scholarly journals Dextran sulphate-induced tau assemblies cause endogenous tau aggregation and propagation in wild-type mice

2020 ◽  
Vol 2 (2) ◽  
Author(s):  
Masami Masuda-Suzukake ◽  
Genjiro Suzuki ◽  
Masato Hosokawa ◽  
Takashi Nonaka ◽  
Michel Goedert ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Tau Protein ◽  
Guanidine Hydrochloride ◽  
Intracerebral Injection ◽  
Dextran Sulphate ◽  
Wild Type ◽  
Synthetic Filament ◽  
Biochemical Analyses ◽  
Tau Aggregation

Abstract Accumulation of assembled tau protein in the central nervous system is characteristic of Alzheimer’s disease and several other neurodegenerative diseases, called tauopathies. Recent studies have revealed that propagation of assembled tau is key to understanding the pathological mechanisms of these diseases. Mouse models of tau propagation are established by injecting human-derived tau seeds intracerebrally; nevertheless, these have a limitation in terms of regulation of availability. To date, no study has shown that synthetic assembled tau induce tau propagation in non-transgenic mice. Here we confirm that dextran sulphate, a sulphated glycosaminoglycan, induces the assembly of recombinant tau protein into filaments in vitro. As compared to tau filaments induced by heparin, those induced by dextran sulphate showed higher thioflavin T fluorescence and lower resistance to guanidine hydrochloride, which suggests that the two types of filaments have distinct conformational features. Unlike other synthetic filament seeds, intracerebral injection of dextran sulphate-induced assemblies of recombinant tau caused aggregation of endogenous murine tau in wild-type mice. AT8-positive tau was present at the injection site 1 month after injection, from where it spread to anatomically connected regions. Induced tau assemblies were also stained by anti-tau antibodies AT100, AT180, 12E8, PHF1, anti-pS396 and anti-pS422. They were thioflavin- and Gallyas-Braak silver-positive, indicative of amyloid. In biochemical analyses, accumulated sarkosyl-insoluble and hyperphosphorylated tau was observed in the injected mice. In conclusion, we revealed that intracerebral injection of synthetic full-length wild-type tau seeds prepared in the presence of dextran sulphate caused tau propagation in non-transgenic mice. These findings establish that propagation of tau assemblies does not require tau to be either mutant and/or overexpressed.

scholarly journals Abrogation of Neuraminidase Reduces Biofilm Formation, Capsule Biosynthesis, and Virulence of Porphyromonas gingivalis

2011 ◽  
Vol 80 (1) ◽  
pp. 3-13 ◽  
Author(s):  
Chen Li ◽  
Kurniyati ◽  
Bo Hu ◽  
Jiang Bian ◽  
Jianlan Sun ◽  
...  
Keyword(s):  
Porphyromonas Gingivalis ◽  
Biofilm Formation ◽  
Adult Population ◽  
Transcriptional Analysis ◽  
Wild Type ◽  
Content Type ◽  
Multiple Organs ◽  
Biochemical Analyses

ABSTRACTThe oral bacteriumPorphyromonas gingivalisis a key etiological agent of human periodontitis, a prevalent chronic disease that affects up to 80% of the adult population worldwide.P. gingivalisexhibits neuraminidase activity. However, the enzyme responsible for this activity, its biochemical features, and its role in the physiology and virulence ofP. gingivalisremain elusive. In this report, we found thatP. gingivalisencodes a neuraminidase, PG0352 (SiaPg). Transcriptional analysis showed thatPG0352is monocistronic and is regulated by a sigma70-like promoter. Biochemical analyses demonstrated that SiaPgis an exo-α-neuraminidase that cleaves glycosidic-linked sialic acids. Cryoelectron microscopy and tomography analyses revealed that thePG0352deletion mutant (ΔPG352) failed to produce an intact capsule layer. Compared to the wild type,in vitrostudies showed that ΔPG352 formed less biofilm and was less resistant to killing by the host complement.In vivostudies showed that while the wild type caused a spreading type of infection that affected multiple organs and all infected mice were killed, ΔPG352 only caused localized infection and all animals survived. Taken together, these results demonstrate that SiaPgis an important virulence factor that contributes to the biofilm formation, capsule biosynthesis, and pathogenicity ofP. gingivalis, and it can potentially serve as a new target for developing therapeutic agents againstP. gingivalisinfection.

