Tuesday, January 08, 2008

Tubulovesicular structures are a consistent (and unexplained) finding in the brains of humans with prion diseases

Tuesday, January 08, 2008 Tubulovesicular structures are a consistent (and unexplained) finding in the brains of humans with prion diseases Virus Res. 2007 Dec 27; : 18164506

Tubulovesicular structures are a consistent (and unexplained) finding in the brains of humans with prion diseases.

[My paper] Pawel P Liberski , Beata Sikorska , Jean-Jacques Hauw , Nicolas Kopp , Nathalie Streihenberger , Pierric Giraud , Herbert Budka , J W Boellaard , Paul Brown

Creutzfeldt-Jakob disease (CJD), Gerstmann-Sträussler-Scheinker disease (GSS) and Fatal Familial Insomnia (FFI) are slow neurodegenerative disorders classified as transmissible spongiform encephalopathies (TSEs) or prion diseases, which appear in sporadic, hereditary or environmentally acquired forms. Tubulovesicular structures (TVS) are ultrastructural particles of unknown origin and chemical composition found in the brains of both animal and human forms of transmissible spongiform encephalopathies or prion diseases. In this paper, we report the results of a search for TVS in a total of 13 cases of sporadic Creutzfeldt-Jakob disease, three cases of Gerstmann-Sträussler-Scheinker disease, two cases of Fatal Familial Insomnia, and individual cases of familial, iatrogenic, and variant CJD (vCJD). TVS were found in all but one sporadic and one familial case of CJD. As controls, we examined 15 cases of Alzheimer's disease (AD), two cases of Pick's disease, and one case of multiple system atrophy. TVS were not present in any of these cases. This study confirms the TSE-specificity of TVS, the morphology of which suggests a possible pathogenetic role and relationship to recently described virion-like arrays of 25nm particles in scrapie-infected tissue cultures. http://lib.bioinfo.pl/auth:Brown,P

Neuroscience Cells infected with scrapie and Creutzfeldt-Jakob disease agents produce intracellular 25-nm virus-like particles ( infection neuroectodermal cultures virion ultrastructure prion amyloid retrovirus )

Laura Manuelidis *, Zhoa-Xue Yu, Nuria Banquero, and Brian Mullins Yale Medical School, 333 Cedar Street, New Haven, CT 06510

Communicated by Sheldon Penman, Massachusetts Institute of Technology, Cambridge, MA, December 11, 2006 (received for review October 10, 2006)

We had repeatedly found 25-nm-diameter virus-like particles in highly infectious brain fractions with little prion protein (PrP), and therefore we searched for similar virus-like particles in situ in infected cell lines with high titers. Neuroblastoma cells infected with the 22L strain of scrapie as well as hypothalamic GT cells infected with the FU Creutzfeldt-Jakob disease agent, but not parallel mock controls, displayed dense 25-nm virus-like particles in orthogonal arrays. These particles had no relation to abnormal PrP amyloid in situ, nor were they labeled by PrP antibodies that faithfully recognized rough endoplasmic reticulum membranes and amyloid fibrils, the predicted sites of normal and pathological intracellular PrP. Additionally, phorbol ester stimulated the production of abnormal PrP gel bands by >5-fold in infected N2a + 22L cells, yet this did not increase either the number of virus-like arrays or the infectious titer of these cells. Thus, the 25-nm infection-associated particles could not be prions. Synaptic differentiation and neurodegeneration, as well as retroviruses that populate the rough endoplasmic reticulum of neuroblastoma cells, were not required for particle production. The 25-nm particle arrays in cultured cells strongly resembled those first described in 1968 in synaptic regions of scrapie-infected brain and subsequently identified in many natural and experimental TSEs. The high infectivity of comparable, isolated virus-like particles that show no intrinsic PrP by antibody labeling, combined with their loss of infectivity when nucleic acid-protein complexes are disrupted, make it likely that these 25-nm particles are the causal TSE virions that induce late-stage PrP brain pathology.

---------------------------------------------------------------------------- ----

Author contributions: L.M. designed research; L.M., Z.-X.Y., N.B., and B.M. performed research; L.M., Z.-X.Y., and N.B. analyzed data; and L.M. wrote the paper.

