Browsing by Person "Barron, Rona"

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The 101L mutation in murine PrP can alter transmission across three species barriers
(2002) Barron, Rona; Jamieson, Elizabeth; Thomson, Val; Melton, David W.; Will, Robert; Ironside, James; Manson, Jean C.
Accumulation of prion protein in the brain that is not associated with transmissible disease
(National Academy of Sciences, 2007-03-13) Piccardo, Pedro; King, Declan; Ghetti, Bernardino; Manson, Jean C.; Barron, Rona
Prion diseases or transmissible spongiform encephalopathies are characterized histopathologically by the accumulation of prion protein (PrP) ranging from diffuse deposits to amyloid plaques. Moreover, pathologic PrP isoforms (PrPSc) are detected by immunoblot analysis and used both as diagnostic markers of disease and as indicators of the presence of infectivity in tissues. It is not known which forms of PrP are associated with infectivity. To address this question, we performed bioassays using human brain extracts from two cases with phenotypically distinct forms of familial prion disease (Gerstmann-Sträussler-Scheinker P102L). Both cases had PrP accumulations in the brain, but each had different PrPSc isoforms. Only one of the brains had spongiform degeneration. Tissue from this case transmitted disease efficiently to transgenic mice (Tg PrP101LL), resulting in spongiform encephalopathy. In contrast, inoculation of tissue from the case with no spongiform degeneration resulted in almost complete absence of disease transmission but elicited striking PrP-amyloid deposition in several recipient mouse brains. Brains of these mice failed to transmit any neurological disease on passage, but PrP-amyloid deposition was again observed in the brains of recipient mice. These data suggest the possible isolation of an infectious agent that promotes PrP amyloidogenesis in the absence of a spongiform encephalopathy. Alternatively, the infectious agent may be rendered nonpathogenic by sequestration in amyloid plaques, or PrP amyloid can seed amyloid accumulation in the brain, causing a proteinopathy that is unrelated to prion disease. Formation of PrP amyloid may therefore not necessarily be a reliable marker of transmissible spongiform encephalopathy infectivity.
Bovine PrP expression levels in transgenic mice influence transmission characteristics of atypical bovine spongiform encephalopathy
(Microbiology Society, 2012-05-01) Wilson, Rona; Hart, Patricia; Piccardo, Pedro; Hunter, Nora; Casalone, Cristina; Baron, Thierry; Barron, Rona
Until recently, transmissible spongiform encephalopathy (TSE) disease in cattle was thought to be caused by a single agent strain, bovine spongiform encephalopathy (BSE) (classical BSE or BSE-C). However, due to the initiation of a large-scale surveillance programme throughout Europe, two atypical BSE strains, bovine amyloidotic spongiform encephalopathy (BASE, also named BSE-L) and BSE-H have since been discovered. These atypical BSE isolates have been previously transmitted to a range of transgenic mouse models overexpressing PrP from different species at different levels, on a variety of genetic backgrounds. To control for genetic background and expression level in the analysis of these isolates, we performed here a comprehensive comparison of the neuropathological and molecular properties of all three BSE agents (BASE, BSE-C and BSE-H) upon transmission into the same gene-targeted transgenic mouse line expressing the bovine prion protein (Bov6) and a wild-type control of the same genetic background. Significantly, upon challenge with these BSE agents, we found that BASE did not produce shorter survival times in these mice compared with BSE-C, contrary to previous studies using overexpressing bovine transgenic mice. Amyloid plaques were only present in mice challenged with atypical BSE and neuropathological features, including intensity of PrP deposition in the brain and severity of vacuolar degeneration were less pronounced in BASE compared with BSE-C-challenged mice.
Changing a single amino acid in the N-terminus of murine PrP alters incubation time across three species barriers
(EMBO Press, 2001-09-17) Barron, Rona; Thomson, Val; Jamieson, Elizabeth; Melton, David W.; Ironside, James; Will, Robert; Manson, Jean C.
