Tabela com principais espec.
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H||IP, ICC, IHC, ELISA, WB, DB||Rb||Serum||Polyclonal Antibody|
|Presentation||Unpurified rabbit polyclonal antibody serum containing 0.05% sodium azide.|
|Safety Information according to GHS|
|Material Size||100 µL|
|Título||Número do lote|
|Anti-Amyloid Fibrils OC - 2147135||2147135|
|Anti-Amyloid Fibrils OC - 2424776||2424776|
|Anti-Amyloid Fibrils OC - 1966929||1966929|
|Anti-Amyloid Fibrils OC - 1993788||1993788|
|Anti-Amyloid Fibrils OC - 2022060||2022060|
|Anti-Amyloid Fibrils OC - 2038298||2038298|
|Anti-Amyloid Fibrils OC - 2073638||2073638|
|Anti-Amyloid Fibrils OC - 2287786||2287786|
|Anti-Amyloid Fibrils OC - 2345063||2345063|
|Anti-Amyloid Fibrils OC - NG1648320||NG1648320|
|Anti-Amyloid Fibrils OC - NG1732175||NG1732175|
|Anti-Amyloid Fibrils OC - NG1865063||NG1865063|
|Anti-Amyloid Fibrils OC - NG1916160||NG1916160|
|Anti-Amyloid Fibrils OC -2489077||2489077|
|Anti-Amyloid Fibrils OC -2554096||2554096|
|Anti-Amyloid Fibrils OC -2587247||2587247|
|Anti-Amyloid Fibrils OC -2620587||2620587|
|Anti-Amyloid Fibrils OC -2649156||2649156|
|Anti-Amyloid Fibrils OC -2682434||2682434|
|Anti-Amyloid Fibrils OC -2726819||2726819|
|Anti-Amyloid Fibrils OC -2772691||2772691|
|Anti-Amyloid Fibrils OC -2834757||2834757|
Referências | 12 Disponível | Ver todas as referências
|Visão geral das referências||Aplicação||Pub Med ID|
|Characterization of a Novel Mouse Model of Alzheimer's Disease--Amyloid Pathology and Unique β-Amyloid Oligomer Profile. |
Liu, P; Paulson, JB; Forster, CL; Shapiro, SL; Ashe, KH; Zahs, KR
PloS one 10 e0126317 2015
Amyloid plaques composed of β-amyloid (Aβ) protein are a pathological hallmark of Alzheimer's disease. We here report the generation and characterization of a novel transgenic mouse model of Aβ toxicity. The rTg9191 mice harbor a transgene encoding the 695 amino-acid isoform of human amyloid precursor protein (APP) with the Swedish and London mutations (APPNLI) linked to familial Alzheimer's disease, under the control of a tetracycline-response element, as well as a transgene encoding the tetracycline transactivator, under the control of the promoter for calcium-calmodulin kinase IIα. In these mice, APPNLI is expressed at a level four-fold that of endogenous mouse APP and its expression is restricted to forebrain regions. Transgene expression was suppressed by 87% after two months of doxycycline administration. Histologically, we showed that (1) Aβ plaques emerged in cerebral cortex and hippocampus as early as 8 and 10.5-12.5 months of age, respectively; (2) plaque deposition progressed in an age-dependent manner, occupying up to 19% of cortex at ~25 months of age; and (3) neuropathology--such as abnormal neuronal architecture, tau hyperphosphorylation and misfolding, and neuroinflammation--was observed in the vicinity of neuritic plaques. Biochemically, we determined total Aβ production at varied ages of mice, and we showed that mice produced primarily fibrillar Aβ assemblies recognized by conformation-selective OC antibodies, but few non-fibrillar oligomers (e.g., Aβ*56) detectable by A11 antibodies. Finally, we showed that expression of the tetracycline transactivator resulted in reduced brain weight and smaller dentate-gyrus size. Collectively, these data indicate that rTg9191 mice may serve as a model for studying the neurological effects of the fibrillar Aβ assemblies in situ.
