•We examined methylene blue treatment in the rTg4510 mouse model of tauopathy.•Methylene blue treatment does not clear existing neurofibrillary tangles in vivo.•Neither PHF1 positive nor Gallyas ...positive tangles were cleared by the treatment.
Alzheimer's disease is characterized pathologically by aggregation of amyloid beta into senile plaques and aggregation of pathologically modified tau into neurofibrillary tangles. While changes in amyloid processing are strongly implicated in disease initiation, the recent failure of amyloid-based therapies has highlighted the importance of tau as a therapeutic target. “Tangle busting” compounds including methylene blue and analogous molecules are currently being evaluated as therapeutics in Alzheimer's disease. Previous studies indicated that methylene blue can reverse tau aggregation in vitro after 10min, and subsequent studies suggested that high levels of drug reduce tau protein levels (assessed biochemically) in vivo. Here, we tested whether methylene blue could remove established neurofibrillary tangles in the rTg4510 model of tauopathy, which develops robust tangle pathology. We find that 6 weeks of methylene blue dosing in the water from 16 months to 17.5 months of age decreases soluble tau but does not remove sarkosyl insoluble tau, or histologically defined PHF1 or Gallyas positive tangle pathology. These data indicate that methylene blue treatment will likely not rapidly reverse existing tangle pathology.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Misfolding of tau proteins into prions and their propagation along neural circuits are thought to result in neurodegeneration causing Alzheimer’s disease, progressive supranuclear palsy, chronic ...traumatic encephalopathy, and other tauopathies. Little is known about the molecular processes mediating tau prion replication and spreading in different brain regions. Using transgenic (Tg) mice with a neuronal promoter driving expression of human mutant (P301S) tau, we found that tau prion formation and histopathologic deposition is largely restricted to the hindbrain. Unexpectedly, tau mRNA and protein levels did not differ between the forebrain and hindbrain, suggesting that other factors modulating the conversion of tau into a prion exist and are region specific. Using a cell-based prion propagation assay, we discovered that tau prion replication is suppressed by forebrain-derived inhibitors, one of which is sortilin, a lysosomal sorting receptor. We also show that sortilin expression is higher in the forebrain than the hindbrain across the life span of the Tg mice, suggesting that sortilin, at least in part, inhibits forebrain tau prion replication in vivo. Our findings provide evidence for selective vulnerability in mice resulting in highly regulated levels of tau prion propagation, thus affording a model for identification of additional molecules that could mitigate the levels of tau prions in human tauopathies.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Although the discovery of the prion protein (PrP) resulted from its co-purification with scrapie infectivity in Syrian hamsters, work with genetically defined and genetically modified mice proved ...crucial for understanding the fundamental processes involved not only in prion diseases caused by PrP misfolding, aggregation, and spread but also in other, much more common, neurodegenerative brain diseases. In this review, we focus on methodological and conceptual approaches used to study scrapie and related PrP misfolding diseases in mice and how these approaches have advanced our understanding of related disorders including Alzheimer's and Parkinson's disease.
Glial cytoplasmic inclusions (GCIs) containing aggregated and hyperphosphorylated alpha-synuclein are the signature neuropathological hallmark of multiple system atrophy (MSA). Native alpha-synuclein ...can adopt a prion conformation that self-propagates and spreads throughout the brain ultimately resulting in neurodegeneration. A growing body of evidence argues that, in addition to oligodendrocytes, astrocytes contain alpha-synuclein inclusions in MSA and other alpha-synucleinopathies at advanced stages of disease. To study the role of astrocytes in MSA, we added MSA brain homogenate to primary cultures of astrocytes from transgenic (Tg) mouse lines expressing human alpha-synuclein. Astrocytes from four Tg lines, expressing either wild-type or mutant (A53T or A30P) human alpha-synuclein, propagated and accumulated alpha-synuclein prions. Furthermore, we found that MSA-infected astrocytes formed two morphologically distinct alpha-synuclein inclusions: filamentous and granular. Both types of cytoplasmic inclusions shared several features characteristic of alpha-synuclein inclusions in synucleinopathies: hyperphosphorylation preceded by aggregation, ubiquitination, thioflavin S-positivity, and co-localization with p62. Our findings demonstrate that human alpha-synuclein forms distinct inclusion morphologies and propagates within cultured Tg astrocytes exposed to MSA prions, indicating that alpha-synuclein expression determines the tropism of inclusion formation in certain cells. Thus, our work may prove useful in elucidating the role of astrocytes in the pathogenic mechanisms that feature in neurodegeneration caused by MSA prions. Keywords: MSA, Prion, Astrocytes, alpha-Synuclein, Proteinopathies
Overproduction of the amyloid beta (Abeta) peptide is a key factor in the pathogenesis of Alzheimer's disease (AD), but the mechanisms of its pathogenic effects have not been defined. Patients with ...AD have cerebrovascular alterations attributable to the deleterious effects of Abeta on cerebral blood vessels. We report here that NADPH oxidase, the major source of free radicals in blood vessels, is responsible for the cerebrovascular dysregulation induced by Abeta. Thus, the free-radical production and the associated alterations in vasoregulation induced by Abeta are abrogated by the NADPH oxidase peptide inhibitor gp91ds-tat and are not observed in mice lacking the catalytic subunit of NADPH oxidase (gp91phox). Furthermore, oxidative stress and cerebrovascular dysfunction do not occur in transgenic mice overexpressing the amyloid precursor protein but lacking gp91phox. The mechanisms by which NADPH oxidase-derived radicals mediate the cerebrovascular dysfunction involve reduced bioavailability of nitric oxide. Thus, a gp91phox-containing NADPH oxidase is the critical link between Abeta and cerebrovascular dysfunction, which may underlie the alteration in cerebral blood flow regulation observed in AD patients.
Mesopelagic zone ecology and biogeochemistry – a synthesis Robinson, Carol; Steinberg, Deborah K.; Anderson, Thomas R. ...
Deep-sea research. Part 2. Topical studies in oceanography,
08/2010, Volume:
57, Issue:
16
Journal Article, Conference Proceeding
Peer reviewed
The mesopelagic zone is the oceanic region through which carbon and other elements must pass in order to reach deeper waters or the sea floor. However, the food web interactions that occur in the ...mesopelagic zone are difficult to measure and so, despite their crucial importance to global elemental cycles, are not very well known. Recent developments in technology and new approaches have advanced the study of the variability in and controls upon the distribution and diversity of organisms in the mesopelagic zone, including the roles of respiration, recycling, and repackaging of particulate and dissolved organic material. However, there are remarkably few syntheses of the ecology and biogeochemistry of the microbes and metazoa that permanently reside or habitually visit this ‘twilight zone’. Without this synthesis, it is difficult to assess the impact of ongoing changes in ocean hydrography and chemistry, due to increasing atmospheric carbon dioxide levels, on the biological carbon pump. This paper reviews what is known about the distribution of microbes and metazoa in the mesopelagic zone in relation to their activity and impact on global biogeochemical cycles. Thus, gaps in our knowledge are identified and suggestions made for priority research programmes that will improve our ability to predict the effects of climate change on carbon sequestration.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
ENU mutagenesis is a powerful method for generating novel lines of mice that are informative with respect to both fundamental biological processes and human disease. Rapid developments in genomic ...technology have made the task of identifying causal mutations by positional cloning remarkably efficient. One limitation of this approach remains the mutation frequency achievable using standard treatment protocols, which currently generate approximately 1-2 sequence changes per megabase when optimized. In this study we used two strategies to attempt to increase the number of mutations induced by ENU treatment. One approach employed mice carrying a mutation in the DNA repair enzyme Msh6. The second strategy involved injection of ENU to successive generations of mice. To evaluate the number of ENU-induced mutations, single mice or pooled samples were analyzed using whole exome sequencing. The results showed that there is considerable variability in the induced mutation frequency using these approaches, but an overall increase in ENU-induced variants from one generation to another was observed. The analysis of the mice deficient for Msh6 also showed an increase in the ENU-induced variants compared to the wild-type ENU-treated mice. However, in both cases the increase in ENU-induced mutation frequency was modest.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
A systems approach to prion disease Hood, Leroy E; Price, Nathan D; Pitstick, Rose ...
Molecular systems biology,
2009, Volume:
5, Issue:
1
Journal Article
Peer reviewed
Open access
Prions cause transmissible neurodegenerative diseases and replicate by conformational conversion of normal benign forms of prion protein (PrPC) to disease‐causing PrPSc isoforms. A systems approach ...to disease postulates that disease arises from perturbation of biological networks in the relevant organ. We tracked global gene expression in the brains of eight distinct mouse strain–prion strain combinations throughout the progression of the disease to capture the effects of prion strain, host genetics, and PrP concentration on disease incubation time. Subtractive analyses exploiting various aspects of prion biology and infection identified a core of 333 differentially expressed genes (DEGs) that appeared central to prion disease. DEGs were mapped into functional pathways and networks reflecting defined neuropathological events and PrPSc replication and accumulation, enabling the identification of novel modules and modules that may be involved in genetic effects on incubation time and in prion strain specificity. Our systems analysis provides a comprehensive basis for developing models for prion replication and disease, and suggests some possible therapeutic approaches.
Synopsis
A systems approach to disease postulates that disease arises from the pathological perturbation (genetic and/or environmental) of one or more biological networks in the relevant organ and hence to understand a disease one must study the dynamical changes in relevant biological networks during disease progression. We applied the systems approach analyzing brain transcriptomes to the experimentally tractable neurodegenerative diseases caused by prion infection of mice. Prions are unique among transmissible, disease‐causing agents in that they replicate by conformational conversion of normal benign forms of prion protein (PrPC) to disease‐specific PrPSc isoforms. Neuropathological features common to all prion diseases in mammals, which include bovine spongiform encephalopathy (BSE) in cows, Creutzfeldt–Jakob disease (CJD) in humans, and scrapie in sheep, can be conveniently subdivided into four well‐defined pathological processes: prion replication and PrPSc accumulation (Prusiner, 2003), synaptic degeneration (Ishikura et al, 2005), microglia and astrocyte activation (Rezaie and Lantos, 2001; Perry et al, 2002), and neuronal cell death (Liberski et al, 2004). Data on pathological changes in prion disease have been derived in multiple laboratories that have viewed prion‐induced neurodegeneration from different perspectives and with different preconceptions. Our comprehensive and independent systems analysis of the brain transcriptomes in normal and prion‐infected mice provides gene expression correlates with pathological information and will aid in organizing the current abundance of data fragments into a coherent pathogenic model of prion disease.
We tracked global gene expression in the brains of eight distinct mouse strain–prion strain combinations at 8–10 time points throughout the incubation periods (60–350 days) to capture the effects of prion strain, host genetics, and PrP concentration on disease incubation time (Figure 1). Approximately 7400 genes were differentially expressed genes (DEGs) in one or more of the combinations. Subtractive analyses using three inbred mouse strains and two prion strains reduced the data dimensionality from 7400 to a core of 333 DEGs that reflected effects of prion strain and Prnp genotype that appeared central to prion disease. Of these, 178 had not previously been reported to change in prion‐infected mice. Gene expression results were combined with temporal patterns of PrPSc accumulation, pathology, gene ontology, protein interactions, and cell‐specific gene expression data to generate hypothetical dynamic protein networks that could be associated with known pathological events in disease progression; 231 DEGs were mapped into these networks. Figure 4 is a snapshot of one of these networks (PrPSc accumulation) in a single mouse strain–prion strain combination at 14 weeks after inoculation, before any clinical signs are apparent. This figure includes a histoblot to track regional deposition of proteinase K‐resistant PrPSc in the brain; histoblots were collected at each time point for each prion strain–mouse strain combination. The previously unidentified DEGs and those that could not be readily assigned to networks likely encode previously unidentified aspects of prion disease and subsets of these may reveal involvement of modules reflecting androgenic steroid, iron, or arachidonate/prostaglandin metabolism. All data and tools used in these studies are available online in a prion disease database (http://prion.systemsbiology.net)
Grouping mice in the five core prion strain–mouse strain combinations according to differences in incubation time revealed 55 DEGs, the expression of which was significantly enriched only in groups with short incubation times (B6‐RML, B6.I‐301V, and FVB‐RML). Similarly, grouping according to prion strain (RML or 301V) identified 39 DEGs enriched in RML prion‐inoculated mice. Interestingly, the emphasis on pathways such as cholesterol metabolism or glycosaminoglycan biosynthesis as central to prion disease may reflect the widespread use of RML and related prion isolates in short incubation time mice and in cell culture. The five core mouse strain–prion strain combinations emphasize incubation time differences reflecting interactions of PrPSc with PrPC encoded by alternative alleles of Prnp. PrPC concentration can also affect incubation time, and differential gene expression was explored in FVB.129‐Prnptm1Zrch/wt (0/+) mice that express half the normal amount of PrP and have a very long RML incubation time and in FVB‐Tg(PrP‐A)4053 (Tg4053) mice that overexpress PrP and have a very short incubation time. Among the 333 shared DEGs gleaned from five prion–wild‐type mouse strain combinations, 311 DEGs also were changed in Prnp (0/+) mice (summarized in Figure 1). In contrast, Tg4053 mice PrP showed significant changes in only 125 of the 333 DEGs in the shared set. Prominent shared DEGs in most of the key shared modules exhibited patterns in Tg4053 mice that were similar to the core groups, though generally with differentials of smaller magnitude and closer in time to clinical illness than all other combinations of prions and mice. Perplexingly, prion‐infected Tg4053 also had a unique set of highly significant DEGs that were not seen in any other mouse–prion combination.