The PrP gene of the host exerts a major influence over the outcome of transmissible spongiform encephalopathy (TSE) disease, but the mechanism by which this is achieved is not understood. We have introduced a specific mutation into the endogenous murine PrP gene using gene targeting to produce transgenic mice with a single amino acid alteration (proline to leucine) at amino acid position 101 in their PrP protein (P101L). The effect of this alteration on incubation time, targeting and PrPSc formation has been studied in TSE-infected animals. Transgenic mice carrying the P101L mutation in PrP have remarkable differences in incubation time and targeting of central nervous system pathology compared with wild-type littermates, following inoculation with infectivity from human, hamster, sheep and murine sources. This single mutation can alter incubation time across three species barriers in a strain-dependent manner. These findings suggest a critical role for the structurally ‘flexible’ region of PrP in agent replication and targeting of TSE pathology.
Characterization of an unusual transmissible spongiform encephalopathy in goat by transmission in knock-in transgenic mice
(Microbiology Society, 2013-08-01) Wilson, Rona; King, Declan; Hunter, Nora; Goldmann, Wilfred; Barron, Rona
Bovine spongiform encephalopathy (BSE) is a fatal neurodegenerative disorder of cattle, and its transmission to humans through contaminated food is thought to be the cause of the variant form of Creutzfeldt–Jakob disease. BSE is believed to have spread from the recycling in cattle of ruminant tissue in meat and bone meal (MBM). However, during this time, sheep and goats were also exposed to BSE-contaminated MBM. Both sheep and goats are experimentally susceptible to BSE, and while there have been no reported natural BSE cases in sheep, two goat BSE field cases have been documented. While cases of BSE are rare in small ruminants, the existence of scrapie in both sheep and goats is well established. In the UK, during 2006–2007, a serious outbreak of clinical scrapie was detected in a large dairy goat herd. Subsequently, 200 goats were selected for post-mortem examination, one of which showed biochemical and immunohistochemical features of the disease-associated prion protein (PrPTSE) which differed from all other infected goats. In the present study, we investigated this unusual case by performing transmission bioassays into a panel of mouse lines. Following characterization, we found that strain properties such as the ability to transmit to different mouse lines, lesion profile pattern, degree of PrP deposition in the brain and biochemical features of this unusual goat case were neither consistent with goat BSE nor with a goat scrapie herdmate control. However, our results suggest that this unusual case has BSE-like properties and highlights the need for continued surveillance.
Chronic wasting disease and atypical forms of bovine spongiform encephalopathy and scrapie are not transmissible to mice expressing wild-type levels of human prion protein
(Microbiology Society, 2012-07-01) Wilson, Rona; Plinston, Chris; Hunter, Nora; Casalone, Cristina; Corona, Cristiano; Tagliavini, Fabrizio; Suardi, SIlvia; Ruggerone, Margherita; Graziano, Silvia; Sbriccoli, Marco; Cardone, Franco; Pocchiari, Maurizio; Ingrosso, Loredana; Baron, Thierry; Richt, Juergen; Andreoletti, Olivier; Simmons, Marion; Lockey, Richard; Manson, Jean C.; Barron, Rona
The association between bovine spongiform encephalopathy (BSE) and variant Creutzfeldt–Jakob disease (vCJD) has demonstrated that cattle transmissible spongiform encephalopathies (TSEs) can pose a risk to human health and raises the possibility that other ruminant TSEs may be transmissible to humans. In recent years, several novel TSEs in sheep, cattle and deer have been described and the risk posed to humans by these agents is currently unknown. In this study, we inoculated two forms of atypical BSE (BASE and H-type BSE), a chronic wasting disease (CWD) isolate and seven isolates of atypical scrapie into gene-targeted transgenic (Tg) mice expressing the human prion protein (PrP). Upon challenge with these ruminant TSEs, gene-targeted Tg mice expressing human PrP did not show any signs of disease pathology. These data strongly suggest the presence of a substantial transmission barrier between these recently identified ruminant TSEs and humans.
Differential protein expression profiling in BSE disease
(Taylor and Francis, 2010-08-10) Barr, Janice B.; Waddington, Dave; Barron, Rona
Bovine spongiform encephalopathy (BSE) is a fatal neurodegenerative disease affecting cattle. Current tests for the detection of BSE are based solely on the only definitive marker of the disease, an abnormal conformer (PrPd), of the host encoded prion protein (PrPc). Recent evidence that other transmissible spongiform encephalopathy diseases can be present in the absence of PrPd, coupled with the need to establish pre-mortem diagnostic assays have led to a search for alternative diagnostic approaches. In this study we apply differential protein expression profiling for the prediction of BSE disease in post-mortem bovine brain tissue. The protein profiles of groups of 27 BSE diseased cattle were compared with 28 control animals. Analysis using statistical learning (and linear discriminant analysis) techniques established protein markers of disease with good predictive power (sensitivity 85% and specificity 71%). Further work will be required to test the predictive markers in a wider range of diseases, particularly other neurological conditions.
