Complex proteinopathy with accumulations of prion protein, hyperphosphorylated tau, α-synuclein and ubiquitin in experimental bovine spongiform encephalopathy of monkeys

Abstract

Proteins aggregate in several slowly progressive neurodegenerative diseases called 'proteinopathies'. Studies with cell cultures and transgenic mice overexpressing mutated proteins suggested that aggregates of one protein induced misfolding and aggregation of other proteins as well - a possible common mechanism for some neurodegenerative diseases. However, most proteinopathies are 'sporadic', without gene mutation or overexpression. Thus, proteinopathies in WT animals genetically close to humans might be informative. Squirrel monkeys infected with the classical bovine spongiform encephalopathy agent developed an encephalopathy resembling variant Creutzfeldt-Jakob disease with accumulations not only of abnormal prion protein (PrP(TSE)), but also three other proteins: hyperphosphorylated tau (p-tau), α-synuclein and ubiquitin; β-amyloid protein (Aβ) did not accumulate. Severity of brain lesions correlated with spongiform degeneration. No amyloid was detected. These results suggested that PrP(TSE) enhanced formation of p-tau and aggregation of α-synuclein and ubiquitin, but not Aβ, providing a new experimental model for neurodegenerative diseases associated with complex proteinopathies.

Fig. 1.

SQs inoculated with classical BSE (SQ-BSE) develop severe TSE. (a) Spongiform degeneration, (b) astrogliosis, (c) PrP^TSE deposition and (d) ubiquitin. Immunopositivity in the frontal cortex of a SQ-BSE. (a) Haematoxylin and eosin; (b–d) immunostaining for GFAP, PrP (6H4) and ubiquitin (Ubi-1), respectively. Bars, 100 µm.

Fig. 2.

Accumulation of p-tau protein in the cerebrum of SQ-BSE. Granular and rod-shaped tau in the frontal cortex of SQ-BSE. (a–d, f) Immunopositivity observed with anti-tau antibodies AT8, AT100, AT180 and AT270, and anti-4-repeat tau, RD4, respectively. (e) No immunopositivity was observed in tissue sections probed with anti-3-repeat tau, RD3. Bars, 100 µm.

Fig. 3.

α-Synucleinopathy in the cerebrum of SQ-BSE. (a, c, d) Granular α-synuclein immunopositivity in the frontal cortex of SQ-BSE with severe spongiform degeneration. (b) No α-synuclein accumulation in tissue sections of the temporal cortex (area without spongiform degeneration or PrP^TSE deposition) of the same animal shown in (a, c, d). (e) Fine granular and evenly distributed synaptophysin immunoreactivity in the temporal cortex (area without spongiform degeneration or PrP^TSE deposition) of SQ-BSE. (f) Reduced and disorganized synaptophysin immunoreactivity in frontal cortex of SQ-BSE with severe TSE. Bars, 100 µm (a, b, e, f); 50 µm (c, d).

Fig. 4.

p-Tau and α-synuclein co-localized in brain of SQ-BSE. (a) Small punctate α-synuclein (black arrows), (b) large granular and rod-shaped p-tau (red arrow heads), and (c) co-localization of α-synuclein (black arrows) and p-tau (red arrow heads) in the frontal cortex of SQ-BSE and severe spongiform degeneration. Tau was detected with antibody AT8 and α-synuclein was detected with antibody 4D6. Double staining was performed using Vectastain Elite ABC Kit (Vector Laboratories). (d) No immunopositivity was observed in tissue sections probed with normal mouse serum. Bars, 50 µm.