The authors declare no conflict of interest.

*To whom correspondence should be addressed.

Laura Manuelidis, E-mail: laura.manuelidis@yale.edu

www.pnas.org/cgi/doi/10.1073/pnas.0610999104


http://www.pnas.org/cgi/content/abstract/0610999104v1


please see ;

http://info.med.yale.edu/neurosci/faculty/manuelidis_main.html


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From: TSS Subject: Disease-specific particles without prion protein in prion diseases – phenomenon or epiphenomenon? Date: July 13, 2007 at 9:49 am PST

Disease-specific particles without prion protein in prion diseases – phenomenon or epiphenomenon?

P. P. Liberski**Department of Molecular Pathology and Neuropathology, Medical University of Lodz, Lodz, Poland; and Pawel P. Liberski, Department of Molecular Pathology and Neuropathology, Medical University of Lodz, Czechoslowacka Street 8/10, PL 92-216 Lodz, Poland. Tel: +42-679-14-77; Fax: +42-679-14-77; E-mail: ppliber@csk.am.lodz.pl and P. Brown††Refired, Bethesda, Maryland, USA *Department of Molecular Pathology and Neuropathology, Medical University of Lodz, Lodz, Poland; and †Refired, Bethesda, Maryland, USA Pawel P. Liberski, Department of Molecular Pathology and Neuropathology, Medical University of Lodz, Czechoslowacka Street 8/10, PL 92-216 Lodz, Poland. Tel: +42-679-14-77; Fax: +42-679-14-77; E-mail: ppliber@csk.am.lodz.pl P. P. Liberski and P. Brown (2007) Neuropathology and Applied Neurobiology 33, 395–397

Disease-specific particles without prion protein in prion diseases – phenomenon or epiphenomenon?

Abstract

The search for the cause of transmissible spongiform encephalopathies (TSEs) has a long and tortuous history. In a recent paper, 25-nm virus-like particles were identified that were consistently observed in cell cultures infected with Creutzfeldt-Jakob disease (CJD) and scrapie; they are similar to, or even identical with, the virus-like tubulovesicular structures (TVS)

found in experimental scrapie as early as in 1968, and subsequently in all naturally occurring and experimentally induced TSEs. These particles have been viewed with caution by the scientific community because of the unverified or uninterpretable record of virus-like structures reported over the years in TSEs. TVS are spherical or tubular particles of approximate diameter 25–37 nm. They are smaller than synaptic vesicles, but larger than many particulate structures of the central nervous system, such as glycogen granules. Their electron density is higher compared with synaptic vesicles, and in experimental murine scrapie, they form paracrystalline arrays. None of these observations distinguish between TVS as an entity critical to the infectious process, or as a highly specific ultrastructural epiphenomenon, but their consistent presence in all TSEs demands further research.

http://www.blackwell-synergy.com/doi/abs/10.1111/j.1365-2990.2007.00867.x


Confusious is confused again ???

An epiphenomenon is a secondary phenomenon that occurs alongside a primary phenomenon.

Often, a causal relationship between the phenomena is implied: the epiphenomenon is a consequence of the primary phenomenon. In medicine, this relationship is typically not implied: an epiphenomenon may occur independently, and is merely called an epiphenomenon because it is not the primary phenomenon under study. (A side-effect is a specific kind of epiphenomenon that does occur as a consequence.)

In philosophy of mind, epiphenomenalism is the view that mental phenomena are caused by physical phenomena, and cannot cause anything themselves. It was probably first mentioned by T. H. Huxley in 1874.

Marvelous that the authors ignore the fact that Manuelidis et al first reported dense 25nm diameter particles (by field flow fractionation in 1992) in highly infectious brain fractions with little PrP. Unlike PrP, these viruslike particles have not been found in uninfected brains. Of course they are important to characterize. Though the authors seem to dismiss them as epiphenomena. Could the dense 25nm particles be the Phenomenon, with the PrP the Epiphenomenon ? which came first, the horse or the cart ???