Differential protein profiling as a potential multi-marker approach for TSE diagnosis
(BMC, 2009-11-27) Barr, Janice B.; Watson, Michael; Head, Mark W.; Ironside, James W.; Harris, Nathan; Hogarth, Caroline; Fraser, Janet R.; Barron, Rona
This "proof of concept" study, examines the use of differential protein expression profiling using surface enhanced laser desorption and ionisationtime of flight mass spectrometry (SELDI-TOF) for the diagnosis of TSE disease. Spectral output from all proteins selectively captured from individual murine brain homogenate samples, are compared as "profiles" in groups of infected and non-infected animals. Differential protein expression between groups is thus highlighted and statistically significant protein "peaks" used to construct a panel of disease specific markers. Studies at both terminal stages of disease and throughout the time course of disease have shown a disease specific protein profile or "disease fingerprint" which could be used to distinguish between groups of TSE infected and uninfected animals at an early time point of disease. Results Our results show many differentially expressed proteins in diseased and control animals, some at early stages of disease. Three proteins identified by SELDI-TOF analysis were verified by immunohistochemistry in brain tissue sections. We demonstrate that by combining the most statistically significant changes in expression, a panel of markers can be constructed that can distinguish between TSE diseased and normal animals. Conclusion Differential protein expression profiling has the potential to be used for the detection of disease in TSE infected animals. Having established that a "training set" of potential markers can be constructed, more work would be required to further test the specificity and sensitivity of the assay in a "testing set". Based on these promising results, further studies are being performed using blood samples from infected sheep to assess the potential use of SELDI-TOF as a pre-mortem blood based diagnostic.
Dissociation between Transmissible Spongiform Encephalopathy (TSE) Infectivity and Proteinase K-Resistant PrPSc Levels in Peripheral Tissue from a Murine Transgenic Model of TSE Disease
(American Society for Microbiology, 2013-04-24) Dobie, Karen; Barron, Rona
Most current diagnostic tests for transmissible spongiform encephalopathies (TSE) rely on the presence of proteinase K (PK)-resistant PrPSc (PrP-res) in postmortem tissues as an indication of TSE disease. However, a number of studies have highlighted a discrepancy between TSE infectivity and PrP-res levels in both natural and experimental cases of TSE disease. Previously, we have shown high TSE infectivity levels in the brain tissue of mice that have a clinical TSE disease with associated vacuolar pathology but little or no detectable PrP-res. Here, the levels of TSE infectivity and PrP-res within a peripheral tissue of this mouse model were investigated. Biochemical analysis showed that low levels of PrP-res were present in the spleen tissue in comparison to the levels observed in the spleen of mice infected with ME7 or 79A. However, upon subpassage of brain and spleen tissue from clinically ill mice with little or no PrP-res detectable, similar short incubation periods to disease were observed, indicating that infectivity levels were similarly high in both tissues. Thus, the discrepancy between PrP-res and TSE infectivity was also present in the peripheral tissues of this disease model. This result indicates that peripheral tissues can contain higher levels of infectivity given the correct combination of host species, PrP genotype, and TSE agent. Therefore, the assumption that the levels of peripheral infectivity are lower than those in the central nervous system is not always correct, and this could have implications for current food safety regulations.