scholarly journals Effects of resveratrol and morin on insoluble tau in tau transgenic mice

2018 ◽  
Vol 9 (1) ◽  
pp. 54-60 ◽  
Author(s):  
Kwun Chung Yu ◽  
Ping Kwan ◽  
Stanley K.K. Cheung ◽  
Amy Ho ◽  
Larry Baum
Keyword(s):  
Transgenic Mice ◽  
Motor Function ◽  
Tau Protein ◽  
Late Stage ◽  
Animal Studies ◽  
Paired Helical Filaments ◽  
Hyperphosphorylated Tau ◽  
Total Tau ◽  
Tau Aggregation ◽  
Do So

Abstract Tauopathies are neurodegenerative diseases, including Alzheimer’s disease (AD) and frontotemporal dementia (FTD), in which tau protein aggregates within neurons. An effective treatment is lacking and is urgently needed. We evaluated two structurally similar natural compounds, morin and resveratrol, for treating tauopathy in JNPL3 P301L mutant human tau overexpressing mice. Rotarod tests were performed to determine effects on motor function. After treatment from age 11 to 14 months, brains of 26 mice were collected to quantify aggregated hyperphosphorylated tau by Thioflavin T and immunohistochemistry (IHC) and to quantify total tau (HT7 antibody) and hyperphosphorylated tau (AT8 antibody) in homogenates and a fraction enriched for paired helical filaments. Resveratrol reduced the level of total hyperphosphorylated tau in IHC sections (p=0.036), and morin exhibited a tendency to do so (p=0.29), while the two drugs tended to increase the proportion of solubilizable tau that was hyperphosphorylated, as detected in blots. Neither resveratrol nor morin affected motor function. One explanation of these results is that the drugs might interrupt a late stage in tau aggregation, after small aggregates have formed but before further aggregation has occurred. Further animal studies would be informative to explore the possible efficacy of morin or resveratrol for treating tauopathies.

scholarly journals Mice that overexpress Cu/Zn superoxide dismutase are resistant to allergen-induced changes in airway control

2000 ◽  
Vol 279 (2) ◽  
pp. L350-L359 ◽  
Author(s):  
Gary L. Larsen ◽  
Carl W. White ◽  
Katsuyuki Takeda ◽  
Joan E. Loader ◽  
Dee Dee H. Nguyen ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Smooth Muscle ◽  
Transgenic Mice ◽  
Neural Control ◽  
Electrical Field Stimulation ◽  
Tracheal Smooth Muscle ◽  
Wild Type ◽  
Airway Control ◽  
Induced Changes

Within the respiratory epithelium of asthmatic patients, copper/zinc-containing superoxide dismutase (Cu/Zn SOD) is decreased. To address the hypothesis that lung Cu/Zn SOD protects against allergen-induced injury, wild-type and transgenic mice that overexpress human Cu/Zn SOD were either passively sensitized to ovalbumin (OVA) or actively sensitized by repeated airway exposure to OVA. Controls included nonsensitized wild-type and transgenic mice given intravenous saline or airway exposure to saline. After aerosol challenge to saline or OVA, segments of tracheal smooth muscle were obtained for in vitro analysis of neural control. In response to electrical field stimulation, wild-type sensitized mice challenged with OVA had significant increases in cholinergic reactivity. Conversely, sensitized transgenic mice challenged with OVA were resistant to changes in neural control. Stimulation of tracheal smooth muscle to elicit acetylcholine release showed that passively sensitized wild-type but not transgenic mice released more acetylcholine after OVA challenge. Function of the M2 muscarinic autoreceptor was preserved in transgenic mice. These results demonstrate that murine airways with elevated Cu/Zn SOD were resistant to allergen-induced changes in neural control.

scholarly journals Prion replication without host adaptation during interspecies transmissions