|Cytotoxic helix-rich oligomer formation by melittin and pancreatic polypeptide. |
Singh, PK; Ghosh, D; Tewari, D; Mohite, GM; Carvalho, E; Jha, NN; Jacob, RS; Sahay, S; Banerjee, R; Bera, AK; Maji, SK
PloS one 10 e0120346 2015
Conversion of amyloid fibrils by many peptides/proteins involves cytotoxic helix-rich oligomers. However, their toxicity and biophysical studies remain largely unknown due to their highly dynamic nature. To address this, we chose two helical peptides (melittin, Mel and pancreatic polypeptide, PP) and studied their aggregation and toxicity. Mel converted its random coil structure to oligomeric helical structure upon binding to heparin; however, PP remained as helix after oligomerization. Interestingly, similar to Parkinson's associated α-synuclein (AS) oligomers, Mel and PP also showed tinctorial properties, higher hydrophobic surface exposure, cellular toxicity and membrane pore formation after oligomerization in the presence of heparin. We suggest that helix-rich oligomers with exposed hydrophobic surface are highly cytotoxic to cells irrespective of their disease association. Moreover as Mel and PP (in the presence of heparin) instantly self-assemble into stable helix-rich amyloidogenic oligomers; they could be represented as models for understanding the biophysical and cytotoxic properties of helix-rich intermediates in detail.
|Amyloid properties of the mouse egg zona pellucida. |
Egge, N; Muthusubramanian, A; Cornwall, GA
PloS one 10 e0129907 2015
The zona pellucida (ZP) surrounding the oocyte is an extracellular fibrillar matrix that plays critical roles during fertilization including species-specific gamete recognition and protection from polyspermy. The mouse ZP is composed of three proteins, ZP1, ZP2, and ZP3, all of which have a ZP polymerization domain that directs protein fibril formation and assembly into the three-dimensional ZP matrix. Egg coats surrounding oocytes in nonmammalian vertebrates and in invertebrates are also fibrillar matrices and are composed of ZP domain-containing proteins suggesting the basic structure and function of the ZP/egg coat is highly conserved. However, sequence similarity between ZP domains is low across species and thus the mechanism for the conservation of ZP/egg coat structure and its function is not known. Using approaches classically used to identify amyloid including conformation-dependent antibodies and dyes, X-ray diffraction, and negative stain electron microscopy, our studies suggest the mouse ZP is a functional amyloid. Amyloids are cross-β sheet fibrillar structures that, while typically associated with neurodegenerative and prion diseases in mammals, can also carry out functional roles in normal cells without resulting pathology. An analysis of the ZP domain from mouse ZP3 and ZP3 homologs from five additional taxa using the algorithm AmylPred 2 to identify amyloidogenic sites, revealed in all taxa a remarkable conservation of regions that were predicted to form amyloid. This included a conserved amyloidogenic region that localized to a stretch of hydrophobic amino acids previously shown in mouse ZP3 to be essential for fibril assembly. Similarly, a domain in the yeast protein α-agglutinin/Sag 1p, that possesses ZP domain-like features and which is essential for mating, also had sites that were predicted to be amyloidogenic including a hydrophobic stretch that appeared analogous to the critical site in mouse ZP3. Together, these studies suggest that amyloidogenesis may be a conserved mechanism for ZP structure and function across billions of years of evolution.
|Microglia constitute a barrier that prevents neurotoxic protofibrillar Aβ42 hotspots around plaques. |
Condello, C; Yuan, P; Schain, A; Grutzendler, J
Nature communications 6 6176 2015
In Alzheimer's disease (AD), β-amyloid (Aβ) plaques are tightly enveloped by microglia processes, but the significance of this phenomenon is unknown. Here we show that microglia constitute a barrier with profound impact on plaque composition and toxicity. Using high-resolution confocal and in vivo two-photon imaging in AD mouse models, we demonstrate that this barrier prevents outward plaque expansion and leads to compact plaque microregions with low Aβ42 affinity. Areas uncovered by microglia are less compact but have high Aβ42 affinity, leading to the formation of protofibrillar Aβ42 hotspots that are associated with more severe axonal dystrophy. In ageing, microglia coverage is reduced leading to enlarged protofibrillar Aβ42 hotspots and more severe neuritic dystrophy. CX3CR1 gene deletion or anti-Aβ immunotherapy causes expansion of microglia coverage and reduced neuritic dystrophy. Failure of the microglia barrier and the accumulation of neurotoxic protofibrillar Aβ hotspots may constitute novel therapeutic and clinical imaging targets for AD.