Dissociation of Prion Protein Amyloid Seeding from Transmission of a Spongiform Encephalopathy
(American Society for Microbiology, 2013-11-15) Piccardo, Pedro; King, Declan; Telling, Glenn; Manson, Jean C.; Barron, Rona
Misfolding and aggregation of proteins are common pathogenic mechanisms of a group of diseases called proteinopathies. The formation and spread of proteinaceous lesions within and between individuals were first described in prion diseases and proposed as the basis of their infectious nature. Recently, a similar “prion-like” mechanism of transmission has been proposed in other neurodegenerative diseases such as Alzheimer's disease. We investigated if misfolding and aggregation of corrupted prion protein (PrPTSE) are always associated with horizontal transmission of disease. Knock-in transgenic mice (101LL) expressing mutant PrP (PrP-101L) that are susceptible to disease but do not develop any spontaneous neurological phenotype were inoculated with (i) brain extracts containing PrPTSE from healthy 101LL mice with PrP plaques in the corpus callosum or (ii) brain extracts from mice overexpressing PrP-101L with neurological disease, severe spongiform encephalopathy, and formation of proteinase K-resistant PrPTSE. In all instances, 101LL mice developed PrP plaques in the area of inoculation and vicinity in the absence of clinical disease or spongiform degeneration of the brain. Importantly, 101LL mice did not transmit disease on serial passage, ruling out the presence of subclinical infection. Thus, in both experimental models the formation of PrPTSE is not infectious. These results have implications for the interpretation of tests based on the detection of protein aggregates and suggest that de novo formation of PrPTSE in the host does not always result in a transmissible prion disease. In addition, these results question the validity of assuming that all diseases due to protein misfolding can be transmitted between individuals.
Gene Targeting the PrP Gene
(Horizon Bioscience, 2004) Barron, Rona; Manson, Jean C.
A gene-targeted mouse model of P102L Gerstmann-Sträussler-Scheinker syndrome
(Elsevier, 2003-03) Barron, Rona; Manson, Jean C.
Glycosylation of PrPC Determines Timing of Neuroinvasion and Targeting in the Brain following Transmissible Spongiform Encephalopathy Infection by a Peripheral Route
(American Society for Microbiology, 2010-04-01) Cancellotti, Enrico; Bradford, Barry M.; Tuzi, Nadia L.; Hickey, Raymond D.; Brown, Debbie; Brown, Karen L.; Barron, Rona; Kisielewski, Dorothy; Piccardo, Pedro; Manson, Jean C.
Transmissible spongiform encephalopathy (TSE) infectivity naturally spreads from site of entry in the periphery to the central nervous system where pathological lesions are formed. Several routes and cells within the host have been identified as important for facilitating the infectious process. Expression of the glycoprotein cellular PrP (PrPC) is considered a key factor for replication of infectivity in the central nervous system (CNS) and its transport to the brain, and it has been suggested that the infectious agent propagates from cell to cell via a domino-like effect. However, precisely how this is achieved and what involvement the different glycoforms of PrP have in these processes remain to be determined. To address this issue, we have used our unique models of gene-targeted transgenic mice expressing different glycosylated forms of PrP. Two TSE strains were inoculated intraperitoneally into these mice to assess the contribution of diglycosylated, monoglycosylated, and unglycosylated PrP in spreading of infectivity to the brain. This study demonstrates that glycosylation of host PrP has a profound effect in determining the outcome of disease. Lack of diglycosylated PrP slowed or prevented disease onset after peripheral challenge, suggesting an important role for fully glycosylated PrP in either the replication of the infectious agent in the periphery or its transport to the CNS. Moreover, mice expressing unglycosylated PrP did not develop clinical disease, and mice expressing monoglycosylated PrP showed strikingly different neuropathologic features compared to those expressing diglycosylated PrP. This demonstrates that targeting in the brain following peripheral inoculation is profoundly influenced by the glycosylation status of host PrP.
High Titers of Transmissible Spongiform Encephalopathy Infectivity Associated with Extremely Low Levels of PrPSc in Vivo
(American Society for Biochemistry and Molecular Biology, 2007-10-08) Barron, Rona; Campbell, Susan L.; King, Declan; Bellon, Anne; Chapman, Karen E.; Williamson, R. Anthony; Manson, Jean C.
Diagnosis of transmissible spongiform encephalopathy (TSE) disease in humans and ruminants relies on the detection in post-mortem brain tissue of the protease-resistant form of the host glycoprotein PrP. The presence of this abnormal isoform (PrPSc) in tissues is taken as indicative of the presence of TSE infectivity. Here we demonstrate conclusively that high titers of TSE infectivity can be present in brain tissue of animals that show clinical and vacuolar signs of TSE disease but contain low or undetectable levels of PrPSc. This work questions the correlation between PrPSc level and the titer of infectivity and shows that tissues containing little or no proteinase K-resistant PrP can be infectious and harbor high titers of TSE infectivity. Reliance on protease-resistant PrPSc as a sole measure of infectivity may therefore in some instances significantly underestimate biological properties of diagnostic samples, thereby undermining efforts to contain and eradicate TSEs.