We have demonstrated here the power of comprehensive, global systems approaches to diseases as complex as prion infection, even when the data sets are restricted to gene expression profiles, and involve whole brain. The efficacy of using several strain combinations, prion and genetic backgrounds as biological filters to identify the network signals that are important for various disease‐related processes is a striking lesson from our study. The new modules that have been connected to the disease, the strong alignment of the specific pathogenic processes with network changes, and the range of novel and sharpened hypotheses illustrate the power of this approach. We have confidence that with the addition of other data types, the attribution of network processes to brain regions, and the specific testing of hypotheses suggested here, that the systems medicine of prion disease (and other neurodegenerative diseases) will advance rapidly. This study also provided new insights into the power of systems approaches to formulate new strategies for blood diagnosis and treatment.
A systems approach was applied to neurodegenerative disease caused by prions, which are transmissible agents that replicate by conformational conversion of normal, benign forms of prion protein (PrPC) to disease‐specific PrPSc isoforms.
Analysis of brain transcriptomes and regional PrPSc accumulation in eight distinct mouse strain‐prion strain combinations at 8 to 10 time points across their incubation periods captured the effects of prion strain, host genetics, and PrP concentration on pathogenesis and disease incubation time.
We constructed hypothetical networks corresponding to four signal features of prion disease‐prion accumulation and replication, microglial and astrocytic activation, degeneration of axons and presynaptic boutons, and neural cell death.
A core of 333 genes showing shared dynamic differential expression appeared to encode the heart of prion disease; two‐thirds of these genes encoded known pathological events of prion disease and one third encoded novel, previously unknown, aspects of this disease.
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FZAB, GIS, IJS, IZUM, KILJ, NLZOH, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK
Pituitary Prolactin (PRL) and Growth Hormone (GH) are separately controlled and sub-serve different purposes. Surprisingly, we demonstrate that extra-pituitary expression in the adult mammalian ...central nervous system (CNS) is coordinated at mRNA and protein levels. However this was not a uniform effect within populations, such that wide inter-individual variation was superimposed on coordinate PRL/GH expression. Up to 44% of individuals in healthy cohorts of mice and rats showed protein levels above the norm and coordinated expression of PRL and GH transcripts above baseline occurred in the amygdala, frontal lobe and hippocampus of 10% of human subjects. High levels of PRL and GH present in post mortem tissue were often presaged by altered responses in fear conditioning and stress induced hyperthermia behavioral tests. Our data define a common phenotype polymorphism in healthy mammalian brains, and, given the pleiotropic effects known for circulating PRL and GH, further consequences of coordinated CNS over-expression may await discovery.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
A growing body of evidence suggests a relationship between oxidative stress and β‐amyloid (Aβ) peptide accumulation, a hallmark in the pathogenesis of Alzheimer's disease (AD). However, a direct ...causal relationship between oxidative stress and Aβ pathology has not been established in vivo. Therefore, we crossed mice with a knockout of one allele of manganese superoxide dismutase (MnSOD), a critical antioxidant enzyme, with Tg19959 mice, which overexpress a doubly mutated human β‐amyloid precursor protein (APP). Partial deficiency of MnSOD, which is well established to cause elevated oxidative stress, significantly increased brain Aβ levels and Aβ plaque burden in Tg19959 mice. These results indicate that oxidative stress can promote the pathogenesis of AD and further support the feasibility of antioxidant approaches for AD therapy.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
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