|Forebrain microglia from wild-type but not adult 5xFAD mice prevent amyloid-β plaque formation in organotypic hippocampal slice cultures. |
Hellwig, S; Masuch, A; Nestel, S; Katzmarski, N; Meyer-Luehmann, M; Biber, K
Scientific reports 5 14624 2015
The role of microglia in amyloid-β (Aβ) deposition is controversial. In the present study, an organotypic hippocampal slice culture (OHSC) system with an in vivo-like microglial-neuronal environment was used to investigate the potential contribution of microglia to Aβ plaque formation. We found that microglia ingested Aβ, thereby preventing plaque formation in OHSCs. Conversely, Aβ deposits formed rapidly in microglia-free wild-type slices. The capacity to prevent Aβ plaque formation was absent in forebrain microglia from young adult but not juvenile 5xFamilial Alzheimer's disease (FAD) mice. Since no loss of Aβ clearance capacity was observed in both wild-type and cerebellar microglia from 5xFAD animals, the high Aβ1-42 burden in the forebrain of 5xFAD animals likely underlies the exhaustion of microglial Aβ clearance capacity. These data may therefore explain why Aβ plaque formation has never been described in wild-type mice, and point to a beneficial role of microglia in AD pathology. We also describe a new method to study Aβ plaque formation in a cell culture setting.
|Functional amyloids in the mouse sperm acrosome. |
Guyonnet, B; Egge, N; Cornwall, GA
Molecular and cellular biology 34 2624-34 2014
The acrosomal matrix (AM) is an insoluble structure within the sperm acrosome that serves as a scaffold controlling the release of AM-associated proteins during the sperm acrosome reaction. The AM also interacts with the zona pellucida (ZP) that surrounds the oocyte, suggesting a remarkable stability that allows its survival despite being surrounded by proteolytic and hydrolytic enzymes released during the acrosome reaction. To date, the mechanism responsible for the stability of the AM is not known. Our studies demonstrate that amyloids are present within the sperm AM and contribute to the formation of an SDS- and formic-acid-resistant core. The AM core contained several known amyloidogenic proteins, as well as many proteins predicted to form amyloid, including several ZP binding proteins, suggesting a functional role for the amyloid core in sperm-ZP interactions. While stable at pH 3, at pH 7, the sperm AM rapidly destabilized. The pH-dependent dispersion of the AM correlated with a change in amyloid structure leading to a loss of mature forms and a gain of immature forms, suggesting that the reversal of amyloid is integral to AM dispersion.
|Calcium ions promote superoxide dismutase 1 (SOD1) aggregation into non fibrillar amyloid: a link to toxic effects of calcium overload in amyotrophic lateral sclerosis (ALS)? |
Leal, Sónia S, et al.
J. Biol. Chem., (2013) 2013
Imbalance in metal ion homeostasis is a hallmark in neurodegenerative conditions involving protein deposition, and amyotrophic lateral sclerosis (ALS) is no exception. In particular, Ca(2+) dysregulation has been shown to correlate with superoxide dismutase-1 (SOD1) aggregation in a cellular model of ALS. Here we present evidence that SOD1 aggregation is enhanced and modulated by Ca(2+). We show that at physiological pH, Ca(2+) induces conformational changes that increase SOD1 β-sheet content, as probed by far UV-CD and ATR-FTIR, and enhances SOD1 hydrophobicity as probed by ANS fluorescence emission. Moreover, dynamic light scattering analysis showed that Ca(2+) boosts the onset of SOD1 aggregation. In agreement, Ca(2+) decreases SOD1 critical concentration and nucleation time during aggregation kinetics, as evidenced by Thioflavin-T fluorescence emission. ATR FT-IR analysis showed that Ca(2+) induced aggregates consisting preferentially of antiparallel β-sheets, thus suggesting a modulation effect on the aggregation pathway. Transmission electron microscopy and analysis with conformational anti-fibrils and anti-oligomers antibodies showed that oligomers and amyloidogenic aggregates constitute the prevalent morphology of Ca(2+) induced aggregates, thus indicating that Ca(2+) diverts SOD1 aggregation from fibrils towards amorphous aggregates. Interestingly, the same heterogeneity of conformations is found in ALS derived protein inclusions. We thus hypothesize that transient variations and dysregulation of cellular Ca(2+) levels contribute to the formation of SOD1 aggregates in ALS patients. In this scenario, Ca(2+) may be considered as a pathogenic effector in the formation of ALS proteinaceous inclusions.
|Intrinsically disordered and aggregation prone regions underlie β-aggregation in S100 proteins. |
Carvalho, SB; Botelho, HM; Leal, SS; Cardoso, I; Fritz, G; Gomes, CM
PloS one 8 e76629 2013
S100 proteins are small dimeric calcium-binding proteins which control cell cycle, growth and differentiation via interactions with different target proteins. Intrinsic disorder is a hallmark among many signaling proteins and S100 proteins have been proposed to contain disorder-prone regions. Interestingly, some S100 proteins also form amyloids: S100A8/A9 forms fibrils in prostatic inclusions and S100A6 fibrillates in vitro and seeds SOD1 aggregation. Here we report a study designed to investigate whether β-aggregation is a feature extensive to more members of S100 family. In silico analysis of seven human S100 proteins revealed a direct correlation between aggregation and intrinsic disorder propensity scores, suggesting a relationship between these two independent properties. Averaged position-specific analysis and structural mapping showed that disorder-prone segments are contiguous to aggregation-prone regions and that whereas disorder is prominent on the hinge and target protein-interaction regions, segments with high aggregation propensity are found in ordered regions within the dimer interface. Acidic conditions likely destabilize the seven S100 studied by decreasing the shielding of aggregation-prone regions afforded by the quaternary structure. In agreement with the in silico analysis, hydrophobic moieties become accessible as indicated by strong ANS fluorescence. ATR-FTIR spectra support a structural inter-conversion from α-helices to intermolecular β-sheets, and prompt ThT-binding takes place with no noticeable lag phase. Dot blot analysis using amyloid conformational antibodies denotes a high diversity of conformers; subsequent analysis by TEM shows fibrils as dominant species. Altogether, our data suggests that β-aggregation and disorder-propensity are related properties in S100 proteins, and that the onset of aggregation is likely triggered by loss of protective tertiary and quaternary interactions.
|S100A6 amyloid fibril formation is calcium-modulated and enhances superoxide dismutase-1 (SOD1) aggregation. |
Botelho, Hugo M, et al.
J. Biol. Chem., (2012) 2012
S100A6 is a small EF-hand calcium- and zinc- binding protein involved in the regulation of cell proliferation and cytoskeletal dynamics. It is overexpressed in neurodegenerative disorders and a proposed marker for Amyotrophic Lateral Sclerosis (ALS). Following on recent reports of amyloid formation by S100 proteins, we investigated the aggregation properties of S100A6. Computational analysis using aggregation predictors Waltz and Zyggregator revealed increased propensity within S100A6 helices HI and HIV. Subsequent analysis of Thioflavin-T binding kinetics under acidic conditions elicited a very fast process with no lag phase and extensive formation of aggregates and stacked fibrils as observed by electron microscopy. Ca(2+) exerted an inhibitory effect on the aggregation kinetics, which could be reverted upon chelation. An FT-IR investigation of the early conformational changes occurring under these conditions showed that Ca(2+) promotes anti-parallel β-sheet conformations that repress fibrillization. At pH 7, Ca(2+) rendered the fibril formation kinetics slower: time-resolved imaging showed that fibril formation is highly suppressed, with aggregates forming instead. In the absence of metals an extensive network of fibrils is formed. S100A6 oligomers, but not fibrils, were found to be cytotoxic, decreasing cell viability by up to 40%. This effect was not observed when the aggregates were formed in the presence of Ca(2+). Interestingly, native S1006 seeds SOD1 aggregation, shortening its nucleation process. This suggests a cross-talk between these two proteins involved in ALS. Overall, these results put forward novel roles for S100 proteins, whose metal-modulated aggregation propensity may be a key aspect in their physiology and function.
|Methylene blue modulates huntingtin aggregation intermediates and is protective in Huntington's disease models. |
Sontag, EM; Lotz, GP; Agrawal, N; Tran, A; Aron, R; Yang, G; Necula, M; Lau, A; Finkbeiner, S; Glabe, C; Marsh, JL; Muchowski, PJ; Thompson, LM
The Journal of neuroscience : the official journal of the Society for Neuroscience 32 11109-19 2012
Huntington's disease (HD) is a devastating neurodegenerative disorder with no disease-modifying treatments available. The disease is caused by expansion of a CAG trinucleotide repeat and manifests with progressive motor abnormalities, psychiatric symptoms, and cognitive decline. Expression of an expanded polyglutamine repeat within the Huntingtin (Htt) protein impacts numerous cellular processes, including protein folding and clearance. A hallmark of the disease is the progressive formation of inclusions that represent the culmination of a complex aggregation process. Methylene blue (MB), has been shown to modulate aggregation of amyloidogenic disease proteins. We investigated whether MB could impact mutant Htt-mediated aggregation and neurotoxicity. MB inhibited recombinant protein aggregation in vitro, even when added to preformed oligomers and fibrils. MB also decreased oligomer number and size and decreased accumulation of insoluble mutant Htt in cells. In functional assays, MB increased survival of primary cortical neurons transduced with mutant Htt, reduced neurodegeneration and aggregation in a Drosophila melanogaster model of HD, and reduced disease phenotypes in R6/2 HD modeled mice. Furthermore, MB treatment also promoted an increase in levels of BDNF RNA and protein in vivo. Thus, MB, which is well tolerated and used in humans, has therapeutic potential for HD.
|Fibril specific, conformation dependent antibodies recognize a generic epitope common to amyloid fibrils and fibrillar oligomers that is absent in prefibrillar oligomers. |
Kayed, Rakez, et al.
Mol Neurodegener, 2: 18 (2007) 2007
ABSTRACT: BACKGROUND: Amyloid-related degenerative diseases are associated with the accumulation of misfolded proteins as amyloid fibrils in tissue. In Alzheimer disease (AD), amyloid accumulates in several distinct types of insoluble plaque deposits, intracellular Abeta and as soluble oligomers and the relationships between these deposits and their pathological significance remains unclear. Conformation dependent antibodies have been reported that specifically recognize distinct assembly states of amyloids, including prefibrillar oligomers and fibrils. RESULTS: We immunized rabbits with a morphologically homogeneous population of Abeta42 fibrils. The resulting immune serum (OC) specifically recognizes fibrils, but not random coil monomer or prefibrillar oligomers, indicating fibrils display a distinct conformation dependent epitope that is absent in prefibrillar oligomers. The fibril epitope is also displayed by fibrils of other types of amyloids, indicating that the epitope is a generic feature of the polypeptide backbone. The fibril specific antibody also recognizes 100,000 x G soluble fibrillar oligomers ranging in size from dimer to greater than 250 kDa on western blots. The fibrillar oligomers recognized by OC are immunologically distinct from prefibrillar oligomers recognized by A11, even though their sizes overlap broadly, indicating that size is not a reliable indicator of oligomer conformation. The immune response to prefibrillar oligomers and fibrils is not sequence specific and antisera of the same specificity are produced in response to immunization with islet amyloid polypeptide prefibrillar oligomer mimics and fibrils. The fibril specific antibodies stain all types of amyloid deposits in human AD brain. Diffuse amyloid deposits stain intensely with anti-fibril antibody although they are thioflavin S negative, suggesting that they are indeed fibrillar in conformation. OC also stains islet amyloid deposits in transgenic mouse models of type II diabetes, demonstrating its generic specificity for amyloid fibrils. CONCLUSION: Since the fibril specific antibodies are conformation dependent, sequence-independent, and recognize epitopes that are distinct from those present in prefibrillar oligomers, they may have broad utility for detecting and characterizing the accumulation of amyloid fibrils and fibrillar type oligomers in degenerative diseases.
|TGFbeta1 activates c-Jun and Erk1 via alphaVbeta6 integrin. |
Luettich, K; Schmidt, C
Molecular cancer 2 33 2003
Transforming growth factor beta (TGFbeta) plays an important role in animal development and many cellular processes. A variety of cellular functions that are required for tumor metastasis are controlled by integrins, a family of cell adhesion receptors. Overexpression of alphaVbeta6 integrin is associated with lymph node metastasis of gastric carcinomas. It has been demonstrated that a full TGFbeta1 signal requires both alphaVbeta6 integrin and SMAD pathway. TGFbeta1 binds to alphaVbeta6 via the DLXXL motif, a freely accessible amino acid sequence in the mature form of TGFbeta1. Binding of mature TGFbeta1 to alphaVbeta6 leads to immobilization and tyrosine phosphorylation of proteins, which are associated with focal adhesions, a hallmark of integrin-mediated signal transduction. Here, we show that binding of mature TGFbeta1 recruits the mitogen-activated protein kinase kinase kinase 1 (MEKK1), a mediator of c-Jun activation, and the extracellular signaling-regulated kinase-1 (Erk1) to focal adhesions. In addition, the p21-activated kinase 1 (PAK1) is associated with focal adhesions and differentially phosphorylated upon TGFbeta1 stimulation. We conclude that TGFbeta1 activates c-Jun via the MEKK1/p38 MAP kinase pathway and influences cytoskeletal organization. These finding may provide a link between TGFbeta1 and the metastatic behavior of cancers.