Mouse Model Of Prion Disease Research Articles

Infectivity versus toxicity

Prions are infectious proteins that can cause deadly diseases in mammals. Detailed measurements of infectivity suggest that there may be distinct infectious and toxic versions of this protein. See Letter p.540 Prion infections have a clinically silent incubation period that can go on for years or even decades, followed by an aggressive, short clinical phase. Experiments in the RML mouse model of prion disease now show that prion propagation in the brain proceeds by two distinct phases: a relatively brief exponential phase that is not rate-limited by prion protein concentration, followed by a plateau phase. Surprisingly, it is the latter that can be very prolonged, accounting for the majority of the clinically silent incubation period. The similar levels of infectivity at the end of the first and second phase suggest that there is a separation between prion infectivity and toxicity. The authors suggest that the prions are not neurotoxic themselves, but catalyse the formation of such species from host-cell-encoded cellular prion protein.

Striatal pathology underlies prion infection-mediated hyperactivity in mice

Although prion diseases are most commonly modeled using the laboratory mouse, the diversity of prion strains, behavioral testing and neuropathological assessments hamper our collective understanding of mouse models of prion disease. Here we compared several commonly used murine strains of prions in C57BL/6J female mice in a detailed home cage behavior detection system and a systematic study of pathological markers and neurotransmitter systems. We observed that mice inoculated with RML or 139A prions develop a severe hyperactivity phenotype in the home cage. A detailed assessment of pathology markers, such as microglial marker IBA1, astroglial marker GFAP and degeneration staining indicate early striatal pathology in mice inoculated with RML or 139A but not in those inoculated with 22L prions. An assessment of neuromodulatory systems including serotonin, dopamine, noradrenalin and acetylcholine showed surprisingly little decline in neuronal cell bodies or their innervations of regions controlling locomotor behavior, except for a small decrease in dopaminergic innervations of the dorsal striatum. These results implicate the dorsal striatum in mediating the major behavioral phenotype of 139A and RML prions. Further, they suggest that measurements of activity may be a sensitive manner in which to diagnose murine prion disease. With respect to neuropathology, our results indicate that pathological stains as opposed to neurotransmitter markers are much more informative and sensitive as markers of prion disease in mouse models.

Manganese chelation therapy extends survival in a mouse model of M1000 prion disease

Previous in vitro and in vivo investigations have suggested manganese (Mn(2+)) may play a role in pathogenesis through facilitating refolding of the normal cellular form of the prion protein into protease resistant, pathogenic isoforms (PrP(Sc)), as well as the subsequent promotion of higher order aggregation of these abnormal conformers. To further explore the role of Mn(2+) in pathogenesis, we undertook a number of studies, including an assessment of the disease modifying effects of chelation therapy in a well-characterized mouse model of prion disease. The di-sodium, calcium derivative of the chelator, cyclohexanediaminetetraacetic acid (Na(2)CaCDTA), was administered intraperitoneally to mice inoculated intra-cerebrally with either high or low-dose inocula, with treatment beginning early (shortly after inoculation) or late (at the usual mid-survival point of untreated mice). Analyses by inductively coupled plasma-mass spectrometry demonstrated brain Mn(2+) levels were selectively reduced by up to 50% in treated mice compared with untreated controls, with copper, iron, zinc and cobalt levels unchanged. In mice administered high-dose inocula, none of the treatment groups displayed an increase in survival although western blot analyses of early intensively treated mice showed reduced brain PrP(Sc) levels; mice infected using low-dose inocula however, showed a significant prolongation of survival (p = 0.002). Although our findings support a role for Mn(2+) in prion disease, further studies are required to more precisely delineate the extent of pathogenic involvement.

Systemic inflammation induces acute working memory deficits in the primed brain: relevance for delirium

Delirium is an acute, severe neuropsychiatric syndrome, characterized by cognitive deficits, that is highly prevalent in aging and dementia and is frequently precipitated by peripheral infections. Delirium is poorly understood and the lack of biologically relevant animal models has limited basic research. Here we hypothesized that synaptic loss and accompanying microglial priming during chronic neurodegeneration in the ME7 mouse model of prion disease predisposes these animals to acute dysfunction in the region of prior pathology upon systemic inflammatory activation. Lipopolysaccharide (LPS; 100 μg/kg) induced acute and transient working memory deficits in ME7 animals on a novel T-maze task, but did not do so in normal animals. LPS-treated ME7 animals showed heightened and prolonged transcription of inflammatory mediators in the central nervous system (CNS), compared with LPS-treated normal animals, despite having equivalent levels of circulating cytokines. The demonstration that prior synaptic loss and microglial priming are predisposing factors for acute cognitive impairments induced by systemic inflammation suggests an important animal model with which to study aspects of delirium during dementia.

Development of oligomeric prion‐protein aggregates in a mouse model of prion disease

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.

