Search

In prion diseases, metal imbalances in brain and/or metal substitutions for copper in prion protein suggest that metal-catalyzed oxidation (MCO) and oxidative stress may affect cellular function and accumulation of protease-resistant prion protein (PrP(Sc)). We examined the effect of metal-induced oxidative stress by Fenton reaction on prion protein with regard to its degradation, insolubility, and infectivity. Precipitation and insolubility of prion protein were induced by Fenton reaction in scrapie-infected brain homogenate. Results showed an increase in hydroxylation products (thiobarbituric acid reactive substances; TBARS) and a decrease of ferrous ion (Fe(2+)) levels after Fenton reaction. Efficiency of metal-induced oxidation was higher for Fe(2+) than Mn(2+). Compared to untreated samples, there was increased susceptibility to proteolytic degradation of PrP(Sc) after treatment with 3.12-12.5 mM Fe(2+)-Mn(2+)/H(2)O(2). Interestingly, we observed that Fenton reaction could extend incubation periods, indicating a decrease in scrapie infectivity. These results suggest that PrP(Sc) hydroxylation and degradation may affect PrP conversion and the pathogenesis of prion diseases.

Scrapie is a fatal, but slow, infectious disease. C57BL/6J, SAMP8 (a strain that develops early senescence), SAMR1 (a strain that is resistant to senescence) and AKR/J (a progenitor of the SAM strains) mice were infected with 22A, 139A, 22L and ME7 scrapie strains. Histopathological stains included haematoxylin and eosin (HE), and periodic acid-Schiff (PAS). Vacuolation was found in the brains of all scrapie-infected mice. The 22A strain caused more extensive vacuolation in the brains of SAMP8 and SAMR1 mice than in C57BL mice. PAS-positive plaques (PP) were found in 22A-infected mice in cortex, corpus callosum, hippocampus, subependymal zone area and thalamus. PP were significantly increased in 22A-infected SAMR1 mice compared to mice from other scrapie-infected strains. Clusters of small, round, homogeneous PAS-positive granular structures (PGS) were found in all mouse strains, especially in aging control and 22A-infected C57BL mice, predominantly in the stratum radiatum of the CA1, CA2 and CA3 areas of the hippocampus. Some of these structures were also observed in stratum oriens and piriform cortex, and in cerebellar Purkinje cell areas. Some of the PGS were associated with astrocytes and blood vessels. Each granule was 1-5 microm in diameter and there were clusters consisting of several to 40 PGS; the sizes of the clusters ranged from 10 to 80 microm in diameter. There were more PGS clusters in uninfected C57BL and AKR mice than in uninfected SAMP8 and SAMR1 mice. PGS were not increased in scrapie-infected mice. These findings suggest that PGS accumulation was more dependent on the genetic information of the mouse strain, whereas PP and vacuolation patterns depended on the scrapie strain-mouse strain combination.

Scrapie is a fatal neurodegenerative disease of sheep and goats. The precise details of neuronal and neurite degeneration in scrapie-infected animals remain unknown. Using specific silver staining methods, we compared the neurodegeneration caused by treatment of rats with kainic acid (KA) or ibogaine (IBO) to the neuropathology observed in mice infected with the C602 strain of scrapie. As reported previously, KA resulted in extensive silver labeling of neurons, especially in the cortex, putamen and hippocampus. IBO silver labeling was observed only in small clusters of Purkinje neurons in the paravermal region of the cerebellum. However, in scrapie-infected mice, a few silver stained neurons (differing from the dark degenerating neurons observed following neurotoxic exposure) were found in layer II of cortex, cingulate cortex, zona incerta, thalamus and hypothalamus. Some silver grains were observed in glial-like cells, especially those in the paraventricular region. Degenerating axons were positive for silver staining and were found in the cortex, cingulate cortex, corpus callosum, habenulae, septum, fornix, thalamus, caudate putamen and a few in fasciculus retroflexus and substantia nigra. Our results suggest that the limbic system is one of the important loci for the neurodegenerative effect of at least some scrapie strains.