TSS et al ;-)

BIOLOGICAL SCIENCES / NEUROSCIENCE Cells infected with scrapie and Creutzfeldt–Jakob disease agents produce intracellular 25-nm virus-like particles

Laura Manuelidis*, Zhoa-Xue Yu, Nuria Barquero, and Brian Mullins

Yale Medical School, 333 Cedar Street, New Haven, CT 06510

Communicated by Sheldon Penman, Massachusetts Institute of Technology, Cambridge, MA, December 11, 2006 (received for review October 10, 2006)

We had repeatedly found 25-nm-diameter virus-like particles in highly infectious brain fractions with little prion protein (PrP), and therefore we searched for similar virus-like particles in situ in infected cell lines with high titers. Neuroblastoma cells infected with the 22L strain of scrapie as well as hypothalamic GT cells infected with the FU Creutzfeldt–Jakob disease agent, but not parallel mock controls, displayed dense 25-nm virus-like particles in orthogonal arrays. These particles had no relation to abnormal PrP amyloid in situ, nor were they labeled by PrP antibodies that faithfully recognized rough endoplasmic reticulum membranes and amyloid fibrils, the predicted sites of normal and pathological intracellular PrP. Additionally, phorbol ester stimulated the production of abnormal PrP gel bands by >5-fold in infected N2a + 22L cells, yet this did not increase either the number of virus-like arrays or the infectious titer of these cells. Thus, the 25-nm infection-associated particles could not be prions. Synaptic differentiation and neurodegeneration, as well as retroviruses that populate the rough endoplasmic reticulum of neuroblastoma cells, were not required for particle production. The 25-nm particle arrays in cultured cells strongly resembled those first described in 1968 in synaptic regions of scrapie-infected brain and subsequently identified in many natural and experimental TSEs. The high infectivity of comparable, isolated virus-like particles that show no intrinsic PrP by antibody labeling, combined with their loss of infectivity when nucleic acid–protein complexes are disrupted, make it likely that these 25-nm particles are the causal TSE virions that induce late-stage PrP brain pathology.

infection neuroectodermal cultures virion ultrastructure prion amyloid retrovirus

---------------------------------------------------------------------------- ---- Author contributions: L.M. designed research; L.M., Z.-X.Y., N.B., and B.M. performed research; L.M., Z.-X.Y., and N.B. analyzed data; and L.M. wrote the paper. The authors declare no conflict of interest.

*To whom correspondence should be addressed. E-mail: laura.manuelidis@yale.edu

© 2007 by The National Academy of Sciences of the USA

http://www.pnas.org/cgi/content/abstract/104/6/1965?hits=10&RESULTFORMAT=&FIRSTINDEX=10&maxtoshow=&HITS=10&fulltext=manuelidis&searchid=1&resourcetype=HWCIT


Prospect A 25 nm virion is the likely cause of transmissible spongiform encephalopathies

Laura Manuelidis * Yale Medical School, New Haven, Connecticut 06510

email: Laura Manuelidis (laura.manuelidis@yale.edu)

*Correspondence to Laura Manuelidis, Yale Medical School, New Haven, CT 06510.

Funded by: NIH-NS12674 DOD-DAMD-17-03-1-0360

Keywords Creutzfeldt-Jakob disease • scrapie • BSE • viral particle • latent infection • prion pathology

Abstract The transmissible spongiform encephalopathies (TSEs) such as endemic sheep scrapie, sporadic human Creutzfeldt-Jakob disease (CJD), and epidemic bovine spongiform encephalopathy (BSE) may all be caused by a unique class of slow viruses. This concept remains the most parsimonious explanation of the evidence to date, and correctly predicted the spread of the BSE agent to vastly divergent species. With the popularization of the prion (infectious protein) hypothesis, substantial data pointing to a TSE virus have been largely ignored. Yet no form of prion protein (PrP) fulfills Koch's postulates for infection. Pathologic PrP is not proportional to, or necessary for infection, and recombinant and amplified prions have failed to produce significant infectivity. Moreover, the wealth of data claimed to support the existence of infectious PrP are increasingly contradicted by experimental observations, and cumbersome speculative notions, such as spontaneous PrP mutations and invisible strain-specific forms of infectious PrP are proposed to explain the incompatible data. The ability of many slow viruses to survive harsh environmental conditions and enzymatic assaults, their stealth invasion through protective host-immune defenses, and their ability to hide in the host and persist for many years, all fit nicely with the characteristics of TSE agents. Highly infectious preparations with negligible PrP contain nucleic acids of 1-5 kb, even after exhaustive nuclease digestion. Sedimentation as well as electron microscopic data also reveal spherical infectious particles of 25-35 nm in diameter. This particle size can accommodate a viral genome of 1-4 kb, sufficient to encode a protective nucleocapsid and/or an enzyme required for its replication. Host PrP acts as a cellular facilitator for infectious particles, and ultimately accrues pathological amyloid features. A most significant advance has been the development of tissue culture models that support the replication of many different strains of agent and can produce high levels of infectivity. These models provide new ways to rapidly identify intrinsic viral and strain-specific molecules so important for diagnosis, prevention, and fundamental understanding. J. Cell. Biochem. 100: 897-915, 2007. © 2006 Wiley-Liss, Inc.

