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Amyloid β oligomers (AβOs) accumulate early in Alzheimer's disease (AD) and experimentally cause memory dysfunction and the major pathologies associated with AD, for example, tau abnormalities, synapse loss, oxidative damage, and cognitive dysfunction. In order to develop the most effective AβO‐targeting diagnostics and therapeutics, the AβO structures contributing to AD‐associated toxicity must be elucidated. Here, we investigate the structural properties and pathogenic relevance of AβOs stabilized by the bifunctional crosslinker 1,5‐difluoro‐2,4‐dinitrobenzene (DFDNB). We find that DFDNB stabilizes synthetic Aβ in a soluble oligomeric conformation. With DFDNB, solutions of Aβ that would otherwise convert to large aggregates instead yield solutions of stable AβOs, predominantly in the 50–300 kDa range, that are maintained for at least 12 days at 37°C. Structures were determined by biochemical and native top–down mass spectrometry analyses. Assayed in neuronal cultures and i.c.v.‐injected mice, the DFDNB‐stabilized AβOs were found to induce tau hyperphosphorylation, inhibit choline acetyltransferase, and provoke neuroinflammation. Most interestingly, DFDNB crosslinking was found to stabilize an AβO conformation particularly potent in inducing memory dysfunction in mice. Taken together, these data support the utility of DFDNB crosslinking as a tool for stabilizing pathogenic AβOs in structure‐function studies. The structures of amyloid beta oligomers (AβOs) most germane to the pathogenesis of Alzheimer's disease (AD) are ill‐defined. Here, we demonstrate the utility of the crosslinker 1,5‐difluoro‐2,4‐dinitrobenzene (DFDNB) as a tool for AβO stabilization in structure–function studies. We find that DFDNB‐linked AβOs are detergent‐insensitive and remain in a soluble conformation with time and with increased concentration. Importantly, DFDNB stabilization does not alter the ability of AβOs to invoke potent responses than in multiple AD‐relevant pathogenic assays. [ABSTRACT FROM AUTHOR]

Brain accumulation of soluble oligomers of the amyloid-β peptide (AβOs) is increasingly considered a key early event in the pathogenesis of Alzheimer's disease (AD). A variety of AβO species have been identified, both in vitro and in vivo, ranging from dimers to 24mers and higher order oligomers. However, there is no consensus in the literature regarding which AβO species are most germane to AD pathogenesis. Antibodies capable of specifically recognizing defined subpopulations of AβOs would be a valuable asset in the identification, isolation, and characterization of AD-relevant AβO species. Here, we report the characterization of a human single chain antibody fragment (scFv) denoted NUsc1, one of a number of scFvs we have identified that stringently distinguish AβOs from both monomeric and fibrillar Aβ. NUsc1 readily detected AβOs previously bound to dendrites in cultured hippocampal neurons. In addition, NUsc1 blocked AβO binding and reduced AβO-induced neuronal oxidative stress and tau hyperphosphorylation in cultured neurons. NUsc1 further distinguished brain extracts from AD-transgenic mice from wild type (WT) mice, and detected endogenous AβOs in fixed AD brain tissue and AD brain extracts. Biochemical analyses indicated that NUsc1 targets a subpopulation of AβOs with apparent molecular mass greater than 50 kDa. Results indicate that NUsc1 targets a particular AβO species relevant to AD pathogenesis, and suggest that NUsc1 may constitute an effective tool for AD diagnostics and therapeutics. [ABSTRACT FROM AUTHOR]

Inflammatory activation precedes and correlates with accumulating τ lesions in Alzheimer's disease and tauopathies. However, the relationship between neuroinflammation and etiology of pathologic τ remains elusive. To evaluate whether inflammatory signaling may promote or accelerate neurofibrillary tangle pathology, we explored the effect of recombinant adeno-associated virus (rAAV)-mediated overexpression of a master inflammatory cytokine, IFN-γ, on κ phosphorylation. In initial studies in primary neuroglial cultures, rAAV-mediated expression of IFN-γ did not alter endogenous κ production or paired helical filament κ phosphorylation. Next, we tested the effect of rAAV-mediated expression of IFN-γ in the brains of 2 mouse models of tauopathy: JNPL3 and rTg4510. In both models, IFN-γ increased 1) signal transducer and activator of transcription 1 levels and gliosis, and 2) hyperphosphorylation and conformational alterations of soluble κ compared with control cohorts. However, sarkosyl-insoluble phosphorylated κ levels and ubiquitin staining were unaltered in the IFN-γ cohorts. Notably, IFN-γ-induced κ hyperphosphorylation was associated with release of the inhibitory effect of glycogen synthase kinase 3β function by decreasing Ser9 phosphorylation. Our data suggest that type II IFN signaling can promote κ phosphorylation by modulating cellular kinase activity, though this is insufficient in accelerating neuritic tangle pathology. [ABSTRACT FROM AUTHOR]