Scrapie is a transmissible neurodegenerative disease of sheep and goats. The neuropathological changes include vacuolation, astrocytosis, the development of amyloid plaques in some instances, and neuronal loss. The mechanisms involved in neuronal cell death in scrapie are not known. Recently, we reported the presence of oxidative stress in the brains of scrapie-infected animals and suggested that this is the main mechanism that induces neuronal cell loss. It is known that oxidative stress induced by free radicals is associated with iron accumulation; this association led to an examination of the levels of iron (total iron, Fe(2+) and Fe(3+)) in the brains of control and scrapie-infected mice by biochemical methods. In the scrapie-infected group, both the level of total iron and the Fe(3+) level were significantly increased in cerebral cortex, striatum, and brainstem as compared to the values in the control group. A shift in the ratio of Fe(2+)/Fe(3+) was observed in the same regions of infected mice. Additionally, in this scrapie model, we confirmed the presence of oxidative stress, as evidenced by the increase of free malondialdehyde. These results suggest that iron metabolism is changed and that iron-induced oxidative stress partly contributes to neurodegeneration in scrapie infection.

In scrapie infection, prion protein (PrPSc) is localized in areas where there is neurodegeneration and astrocytosis. It is thought that PrPSc is toxic to neurons and trophic for astrocytes. In our study, paraffin sections from scrapie infected (263K and 139H) and control hamsters were examined with histological and immunocytochemical staining. We found that PrPSc was present in the ependymal cells of both 263K- and 139H-infected hamsters. In 139H-infected hamsters, PrPSc was found in the cytoplasm of neurons in cerebral cortex and in hypothalamic paraventricular (PVN) and supraoptic (SON) nuclei. In contrast, neuronal cytoplasm and nuclei, were positive for PrPSc in most areas such as cortex, hippocampus, and thalamus in 263K-infected hamsters. Many aggregations of PrPSc could be seen in the cortex, hippocampus, substantia nigra and around the Pia mater, corpus callosum, fimbria, ventricles, and blood vessels in sections from 139H- and/or 263K-positive animals. Furthermore, PrPSc was also co-localized with glial fibrillary acidic protein (GFAP) in many reactive astrocytes (approximately 90%) in certain areas such as the hippocampus in 263K-infected hamsters, but not 139H-infected hamsters. The patterns of astrocytosis and PrPSc formation were different between 139H- and 263K-infected hamsters, which may be used for a diagnosis purpose. Our results suggest a hypothesis that multiple cell-types are capable of PrPSc production. Our results also confirm that reactive astrocytes can produce and/or accumulate PrPSc during some scrapie strain infections. The findings suggest a 'snowball effect', that is: astrocytosis might play an important role in amyloidosis, while amyloidosis may induce further astrocytosis at least in 263K-infected hamsters.

Considerable controversy exists concerning the origin and composition of Alzheimer neurofibrillary tangles (ANT) and of paired helical filaments (PHF), the abnormal cytoplasmic fibers which ultrastructurally are the major components of ANT. Thus far, the unusual solubility properties of PHF have hindered the analysis of ANT. A new procedure is presented for isolating purified PHF which are soluble in the presence of sodium dodecyl sulfate. The purification protocol involves differential and rate zonal centrifugation, treatment with the detergents sarcosyl and sulfobetain 3-14, and sonication. The isolated PHF from Alzheimer disease/senile dementia of the Alzheimer type (7 cases) and Down's syndrome (one case) have been characterized structurally by negative-stain electron microscopy, biochemically by PAGE, and immunologically by both the ELISA technique and Western blot analysis using a monoclonal antibody prepared against ANT. Distinct polypeptides were shown to be associated with this structure and not seen in preparations from young and age-matched normal brains.

The 22L strain of scrapie was injected stereotaxically into the cerebellum of C57BL/6J mice to determine its effect on several cerebellar neurotransmitter systems during the early clinical stages of the disease. In this model vacuolar lesions are restricted to the cerebellum with no evidence of vacuolization in other brain regions. Although vacuolar lesions develop throughout all cell layers of the cerebellum, they are most severe in the granule cell layer. Modest but significant (P less than 0.01) reductions in cerebellar weight, glutamate decarboxylase activity, and in the affinity of the N6-[adenine-2,8-3H]cyclohexyladenosine binding sites, were observed in scrapie affected mice. The densities of the high- and low-affinity adenosine receptors were unaffected. Adenosine receptors in the cerebellum are highly localized to the axon terminals of the glutamatergic, GABA receptive granule cells. GABA, benzodiazepine, glutamate, and muscarinic cholinergic receptors were not significantly altered. In addition, the high-affinity uptake of glutamate, and the activity of choline acetyltransferase were not significantly changed. GABA high-affinity uptake was slightly increased. Even though the granule cell layer of the cerebellum had undergone severe vacuolation, only modest neurotransmitter changes were apparent. Although these results suggest a tenuous relationship between scrapie pathology and the integrity of neurotransmitter systems, it is possible that compensatory neurochemical changes in uncompromised neuronal populations may have masked potentially specific neurotransmitter effects.