2017 ◽  
Vol 114 (5) ◽  
pp. 1141-1146 ◽  
Author(s):  
Jifeng Bian ◽  
Vadim Khaychuk ◽  
Rachel C. Angers ◽  
Natalia Fernández-Borges ◽  
Enric Vidal ◽  
...  
Keyword(s):  
Disease Transmission ◽  
Protein Misfolding ◽  
Guanidine Hydrochloride ◽  
Host Adaptation ◽  
Wild Type ◽  
Epidemiological Features ◽  
Significant Barrier ◽  
Prion Transmission ◽  
Selection Of

Adaptation of prions to new species is thought to reflect the capacity of the host-encoded cellular form of the prion protein (PrPC) to selectively propagate optimized prion conformations from larger ensembles generated in the species of origin. Here we describe an alternate replicative process, termed nonadaptive prion amplification (NAPA), in which dominant conformers bypass this requirement during particular interspecies transmissions. To model susceptibility of horses to prions, we produced transgenic (Tg) mice expressing cognate PrPC. Although disease transmission to only a subset of infected TgEq indicated a significant barrier to EqPrPCconversion, the resulting horse prions unexpectedly failed to cause disease upon further passage to TgEq. TgD expressing deer PrPCwas similarly refractory to deer prions from diseased TgD infected with mink prions. In both cases, the resulting prions transmitted to mice expressing PrPCfrom the species of prion origin, demonstrating that transmission barrier eradication of the originating prions was ephemeral and adaptation superficial in TgEq and TgD. Horse prions produced in vitro by protein misfolding cyclic amplification of mouse prions using horse PrPCalso failed to infect TgEq but retained tropism for wild-type mice. Concordant patterns of neuropathology and prion deposition in susceptible mice infected with NAPA prions and the corresponding prion of origin confirmed preservation of strain properties. The comparable responses of both prion types to guanidine hydrochloride denaturation indicated this occurs because NAPA precludes selection of novel prion conformations. Our findings provide insights into mechanisms regulating interspecies prion transmission and a framework to reconcile puzzling epidemiological features of certain prion disorders.

Cuttings of the non-rooting rac tobacco mutant overaccumulate phenolic compounds

2002 ◽  
Vol 29 (1) ◽  
pp. 63 ◽  
Author(s):  
Odile Faivre-Rampant ◽  
Jean-Paul Charpentier ◽  
Claire Kevers ◽  
Jacques Dommes ◽  
Harry Van Onckelen ◽  
...  
Keyword(s):  
Chlorogenic Acid ◽  
Adventitious Roots ◽  
Phenylalanine Ammonia Lyase ◽  
Wild Type ◽  
Growth Limitation ◽  
Phenolic Contents ◽  
Pal Activity ◽  
Culture Cycle ◽  
Biochemical Analyses

The auxin and phenolic contents, as well as phenylalanine ammonia-lyase (PAL) activity, were determined in in vitro cultured shoots of the recalcitrant-to-root rac mutant of tobacco, and compared with wild-type shoots. The mutant and wild-type shoots showed similar auxin changes during the culture cycle, but with higher contents for the mutant. A transient peak of auxin (corresponding to the achievement of the rooting inductive phase) occurred at day 14 in both types of shoots, but earlier in the basal parts of the wild-type stems. The rac shoots contained more phenolics, corresponding with an increased PAL activity. The most abundant phenolic compound found in the two types of tobacco was chlorogenic acid, which was more abundant in the rac shoots. Rutin was also detected at a higher concentration in the mutant shoots. Basal parts of wild-type shoots treated with 10–3 M chlorogenic acid reacted by accumulating auxins and, unlike untreated controls, did not form adventitious roots. The relationships between these biochemical analyses in relation to the growth limitation of the rac mutant, and the inhibition of its root development, are discussed.

Hyperactivable NFAT1 Ameliorates Autoimmune Encephalitis In Vivo.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 711-711
Author(s):  
Srimoyee Ghosh ◽  
Sergei B Koralov ◽  
Irena Stevanovic ◽  
Mark S Sundrud ◽  
Yoshiteru Sasaki ◽  
...  
Keyword(s):  
T Cells ◽  
Transgenic Mice ◽  
Cd4 T Cells ◽  
Th17 Cells ◽  
Autoimmune Encephalitis ◽  
Cell Types ◽  
Wild Type ◽  
Ifn Γ

Abstract Abstract 711 Naïve CD4 T cells differentiate into diverse effector and regulatory subsets to coordinate the adaptive immune response. TH1 and TH2 effector subsets produce IFN-γ and IL-4, respectively, whereas proinflammatory TH17 cells are key regulators of autoimmune inflammation, characteristically produce IL-17 and IL-22 and differentiate in the presence of inflammatory cytokines like IL-6 and IL-21 together with TGF-β. Naive T cells can also differentiate into tissue-protective induced T regulatory (iTreg) cells. NFAT proteins are highly phosphorylated and reside in the cytoplasm of resting cells. Upon dephosphorylation by the Ca2+/calmodulin-dependent serine phosphatase calcineurin, NFAT proteins translocate to the nucleus, where they orchestrate developmental and activation programs in diverse cell types. In this study, we investigated the role of the Ca/NFAT signaling pathway in regulating T cell differentiation and the development of autoimmune diseases. We generated transgenic mice conditionally expressing a hyperactivable version of NFAT1 (AV-NFAT1) from the ROSA26 locus. To restrict AV-NFAT1 expression to the T cell compartment, ROSA26-AV-NFAT1 transgenic mice were bred to CD4-Cre transgenic mice. Naïve CD4 T cells freshly isolated from AV mice produced significantly less IL-2 but increased amounts of the inhibitory cytokine IL-10. To investigate the role of NFAT1 in the generation of TH1, TH2, Tregand TH17 cells, the respective cell types were generated from CD4 T cells of AV mice by in vitro differentiation. T cells from AV-NFAT1 mice exhibited a dysregulation of cytokine expression, producing more IFN-γ and less IL-4. While the numbers of CD4+CD25+ “natural” Treg cells in peripheral lymphoid organs and their in vitro suppressive functions were slightly decreased in AV mice, iTreg generation from CD4+CD25- T cells of AV mice as compared to wild type cells was markedly enhanced. Moreover, TH17 cells generated in vitro from CD4 T cells of AV mice in the presence of IL-6, IL-21 and TGF-β exhibited dramatically increased expression of both IL-10 and IL-17 as compared to wild type controls. To investigate putative NFAT binding sites in the IL-10 and IL-17 gene loci, we performed chromatin immunoprecipitation experiments. We show that NFAT1 can bind at the IL-17 locus at 3 out of 9 CNS regions which are accessible specifically during TH17 but not during TH1 and TH2 differentiation. Furthermore, we provide evidence that NFAT1 binds one CNS region in the IL10-locus in TH17 cells. To verify our observations in vivo, we induced experimental autoimmune encephalitis (EAE) in AV mice and wild type controls with the immunodominant myelin antigen MOG33-55 emulsified in complete Freund‘s adjuvant. While wild type animals showed a normal course of disease with development of tail and hind limb paralysis after approximately 10 days, AV mice showed a markedly weaker disease phenotype with less severe degrees of paralysis and accelerated kinetics of remission. Moreover at the peak of the response, there were fewer CD4+CD25- but more CD4+CD25+ T cells in the CNS of AV animals compared to wild type controls. Surprisingly, these cells produced significantly more IL-2, IL-17 and IFN-γ upon restimulation, even though they displayed decreased disease. In summary, our data provide strong evidence that NFAT1 contributes to the regulation of IL-10 and IL-17 expression in TH17 cells and show that increasing NFAT1 activity can ameliorate autoimmune encephalitis. This could occur in part through upregulation of IL-10 expression as observed in vitro, but is also likely to reflect increased infiltration of regulatory T cells into the CNS as well as increased conversion of conventional T cells into Foxp3+ regulatory T cells within the CNS. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Lysine methylation is an endogenous post-translational modification of tau protein in human brain and a modulator of aggregation propensity

2014 ◽  
Vol 462 (1) ◽  
pp. 77-88 ◽  
Author(s):  
Kristen E. Funk ◽  
Stefani N. Thomas ◽  
Kelsey N. Schafer ◽  
Grace L. Cooper ◽  
Zhongping Liao ◽  
...  
Keyword(s):  
Human Brain ◽  
Tau Protein ◽  
Protein Function ◽  
Lysine Methylation ◽  
Post Translational Modification ◽  
Post Translational Modifications ◽  
Aggregation Propensity ◽  
Normal Human ◽  
Tau Aggregation

Diverse post-translational modifications regulate tau protein function and misfolding. In the present study we identified lysine methylation as a tau post-translational modification in normal human brain, and found it depressed tau aggregation propensity when modelled in vitro.

scholarly journals From Aggregation to Inhibition: N-Amination Converts Amyloidogenic Tau Peptides into Soluble Antagonists of Cellular Seeding

2021 ◽  
Author(s):  
Kamlesh Makwana ◽  
Matthew P. Sarnowski ◽  
Jiyuan Miao ◽  
Yu-Shan Lin ◽  
Juan Del Valle
Keyword(s):  
Small Molecules ◽  
Tau Protein ◽  
Supramolecular Assembly ◽  
Rational Basis ◽  
Protein Assemblies ◽  
Amino Groups ◽  
Minimalist Approach ◽  
Full Complement ◽  
Tau Aggregation

The spread of neurofibrillary tangles resulting from tau protein aggregation is a hallmark of Alzheimer’s and related neurodegenerative diseases. Early oligomerization of tau involves conformational reorganization into parallel b-sheet structures and supramolecular assembly into toxic fibrils. Despite the need for selective inhibitors of tau propagation, b-rich protein assemblies are inherently difficult to target with small molecules. Here, we describe a minimalist approach to mimic the aggregation-prone modules within tau. We carried out a backbone residue scan and show that amide N-amination completely abolishes the tendency of these peptides to self-aggregate, rendering them soluble mimics of ordered b-strands from the tau R2 and R3 domains. Several N-amino peptides (NAPs) inhibit disease-associated tau aggregation and prevent fibril formation <i>in vitro</i>. We further demonstrate that NAPs <b>12</b> and <b>13</b> are effective at blocking the cellular seeding of endogenous tau by interacting with both monomeric or fibrillar forms of extracellular tau. Peptidomimetic <b>12</b> is serum stable, non-toxic to neuronal cells, and selectivity inhibits the aggregation of tau over Ab<sub>­42</sub>. Structural analysis of our lead NAPs shows considerable conformational constraint imposed by the N-amino groups. The enhanced rigidity and full complement of sidechains thus enables NAPs to recognize tau fibrils. The described backbone N-amination approach provides a rational basis for the mimicry of other aggregation-prone peptides that drive pathogenic protein assembly.

scholarly journals From Aggregation to Inhibition: N-Amination Converts Amyloidogenic Tau Peptides into Soluble Antagonists of Cellular Seeding

2021 ◽  
Author(s):  
Kamlesh Makwana ◽  
Matthew P. Sarnowski ◽  
Jiyuan Miao ◽  
Yu-Shan Lin ◽  
Juan Del Valle
Keyword(s):  
Small Molecules ◽  
Tau Protein ◽  
Supramolecular Assembly ◽  
Rational Basis ◽  
Protein Assemblies ◽  
Amino Groups ◽  
Minimalist Approach ◽  
Full Complement ◽  
Tau Aggregation

The spread of neurofibrillary tangles resulting from tau protein aggregation is a hallmark of Alzheimer’s and related neurodegenerative diseases. Early oligomerization of tau involves conformational reorganization into parallel b-sheet structures and supramolecular assembly into toxic fibrils. Despite the need for selective inhibitors of tau propagation, b-rich protein assemblies are inherently difficult to target with small molecules. Here, we describe a minimalist approach to mimic the aggregation-prone modules within tau. We carried out a backbone residue scan and show that amide N-amination completely abolishes the tendency of these peptides to self-aggregate, rendering them soluble mimics of ordered b-strands from the tau R2 and R3 domains. Several N-amino peptides (NAPs) inhibit disease-associated tau aggregation and prevent fibril formation <i>in vitro</i>. We further demonstrate that NAPs <b>12</b> and <b>13</b> are effective at blocking the cellular seeding of endogenous tau by interacting with both monomeric or fibrillar forms of extracellular tau. Peptidomimetic <b>12</b> is serum stable, non-toxic to neuronal cells, and selectivity inhibits the aggregation of tau over Ab<sub>­42</sub>. Structural analysis of our lead NAPs shows considerable conformational constraint imposed by the N-amino groups. The enhanced rigidity and full complement of sidechains thus enables NAPs to recognize tau fibrils. The described backbone N-amination approach provides a rational basis for the mimicry of other aggregation-prone peptides that drive pathogenic protein assembly.

