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Development of oligomeric prion‐protein aggregates in a mouse model of prion disease
Author(s) -
Sasaki Kensuke,
Minaki Haruhiko,
Iwaki Toru
Publication year - 2009
Publication title -
the journal of pathology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.964
H-Index - 184
eISSN - 1096-9896
pISSN - 0022-3417
DOI - 10.1002/path.2576
Subject(s) - proteinase k , gene isoform , western blot , protease , chemistry , blot , prion protein , microbiology and biotechnology , in vitro , biochemistry , size exclusion chromatography , biology , enzyme , disease , pathology , medicine , gene
In prion diseases the normal cellular isoform of prion protein (PrP), denoted PrP C , is converted into an abnormal, pathogenic isoform of PrP (PrP Sc ). Diagnostic tools for prion diseases are conventionally based on the detection of protease‐resistant PrP (PrP res ) after proteinase K digestion. However, recent studies have revealed that protease‐sensitive abnormal PrP (sPrP Sc ) also exists in significant amounts in brains suffering from prion diseases. Here, we designed a simplified size‐exclusion gel chromatography assay, using disposable spin columns to examine PrP aggregates in the course of the disease, without proteinase K digestion. Brain homogenates of NZW mice, inoculated intracranially with Fukuoka‐1 strain, and which died at around 120 days post‐inoculation, were assayed by this gel‐fractionation method and eluted PrP molecules in each fraction were detected by western blot analysis. Oligomeric PrP molecules were well separated from monomers, as predicted. A conventional protease‐digestion assay was also performed to detect PrP res and revealed that the ratio of PrP res to total PrP increased drastically from 105 days. However, the increase of PrP oligomers became significant from 90 days. These PrP oligomers in the early disease stage would, therefore, be sPrP Sc molecules that might affect the disease pathology, such as spongiform change and abnormal PrP deposition. We also observed that the resistance of PrP oligomers to proteinase K and insolubility in phosphotungstic acid precipitation increased with disease progression, which suggests that PrP oligomers are not clearly distinguished from cellular PrP or PrP res but may overlap in a continuous spectrum. Our study casts light on the ambiguity of the definition of PrP Sc and indicates that the abnormality of PrP molecules should be determined from various perspectives, more than protease resistance. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.