---------------------------------------------------------------------------- ---- Received: 12 July 2006; Accepted: 13 July 2006 Digital Object Identifier (DOI)

http://www3.interscience.wiley.com/cgi-bin/abstract/112784723/ABSTRACT?CRETRY=1&SRETRY=0


doi:10.1016/0168-1702(92)90016-3 Copyright © 1992 Published by Elsevier Science B.V.

Analysis of Creutzfeldt-Jakob disease infectious fractions by gel permeation chromatography and sedimentation field flow fractionation

T. Sklaviadis1, R. Dreyer2 and Laura Manuelidis Yale University, School of Medicine, New Hauen, CT 06510, USA Received 17 June 1992; revised 3 September 1992; accepted 8 September 1992. Available online 5 November 2002.

Abstract

Gel permeation chromatography and sedimentation field flow fractionation (SF3) were used to further analyze highly infectious fractions from Creutzfeldt-Jakob disease (CJD) infected hamster brain. These analyses defined the relative molecular mass and physical size of the Creutzfeldt-Jakob disease (CJD) agent with greater precision than previously possible. Highly purified disaggregated fractions yielded single, homogeneous Gaussian peaks with both methods. The relevant analytical peaks contained protein-nucleic acid complexes with an Mrr of ˜ 1.5 × 107 daltons and a mean radius of ˜ 30 nm. The experimental evidence further solidifies the concept of an infectious agent that resembles a viral core rather than a simple protein.

Author Keywords: Scrapie; Creutzfeldt-Jakob Disease; Gel permeation chromatography; Sedimentation field flow fractionation; Agent radius; Molecular mass

1 Present address: Aristotle University of Thessaloniki, School of Health Sciences, Department of Pharmaceutical Sciences, Thessaloniki 54006, Greece.

2 Present address: Miles Laboratories MRC, 400 Morgan Lane, West Haven, CT 06516, USA.

Correspondence to: L. Manuelidis, Yale University, School of Medicine, 310 Cedar St., , New Haven, CT 06510, , USA.

http://www.sciencedirect.com/


1. Sklaviadis, T., Manuelidis, L. and Manuelidis E.E. (1986). Characterization of major peptides in Creutzfeldt-Jakob disease and scrapie. Proc. Natl. Acad. Sci. USA 83:6146-6150.

2. Manuelidis, L., Sklaviadis, T. and Manuelidis, E.E. (1987). Evidence suggesting that PrP is not the infectious agent in Creutzfeldt-Jakob disease. EMBO J 6:341-347.

3. Manuelidis, L., Tesin, D., Sklaviadis, T. and Manuelidis, E.E. (1987). Astrocyte gene expression in Creutzfeldt-Jakob disease. Proc. Natl. Acad. Sci. USA 84:5937-5941.

4. Manuelidis, L., Sklaviadis, T. and Manuelidis, E.E. (1987). On the origin and significance of scrapie associated fibrils. In: Court, L., Dormont, D., Brown, P., Kingbury, D.T. (eds) 2nd International Symposium of Unconventional Virus Diseases of the Central Nervous System. Masson, Paris, pp 489-507.

