Efficient transmission of classical scrapie agent x124 by intralingual route to genetically susceptible sheep with a low dose inoculum.

Scrapie is a naturally occurring prion disease of sheep and goats that results in accumulation of the misfolded prion protein (PrPSc) and progressive neurodegeneration. After inoculation with classical scrapie isolate x124, susceptibility and incubation period are associated with valine at codon 136 (V136) of the prion protein: VRQ/VRQ had the shortest incubation periods, followed by VRQ/ARQ sheep, while ARQ/ARQ sheep only developed disease after inoculation via the intracerebral route. Intralingual inoculation of TSE agents effectively transmits disease similar to intracranial inoculation; therefore, it is possible that oral lesions may facilitate susceptibility to scrapie transmission. In this study, investigated the infectivity of decreasing doses of the x124 scrapie agent (100 mg, 50 mg, 20 mg, and 10 mg) on incubation time and attack rate after experimental intralingual inoculation into VRQ/ARQ sheep. The lowest inoculum dose tested in this study effectively transmitted the x124 scrapie agent in VRQ/ARQ sheep with a 100% attack rate and no significant difference in incubation times among sheep inoculated with varying doses. Moreover, immunohistochemistry and western blot analysis revealed similar biochemical and immunohistochemical features among the four cohorts of sheep irrespective of inoculum dose. This study provides a starting point for further investigation to determine the minimum infectious dose of x124 scrapie in sheep and its effect on attack rate and incubation time, central for assessing the potential risk of scrapie occurrence in sheep flock.

Full restoration of specific infectivity and strain properties from pure mammalian prion protein.

The protein-only hypothesis predicts that infectious mammalian prions are composed solely of PrPSc, a misfolded conformer of the normal prion protein, PrPC. However, protein-only PrPSc preparations lack significant levels of prion infectivity, leading to the alternative hypothesis that cofactor molecules are required to form infectious prions. Here, we show that prions with parental strain properties and full specific infectivity can be restored from protein-only PrPSc in vitro. The restoration reaction is rapid, potent, and requires bank vole PrPC substrate, post-translational modifications, and cofactor molecules. To our knowledge, this represents the first report in which the essential properties of an infectious mammalian prion have been restored from pure PrP without adaptation. These findings provide evidence for a unified hypothesis of prion infectivity in which the global structure of protein-only PrPSc accurately stores latent infectious and strain information, but cofactor molecules control a reversible switch that unmasks biological infectivity.

MiRNA expression profiles in the brains of mice infected with scrapie agents 139A, ME7 and S15.

MicroRNA (miRNA) is a class of non-coding endogenous small-molecule single-stranded RNA that regulates complementary mRNA through degradation or translation of the mRNA targets. Usually, miRNAs show remarkable cell and tissues specificity. Recently, alterations in a set of miRNAs in the brains of patients with certain neurodegenerative diseases, including prion diseases, have been reported. In this study, using deep sequencing technology, miRNA expression profiles in the brains of mice infected with scrapie agents 139A, ME7 and S15 at a terminal stage were comparatively analysed. In total, 57, 94 and 135 differentially expressed miRNAs were identified in the pooled brain samples of 139A-, ME7- and S15-infected mice, respectively, compared with the brains of age-matched normal controls. Among them, 22 were commonly increased and 14 were commonly decreased in the brains of all three infected models. In addition, a reduction in the expression of two novel miRNAs was also commonly observed. Quantitative PCR with reverse transcription analysis of six randomly selected commonly increased and decreased miRNAs in the brains of the three infected mouse models, as well as the two novel miRNAs, verified that the expression patterns were comparable to the deep sequencing data. KEGG analysis of the differentially expressed miRNAs revealed the involvement of similar pathways in all three types of infected animals. Comprehensive analysis of these miRNA profiles not only provides useful clues for understanding prion biology but also is beneficial in the search for possible diagnostic marker(s) for prion diseases.