Host PrP Glycosylation: A Major Factor Determining the Outcome of Prion Infection
(Public Library of Science, 2008-04-15) Tuzi, Nadia L.; Cancellotti, Enrico; Baybutt, Herbert; Blackford, Lorraine; Bradford, Barry; Pinston, Chris; Coghill, Anne; Hart, Patricia; Piccardo, Pedro; Barron, Rona; Manson, Jean C.
The expression of the prion protein (PrP) is essential for transmissible spongiform encephalopathy (TSE) or prion diseases to occur, but the underlying mechanism of infection remains unresolved. To address the hypothesis that glycosylation of host PrP is a major factor influencing TSE infection, we have inoculated gene-targeted transgenic mice that have restricted N-linked glycosylation of PrP with three TSE strains. We have uniquely demonstrated that mice expressing only unglycosylated PrP can sustain a TSE infection, despite altered cellular location of the host PrP. Moreover we have shown that brain material from mice infected with TSE that have only unglycosylated PrPSc is capable of transmitting infection to wild-type mice, demonstrating that glycosylation of PrP is not essential for establishing infection within a host or for transmitting TSE infectivity to a new host. We have further dissected the requirement of each glycosylation site and have shown that different TSE strains have dramatically different requirements for each of the glycosylation sites of host PrP, and moreover, we have shown that the host PrP has a major role in determining the glycosylation state of de novo generated PrPSc.
How does host PrP control TSE disease?
(Springer, 2004-05) Manson, Jean; Barron, Rona; Tuzi, Nadia L.; Baybutt, Herbert; Bishop, Matthew; Cancellotti, Enrico; Hart, Patricia; Jamieson, L.; Aitchison, L.; Gall, E.; Bradford, Barry; King, Declan
PrP is central to the TSE disease process and has been hypothesised to be the infectious agent. Polymorphisms in the PrP gene of a number of species are associated with different incubation times of disease following exposure to an infectious agent and mutations in the human PrP gene can apparently lead to spontaneous genetic disease. Strains of TSE agent are proposed to be generated and maintained through differences in glycosylation or conformation of PrP and the barrier to infection between species is thought to be due to the differences in the sequence of PrP between different species. In order to test these hypotheses, we have introduced specific modifications into the endogenous mouse Prnp gene by gene targeting. The mutated PrP gene is in the correct location under the control of the endogenous Prnp regulatory sequences and thus expressed in the same tissues and amounts as the wild type Prnp gene. This strategy therefore allows the effect of specific mutations in the PrP gene to be assessed. We have introduced mutations into the Prnp gene which prevent glycosylation at each or both of the two N-linked glycosylation sites of PrP and are using TSE infection of these mice to investigate the role of PrP glycosylation in strain targeting and strain determination. We have investigated the role of the sequence of the host PrP gene in determining susceptibility by inserting point mutations or replacing the murine PrP gene with that of human or bovine PrP. This has produced a model of TSE disease which contains high levels of infectivity in the absence of PrPSc and we are using this model to determine the nature of the infectious agent. We have thus established that the gene targeting approach can produce models for TSE disease which address fundamental questions associated with these diseases. We aim to use these models to address central issues including the origin of strains, the species barrier and the nature of the infectious agent.
The Human Urinary Proteome Fingerprint Database UPdb
(Hindawi, 2013-10-09) Husi, Holger; Barr, Janice B.; Skipworth, Richard J. E.; Stephens, Nathan A.; Greig, Carolyn A.; Wackerhage, Henning; Barron, Rona; Fearon, Kenneth C. H.; Ross, James A.