The role of calorie restriction and SIRT1 in prion-mediated neurodegeneration

A central focus of aging research is to determine how calorie restriction (CR) extends lifespan and delays diseases of aging. SIRT1, the mammalian ortholog of Sir2 in yeast, is a longevity factor which mediates dietary restriction in diverse species. In addition, SIRT1 plays a protective role in several models of neurodegenerative disease. We tested the role of SIRT1 in mediating the effects of CR in a mouse model of prion disease. Prion diseases are protein misfolding disorders of the central nervous system with many similarities to other neurodegenerative diseases, including deposition of aggregated protein, gliosis, and loss of synapses and neurons. We report that the onset of prion disease is delayed by CR and in the SIRT1 KO mice fed ad libitum. CR exerts no further effect on the SIRT1 KO strain, suggesting the effects of CR and SIRT1 deletion are mechanistically coupled. In conjunction, SIRT1 is downregulated in certain brain regions of CR mice. The expression of PrP mRNA and protein is reduced in the brains of CR mice and in SIRT1 knockout mice, suggesting a possible mechanism for the delayed onset of disease, as PrP levels are a critical determinant of how quickly mice succumb to prion disease. Surprisingly, CR greatly shortens the duration of clinical symptoms of prion disease and ultimately shortens lifespan of prion-inoculated mice in a manner that is independent of SIRT1. Taken together, our results suggest a more complex interplay between CR, SIRT1, and neurodegenerative diseases than previously appreciated.

Galectin-3 expression is correlated with abnormal prion protein accumulation in murine scrapie

To investigate the involvement of galectin-3 in the process of neurodegeneration in prion diseases, the expression and cellular localization of galectin-3 in the brain were studied in scrapie, a mouse model of prion disease. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analyses showed that the expression of galectin-3 protein and mRNA was induced in scrapie-affected brains, particularly at the time when the abnormal prion protein PrP(Sc) began to accumulate in the brains. Immunohistochemically, immunostaining for galectin-3 was found mainly in B4-isolectin-positive cells (presumably activated microglia/macrophages), but not in astrocytes. Galectin-3 immunoreactivity was localized mainly in areas of PrP(Sc) accumulation and neuronal death in scrapie-infected brains. These findings suggest that the expression of galectin-3 by activated microglia/macrophages in prion disease correlates with abnormal prion protein accumulation.

Decreased cell surface prion protein in mouse models of prion disease

Transmissible spongiform encephalopathies are infectious neurodegenerative diseases caused by prions, composed of ordered aggregates of misfolded cellular prion protein. Neural antigen density of prion protein, Thy-1 and glial fibrillary acidic protein was analyzed using flow cytometry of dissociated mouse brain cells after inoculation with mouse-adapted transmissible spongiform encephalopathy agents. Transmissible spongiform encephalopathy gliosis was demonstrated by increased intracellular immunoreactivity for glial fibrillary acidic protein compared with controls. Immunoreactivity for cell surface prion protein was reduced 2.8-3.8-fold compared with control brain cells, whereas surface Thy-1 protein was reduced 1.5-4-fold. Double-staining protocols revealed loss of brain cells highly immunoreactive for prion protein and Thy-1, with a preferential reduction of prion protein, suggesting that prion protein expression, trafficking or consumption may be affected early in disease.

Migration of dendritic cells into the brain in a mouse model of prion disease

The immune system plays a key role in the dissemination of prion infections from the periphery to the central nervous system (CNS). While follicular dendritic cells are critical for prion replication in lymphoid tissue and subsequent neuroinvasion, myeloid dendritic cells (DCs) have been implicated in both the clearance and propagation of pathological prion protein. Since nothing is known on the ability of DCs to migrate to the CNS during prion diseases, we investigated the immunohistochemical localization of CD205 + DCs in the brain of C57BL/6 mice intraperitoneally infected with the mouse-adapted KFu strain of Gerstmann–Sträussler–Scheinker syndrome, a human genetic prion disorder. In normal brain, CD205 + cells were present in the meninges and choroid plexus, whereas in the majority of mice sacrificed between 120 and 300 days post infection, CD205 + DCs were also detected in the cerebral cortex, subcortical white matter, thalamus and medulla oblongata. These findings demonstrate that DCs can enter the CNS of prion-infected mice, suggesting a possible role for these cells in the pathogenesis of prion disorders.

The acute inflammatory response in CNS following injection of prion brain homogenate or normal brain homogenate.

The neuropathological hallmarks of end-stage prion disease are vacuolation, neuronal loss, astrocytosis and deposition of PrPSc amyloid. We have also shown that there is an inflammatory response in the brains of scrapie-affected mice from 8 weeks post-injection. In this study we have investigated the acute CNS response to the intracerebral injection of scrapie-affected brain homogenate. The ME7 strain of scrapie (Neuropathogenesis Unit, Edinburgh) was used, and control mice were injected with brain homogenate derived from normal C57BL/6 J mice. One microlitre of 10% w/v ME7 (n = 33) and normal brain homogenate (n = 28) was injected stereotaxically into the right dorsal hippocampus. Cryostat sections of brains taken at 1, 2, 5, 7, 14 and 28 days post-injection were examined histologically for neuronal loss, and immunocytochemically to study the inflammatory response. This study shows that ME7 is not acutely neurotoxic in vivo. There is also no difference (ANOVA) in the inflammatory response, which peaked between 2 and 5 days and resolved by 4 weeks after intracerebral injection of either ME7 or normal brain homogenate. The well circumscribed inflammatory response seen previously at 8 weeks is therefore a consequence of a disease process rather than a surgical artefact. This disease process may be related to a localized accumulation of PrPSc sufficient to stimulate an inflammatory response which in turn may contribute to neuronal loss. The role of the inflammatory response in chronic neurodegeneration can be usefully studied using this mouse model of prion disease, and this will undoubtedly shed light on the pathogenic mechanisms underlying other chronic neurodegenerative diseases.