5. Sklaviadis, T.K., Manuelidis, L. and Manuelidis, E.E. (1989). Physical properties of the Creutzfeldt-Jakob disease agent. J. Virol. 63:1212-1222.

6. Murdoch, G.H., Sklaviadis, T., Manuelidis, E.E., Manuelidis, L. (1990). Potential retroviral RNAs in Creutzeldt-Jakob disease. J. Virol. 64:1477-1486.

7. Sklaviadis, T., Akowitz, A., Manuelidis, E.E., Manuelidis, L. (1990). Nuclease treatment results in high specific purification of Creutzfeldt-Jakob disease infectivity with a density characteristic of nucleic acid-protein complexes. Arch. Virol. 112: 215-229.

8. Akowitz, A., Sklaviadis, T., Manuelidis, E.E., Manuelidis, L. (1990). Nuclease-resistant polyadenylated RNAs of significant size are detected by PCR in highly purified Creutzfeldt-Jakob disease preparations. Microbial Pathogenesis 9:33-45.

9. Sklaviadis, T., Dreyer, R., Manuelidis, L.(1992) Analysis of Creutzfedt-Jakob disease infectious fractions by gel permeation chromatography and sedimentation field flow fractionation. Virus Research 26, 241-254.

10. Sklaviadis, T., Akowitz, A., Manuelidis, E.E., Manuelidis, L. (1993) Nucleic acid binding proteins in highly purified Creutzfeldt-Jakob disease preparations. Proc. Natl. Acad. Sci. USA, 90: 5713-5717.

11. Vizirianakis, J.S., Tsiftsoglou A., S., Sklaviadis, T. Developments in slow virus research: Implications of PrP protein involvement in cell growth and differentiation (1993), Bovine Spongiform Encephalopathy. Where are we now? Commision of the European Communities, 14-15 September 1993, Brussels

12. Akowitz, A., Sklaviadis, T., Manuelidis, L. (1994) Endogenous viral complexes with long RNA cosediment with the agent of Creutzfeldt-Jakob Disease. Nucl. Acids. Res., 22: 1101-1107.

13. Manuelidis, T. Sklaviadis, A. Akowitz, W. Fritch (1995) Viral particles are required for infection in neurodegenerative Creutzfeldt-Jakob disease Proc. Natl. Acad. Sci. USA , 92: 5124-5128.

14. Papaconstantinou, E., Karakiulakis, G., Roth, M., Dawson,M., Keyes, P., Papadopoulos,O.,Sklaviadis,T. (1999) Glycan alterations associated with bovine spongiform encephalopathy (Accepted in archives of biochemistry and biophysics)

15. S.Verghese-Nikolakaki, H. Mihaloudi, M. Polymenidou, M. Groschup, G. Papadopoulos, T. Sklaviadis (1999) Expression of the prion protein in the rat forebrain-, an immunohistochemical study. Neuroscience letters 272 9-12

16. S. Leontidis, V. Psychas, S. Argyroudis, A. Giannti-Stefanou, E. Paschaleri-Papadopoulou, T. Manousis, T. Sklaviadis (1999) An overeleven year survey of neurologic diseases of ruminats with special reference to transmissible spongiform encephalopathies (TSEs) in Greece (Accepted for publication Journal of Veterinary Medicine)

17. T. Manousis, M. Sachsamanoglou, P. Toumazos, S. Vergehese-Nikolakaki, O. Papadopoulos, T. Sklaviadis (1999) Western blot detection of PrP Sc in Cyprus sheep with natural scrapie (Accepted for publication The Veterinary Journal)

18. Vassilikioti ,O. Papadopoulos,T. Sklaviadis (1999) Reverse transriptase activity in BSE infectious fractions. (Submitted)

19. Sachsamanoglou, M., Lolis, E., Sklaviadis, T. (1999) Thermostable and chemically induced intermediate conformations of human PrP protein (submitted)

20. T. Manousis, P. Keyes, I. Dexter, R. Green, M. Sachsananoglou, M. Dawson, S. verghese-Nikolakaki, O. Papadopoulos, T. Sklaviadis (1999) Characterization of the murine BSE infectious agent (Submitted Journal of General Virology)

http://www.pharm.auth.gr/prion/Publications.htm


TSS

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