The use of human urine as a diagnostic tool has many advantages, such as ease of sample acquisition and noninvasiveness. However, the discovery of novel biomarkers, as well as biomarker patterns, in urine is hindered mainly by a lack of comparable datasets. To fill this gap, we assembled a new urinary fingerprint database. Here, we report the establishment of a human urinary proteomic fingerprint database using urine from 200 individuals analysed by SELDI-TOF (surface enhanced laser desorption ionisation-time of flight) mass spectrometry (MS) on several chip surfaces (SEND, HP50, NP20, Q10, CM10, and IMAC30). The database currently lists 2490 unique peaks/ion species from 1172 nonredundant SELDI analyses in the mass range of 1500 to 150000. All unprocessed mass spectrometric scans are available as “.xml” data files. Additionally, 1384 peaks were included from external studies using CE (capillary electrophoresis)-MS, MALDI (matrix assisted laser desorption/ionisation), and CE-MALDI hybrids. We propose to use this platform as a global resource to share and exchange primary data derived from MS analyses in urinary research.
Identification and characterization of a putative serine protease expressed in vivo by Mycobacterium avium subsp. paratuberculosis
(Microbiology Society, 1994-08-01) Barron, Rona; Stevenson, Karen; Inglis, Neil F.; Klausen, Joan; Sharp, J. Michael
A putative serine protease expressed in vivo by Mycobacterium avium subsp. paratuberculosis was isolated from a lambda gt11 genomic expression library by screening with serum from a naturally infected sheep. The gene was contained in two overlapping clones, which were shown by antibody elution to encode a protein of 34 kDa in M. a. paratuberculosis. The clones were sequenced and database searches detected a motif identical to the active serine site in trypsin, and 30% homology to the putative serine proteases (HtrA proteins) of Escherichia coli, Salmonella typhimurium, Brucella abortus and Rochalimaea henselae.
Increased Susceptibility of Human-PrP Transgenic Mice to Bovine Spongiform Encephalopathy Infection following Passage in Sheep
(American Society for Microbiology, 2011-01-12) Plinston, Chris; Hart, Patricia; Chong, Angela; Piccardo, Pedro; Hunter, Nora; Foster, James; Manson, Jean C.; Barron, Rona
The risk of the transmission of ruminant transmissible spongiform encephalopathy (TSE) to humans was thought to be low due to the lack of association between sheep scrapie and the incidence of human TSE. However, a single TSE agent strain has been shown to cause both bovine spongiform encephalopathy (BSE) and human vCJD, indicating that some ruminant TSEs are transmissible to humans. While the transmission of cattle BSE to humans in transgenic mouse models has been inefficient, indicating the presence of a significant transmission barrier between cattle and humans, BSE has been transmitted to a number of other species. Here, we aimed to further investigate the human transmission barrier following the passage of BSE in a sheep. Following inoculation with cattle BSE, gene-targeted transgenic mice expressing human PrP showed no clinical or pathological signs of TSE disease. However, following inoculation with an isolate of BSE that had been passaged through a sheep, TSE-associated vacuolation and proteinase K-resistant PrP deposition were observed in mice homozygous for the codon 129-methionine PRNP gene. This observation may be due to higher titers of the BSE agent in sheep or an increased susceptibility of humans to BSE prions following passage through a sheep. However, these data confirm that, contrary to previous predictions, it is possible that a sheep prion is transmissible to humans and that BSE from other species is a public health risk.
Increased susceptibility of transgenic mice expressing human PrP to experimental sheep bovine spongiform encephalopathy is not due to increased agent titre in sheep brain tissue
(Microbiology Society, 2014-08-01) Plinston, Chris; Hart, Patricia; Hunter, Nora; Manson, Jean C.; Barron, Rona
Bovine spongiform encephalopathy (BSE) in cattle and variant Creutzfeldt–Jakob disease in humans have previously been shown to be caused by the same strain of transmissible spongiform encephalopathy agent. It is hypothesized that the agent spread to humans following consumption of food products prepared from infected cattle. Despite evidence supporting zoonotic transmission, mouse models expressing human prion protein (HuTg) have consistently shown poor transmission rates when inoculated with cattle BSE. Higher rates of transmission have however been observed when these mice are exposed to BSE that has been experimentally transmitted through sheep or goats, indicating that humans may potentially be more susceptible to BSE from small ruminants. Here we demonstrate that increased transmissibility of small ruminant BSE to HuTg mice was not due to replication of higher levels of infectivity in sheep brain tissue, and is instead due to other specific changes in the infectious agent.