SEARCH VEGSOURCE:

 

 

Follow Ups | Post Followup | Back to Discussion Board | VegSource
See spam or
inappropriate posts?
Please let us know.
  




From: TSS ()
Subject: Cells infected with scrapie and Creutzfeldt–Jakob disease agents produce intracellular 25-nm virus-like particles
Date: January 31, 2007 at 10:28 am PST

Cells infected with scrapie and Creutzfeldt–Jakob

disease agents produce intracellular 25-nm

virus-like particles

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 N2a22L

cells, yet this did not increase either the number of virus-like arrays

or the infectious titer of these cells. Thus, the 25-nm infectionassociated

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.

snip.

In summation, all of this data provides a clear, consistent,

substantive, and logical alternative to the accepted prion hypothesis.

The causative TSE agent is most consistent with an

exogenous 25-nm virion without intrinsic host PrP. The stimulation

of host innate immune responses by these agents, a

complex set of molecular reactions that precedes the elaboration

of pathologic PrP (9) and one that is not provoked by PrP-res

itself (25), also point to a foreign pathogen rather than some

unpredictably spontaneous mutation in the host’s PrP without

cause. The presence of these particles in many different species

infected with a wide variety of TSE strains is in accord with

Koch’s first requirement (1). It is also improbable that an

identical virus-like structure would be a contaminant or a

secondary coincidental feature of all these different TSE models.

Nevertheless, a more detailed molecular analysis of these

particles will be required to substantiate their causal nature.

Purification of these 25-nm particles from productive tissue

cultures should be informative if the essential infectivity assays

are performed systematically with parallel ultrastructural and

molecular analyses. Animal titrations of infectivity are expensive

and prolonged. However, sustained and reproducible infection

of indicator GT cells by a variety of TSE agents already has

shown that they can rapidly authenticate the presence of agent

in disrupted samples as well as in living cells (4, 17). GT cells also

may be used for testing infectivity of viral nucleic acids as well

as PrP conformers. Rapid assays of infectivity in culture should

facilitate the isolation of infectious particles from host components,

and treatments that modify the production of these

particles in culture may resolve further the infectious structure

from the pathological disease processes it initiates.

Materials and Methods......snip......end........TSS

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


tss



Follow Ups:



Post a Followup

Name:
E-mail: (optional)
Subject:

Comments:

Optional Link URL:
Link Title:
Optional Image URL: