Search PubMed⌕ Search

Biomedical subjects

R A Somerville

Publications and source records attributed to R A Somerville.

At least 37 records · Page 2Linked to original sources
PubMed1992

Are Sinc and the PrP gene congruent? Evidence from PrP gene analysis in Sinc congenic mice.

Congenic mouse strains VM/Dk and VM-Sincs7/Dk differ at the Sinc gene, which controls the incubation period of scrapie in mice; VM/Dk mice are Sincp7p7 and VM-Sincs7/Dk mice are Sincs7s7. Restriction fragment length polymorphism and DNA sequencing analysis demonstrated that the PrP genes also differ in these strains, confirming the close genetic linkage of Sinc and PrP. Using the restriction enzyme HhaI, we have shown that at least 100 kb of DNA flanking the PrP gene differs between the two strains, and therefore the congruence of the Sinc and PrP genes is still not certain.

Animals↗
PubMed1990

Ubiquitin conjugate immunoreactivity in the brains of scrapie infected mice.

Sections of brain from normal mice or clinically-ill mice infected with either the 87V or the ME7 strains of sheep scrapie were immunostained to show the localization of ubiquitin-protein conjugates or a specific marker of disease, the scrapie-associated fibril protein (PrP). In both scrapie models immunoreactive ubiquitin-protein conjugates were seen in thread-like structures found throughout the neuropil, in inclusion bodies within vacuolated neurones, and in areas surrounding anti-PrP positive amyloid plaques. The PrP protein was visualized in diffuse deposits in highly vacuolated parts of the scrapie-affected brain, and focally in amyloid plaques, microglia and neuronal processes. The ubiquitin-protein conjugate staining of scrapie amyloid plaques is very similar to that seen in the plaques of Alzheimer's disease. The ubiquitinated intraneuronal inclusion bodies seen in scrapie resemble the granulovacuolar lesions also seen in Alzheimer's disease, but appear much larger and possibly correspond to material in giant autophagic vacuoles. We suggest that these inclusions may be the result of ubiquitinated abnormal proteins being directed to the lysosomal system, and that scrapie and Alzheimer's disease share at least some common processes of neurodegeneration.

Amyloid↗
PubMed1990

Differential glycosylation of the protein (PrP) forming scrapie-associated fibrils.

PrP is a glycoprotein found in normal brain. In brain affected by scrapie it forms scrapie-associated fibrils (SAF). PrP from SAF shows considerable heterogeneity of size and charge on two-dimensional gels. It separates into six major regions, the three more acidic regions arising as a result of partial proteolytic degradation. The two more basic higher Mr forms (Mr 34,000 and 29,000) of PrP can be reduced in apparent Mr to a lower Mr form (Mr 25,000) with Peptide-N-glycosidase F. In addition, a series of lectins has been found to bind to PrP. Some bind preferentially to the higher Mr forms whereas others bind more strongly to the lower Mr form. Some of the heterogeneity of PrP is therefore due to differential N-glycosylation. We suggest that one or two N-linked carbohydrate chains are bound to the protein causing some of the differences in Mr. The major cause of heterogeneity of PrP is therefore proteolytic cleavage combined with differential glycosylation at the two potential N-glycosylation sites. The glycolipid moiety attached to PrP may be responsible for some lectin binding to all three bands. Using lectins as a probe to study potential differences in N-glycosylation we have looked at their binding to PrP isolated from SAF, from different strains of scrapie and from different regions of the same brain. No major differences in the N-glycan moieties were found.

Animals↗
PubMed1989

Structural and biochemical evidence that scrapie-associated fibrils assemble in vivo.

Scrapie-associated fibrils (SAF) are a ubiquitous pathological feature of brains affected by scrapie and the other scrapie-like agents. They are composed of PrP, a heterogeneous glycoprotein which is also present in normal brain but not as SAF. The PrP protein associated with SAF is partially resistant to proteinase K, whereas the soluble form is not. It has been proposed that SAF do not exist as such in vivo, but rather self-assemble from subunit structures liberated from membranes by detergent extraction during purification. We have purified SAF by a method that does not employ proteinase K. We show that the PrP protein from infected but not uninfected brain is partially resistant to protease digestion before and after detergent extraction. Likewise, SAF can be sheared by sonication before or after detergent extraction. In addition, SAF from mice infected with different strains of scrapie have different sedimentation properties. Since SAF-dependent properties exist before detergent extraction, then so must SAF. They are therefore not a detergent-induced artefact but most probably assemble in vivo.

Animals↗
PubMed1987

Scrapie associated fibrils in the diagnosis of scrapie in sheep.

Previous research has consistently demonstrated by electron microscopy the presence of scrapie associated fibrils in brain extracts prepared from mice and hamsters with clinical signs of experimental scrapie. In the present study similar fibrils were seen in all the brain extracts prepared from 11 Cheviot or Suffolk sheep with natural or experimental scrapie that had been diagnosed clinically and confirmed neuropathologically. They were not found in the brain extracts of nine sheep that did not have scrapie and which included four that had been injected with infected material but did not develop the disease. The presence of such fibrils can therefore be used as an additional diagnostic criterion for natural scrapie in sheep.

Animals↗
PubMed1986

Partial copurification of scrapie-associated fibrils and scrapie infectivity.

The association between scrapie infectivity and scrapie-associated fibrils (SAF) during a partial purification procedure for infectivity was investigated. Scrapie infectivity and SAF can be separated from most membrane components by subcellular fractionation of infected mouse brain to obtain a synaptosomal fraction, followed by detergent treatment and density gradient centrifugation. After different detergent treatments, with either octyl glucoside or sodium N-lauroyl sarcosinate, SAF showed differing sedimentation characteristics but nevertheless cosedimented with scrapie infectivity in both cases. Copurification under two different conditions provides more evidence that SAF may be a form of the infectious agent of scrapie.

Animals↗
PubMed1985

Ultrastructural links between scrapie and Alzheimer's disease.

The discovery of abnormal fibrillar structures, scrapie-associated fibrils (SAF), in fractions with high infectivity from scrapie-infected brains has led to the proposal that SAF are a form of the infectious agent. On the basis of this proposal and on the congophilia shared by SAF and amyloid, it has been speculated elsewhere that the amyloid in Alzheimer's disease is infectious. This speculation is not supported by available evidence and therefore a conventional origin for the amyloid in Alzheimer's disease is favoured--that it originates by partial degradation of a host protein, as occurs in all other forms of amyloidosis characterised so far.

Alzheimer Disease↗
PubMed1985

Brain glutamate decarboxylase and cholinergic enzyme activities in scrapie.

C57BL/6J mice, age 6-8 weeks were inoculated intracerebrally with brain homogenate from mice previously infected with the 139A strain of scrapie; control mice were identically treated with brain homogenate from non-infected normal mice. The activities of choline acetyltransferase (CAT), acetyl cholinesterase (AChE), and glutamic acid decarboxylase (GAD) were determined in the forebrain and hindbrain of these animals after 67, 126 and 151 days post-inoculation. There were no significant differences in the activities of CAT and GAD between scrapie and control mice at early, middle or late stages of the disease in the scrapie-infected animals; there was an about 20% decline in AChE activity in the scrapie brain.

Acetylcholinesterase↗
PubMed1984

Infection-specific particle from the unconventional slow virus diseases.

Scrapie-associated fibrils, first observed in brains of scrapie-infected mice, were also observed in scrapie-infected hamsters and monkeys, in humans with Creutzfeldt-Jakob disease, and in kuru-infected monkeys. These fibrils were not found in a comprehensive series of control brains from humans and animals affected with central nervous system disorders resulting in histopathologies, ultrastructural features, or disease symptoms similar to those of scrapie, kuru, and Creutzfeldt-Jakob disease. These fibrils are also found in preclinical scrapie and in the spleens of scrapie-infected mice; they are a specific marker for the "unconventional" slow virus diseases, and may be the etiological agent.

Alzheimer Disease↗
PubMed1984

The effects of detergents on the composition of postsynaptic densities.

A method of purifying postsynaptic densities (PSD) of Cohen et al. (1977) has been modified, primarily by the substitution of octyl glucoside as the detergent used to solubilize synaptosomal fractions. Subsequent extraction with other detergents resulted in the selective removal of specific polypeptides. In particular sulphobetaine 3-14 removed most of the beta-tubulin but not alpha-tubulin. Sodium N-lauroyl sarcosinate completely destroyed the structural integrity of the PSD when the in vitro formation of intermolecular disulphide bonds was minimized. These results suggest that the structure of PSDs is more labile than previously thought and demonstrate a technique for further examining their composition.

Animals↗
PubMed1984

Decreased high-affinity binding of [3H]muscimol to cerebral synaptic membranes of scrapie-infected mice.

Scrapie is a transmissible disease that results in progressive degeneration of the central nervous system and death. Although scrapie has been studied histopathologically, relatively little is known concerning neurotransmitter alterations. Specific [3H]muscimol binding to whole brain crude synaptic membranes (CSM) from mice clinically affected with scrapie was significantly (p less than 0.01) reduced, to approximately 73% of that of the controls. Of the brain regions examined, binding to only cerebral CSM was significantly (p less than 0.0001) decreased. Scatchard analyses of saturation curves revealed that the high-affinity (KD = 8 +/- 3 nM) site for muscimol was abolished in cerebral CSM from scrapie-infected mice, while the low-affinity site was unaffected. Binding of [3H]flunitrazepam to cerebral CSM was unaffected by scrapie and was stimulated by GABA to the same extent in both scrapie and control mice. These results suggest that scrapie agent 139A in C57BL/6J mice manifests a portion of its CNS pathology via a high-affinity GABA binding site that is unassociated with the benzodiazepine receptor.

Animals↗
PubMed1983

Ultrastructural morphology of amyloid fibrils from neuritic and amyloid plaques.

The structure of partially purified, CNS amyloid fibrils from three different sources have been compared by negative stain EM. The fibrils isolated from brains with senile dementia of Alzheimer type were 4-8 nm in diameter, narrowing every 30-40 nm and apparently composed of two 2-4 nm filaments. The fibrils from a Gerstmann-Sträussler syndrome brain were 7-9 nm in diameter, narrowing every 70-80 nm and with a suggestion that they are composed of two 3-5 nm filaments. The fibrils isolated from 87V scrapie-affected mouse brains were 4-8 nm in diameter with a twist every 15-25 nm presumably composed of two 2-4 nm filaments. The fibrils from the scrapie brains were usually observed in pairs. The shape of the clusters of the isolated amyloid fibrils observed in each disease was similar in negative stain and thin section EM preparations and was related to the characteristic morphology of the amyloid fibrils in the neuritic and amyloid plaques in situ. The structural differences between the CNS amyloid fibrils from the various diseases studied by us may reflect differences in the polypeptides which comprise the fibril and/or a different pathogenesis in the formation of the amyloid fibrils.

Alzheimer Disease↗
PubMed1983

Altered scrapie infectivity estimates by titration and incubation period in the presence of detergents.

During experiments on the purification of scrapie infectivity, changes were found in the dynamics of scrapie titration. After exposure to detergent, infectivity estimates by both endpoint titration and incubation period were altered. The addition of detergent to the diluent used in titration resulted in at least a 100-fold increase in the infectivity estimate. This suggests that the amount of scrapie in a sample, as measured by serial dilution and titration, may be underestimated to different extents, depending on the biochemical milieu of the inoculum. Membrane fractions treated with detergents before dilution exhibited longer incubation periods than untreated fractions for the same number of infectious units of scrapie. This demonstrates that detergent treatments and possibly other biochemical manipulations can cause changes in the response of the host to the inoculum that are not detectable if incubation periods alone are used to estimate scrapie 'titre'.

Animals↗
PubMed1981

Abnormal fibrils from scrapie-infected brain.

Abnormal fibrillary structures, designated "scrapie-associated fibrils" (SAF), have been observed using negative stain techniques in subfractions of brains from scrapie-affected animals. SAF have been observed in all combinations of strain of scrapie agent and strain or species of host examined, regardless of their histopathology, in particular the presence or absence of amyloid plaques. SAF consist either of two or four filaments. They are morphologically dissimilar to the normal brain fibrils--microtubules, neurofilaments, glial filaments, and F actin. However, SAF do bear a resemblance to amyloid.

Amyloid↗
PubMed1980

Sensitivity of scrapie infectivity to detergents and 2-mercaptoethanol.

Exposure of fractions obtained from scrapie-infected mouse brain to Triton X-100 resulted in no change in its infectivity except after sedimentation. Infectivity was reduced after exposure to 2-mercaptoethanol but only in the presence of SDS. These results support the view that protein is an important component of the scrapie agent and that disulfide bonds may play a part in maintaining the structural integrity of the infectious agent.

Animals↗
PubMedDate not supplied

Post-mortem immunodiagnosis of scrapie and bovine spongiform encephalopathy.

Two polyclonal antisera were raised in rabbits against the scrapie-associated fibril protein (PrP) prepared from sheep and mice which were terminally infected with experimental scrapie. The anti-mouse PrP serum identifies the proteins of scrapie-associated fibrils (SAF) from all the host species studied (mouse, hamster, sheep and goat) and bovine spongiform encephalopathy (BSE) fibrils from cow. The anti-sheep PrP serum displays species restricted immunoreactivity. While it identifies several PrP polypeptides from terminally affected sheep, goat and cow material, only the highest molecular weight band is recognised from hamster and there is no detection of mouse PrP. The use of these antisera in routine laboratory testing at post mortem provides a highly sensitive test for scrapie and BSE and may allow the identification of infected animals prior to the onset of clinical signs.

Animals↗