

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