scholarly journals 2313 Characterization of the host pericyte role in glioblastoma angiogenesis

2018 ◽  
Vol 2 (S1) ◽  
pp. 1-1
Author(s):  
Frank Attenello ◽  
Frank Attenello ◽  
Yingxi Wu ◽  
Kathleen Tsung ◽  
William Mack ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Tumor Growth ◽  
Effect Size ◽  
Tumor Size ◽  
Lung Tumor ◽  
Cell Activity ◽  
Feasibility Studies ◽  
Wild Type ◽  
Cerebral Vessel

OBJECTIVES/SPECIFIC AIMS: Glioblastoma (GBM) carries a prognosis of 14.6 months mean survival despite maximal surgical, chemotherapeutic, and radiation therapy. The pericyte is a recently characterized cell shown to be a critical component of cerebral vessel physiology and pathology. Importantly, alterations in pericyte densities have shown resulting changes in breast and lung tumor growth. We leverage transgenic pericyte-deficient mouse models to evaluate resulting behavior of implanted patient-derived GBM. METHODS/STUDY POPULATION: Patient-derived, green fluorescent protein labeled, GBM will be implanted in right frontal bregma of both 6-month old pericyte-deficient (PDGFR+/−) mice and age-matched wild-type littermate controls (IACUC 20755, IRB 16-00929), which are immunosuppressed via daily intraperitoneal cyclosporine injection. In total, 30 mice of both genders are included in tumor and control cohorts. Fixed cortical sections following 3-week period will be stained for pericytes (NG2), endothelium (CD31), hypoxia (piminidazole), and tumor size. One-way ANOVA with will used to compare groups using SAS software (Cary, NC). RESULTS/ANTICIPATED RESULTS: Feasibility studies show robust in vitro growth of patient-derived GBM cells, showing continued growth over 10 cellular division passages. Lentivirally transduced GFP reveals reliable tumor tracking both in vitro and in vivo. Transgenic mice at 6 months display reproducibly decreased pericyte and microvascular density in triplicate. Wild-type mice tolerate tumor injection up to three weeks with visible tumor growth, peritumoral hypervascularity, and no evidence of mouse neural dysfunction. With current cohorts recently implanted with tumor, we anticipate a significant decrease in tumor size with Cohen’s d effect size of 0.5 in GBM implanted in pericyte-deficient mice when compared to control. Effect sizes are based moderate to large (effect size 0.5–0.8) reductions of in vitro GBM growth in vascular gene (TGF-β knockdown studies). In addition, tagged tumor-derived pericytes should comprise a greater proportion of new vasculature in pericyte-knockdown mice to overcome host pericyte depletion. Finally, tumors in transgenic mice should show increased hypoxia from limitations in angiogenesis. DISCUSSION/SIGNIFICANCE OF IMPACT: Feasibility studies show successful tracking of fluorescently tagged-patient derived GBM samples in transgenic mice with decreased vasculature. GBM grafts show no evidence of immunogenic response with cyclosporine protocol. Successful limitation of tumor size with reduced pericyte density will provide support to increasing study of blood-brain barrier, stem cell activity and inflammatory activity of pericyte microenvironments altering GBM behavior. Furthermore, implementation of known pericyte targeted therapies, such as imantinib, can be evaluated for GBM patient treatment efficacy. Studies with assembled clinical translational research scholar mentorship team will allow the principal investigator to develop an independent career with laboratory focused on contributing to improved patient outcomes, translating successful pericyte-targeted results to patient trials.

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