A fundamental question about the early pathogenesis of Alzheimer's disease (AD) concerns how toxic aggregates of amyloid ␤ protein (A␤) are formed from its nontoxic soluble form. We hypothesized previously that GM1 ganglioside-bound A␤ (GA␤) is involved in the process. We now examined this possibility using a novel monoclonal antibody raised against GA␤ purified from an AD brain. Here, we report that GA␤ has a conformation distinct from that of soluble A␤ and initiates A␤ aggregation by acting as a seed. Furthermore, GA␤ generation in the brain was validated by both immunohistochemical and immunoprecipitation studies. These results imply a mechanism underlying the onset of AD and suggest that an endogenous seed can be a target of therapeutic strategy.

The fliD operon of Salmonella typhimurium consists of three flagellar genes, fliD, fliS, and fliT, and is transcribed in this order. It has been shown that an fliD::Tn10 mutation causes an excess export of the flagellum-specific anti-sigma factor, FlgM, resulting in an overexpression of flagellar class 3 operons. In this study, using gene-disruption mutants in the individual genes in the fliD operon, we showed that mutations in any one of the genes in the operon enhanced both FlgM export and the expression of flagellar regulon. This indicates that all three genes in the operon are involved in the negative regulation of FlgM export.

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BackgroundSeveral lines of evidence indicate that memory loss represents a synaptic failure caused by soluble amyloid β (Aβ) oligomers. However, the pathological relevance of Aβ oligomers (AβOs) as the trigger of synaptic or neuronal degeneration, and the possible mechanism underlying the neurotoxic action of endogenous AβOs remain to be determined.ResultsTo specifically target toxic AβOs in vivo, monoclonal antibodies (1A9 and 2C3) specific to them were generated using a novel design method. 1A9 and 2C3 specifically recognize soluble AβOs larger than 35-mers and pentamers on Blue native polyacrylamide gel electrophoresis, respectively. Biophysical and structural analysis by atomic force microscopy (AFM) revealed that neurotoxic 1A9 and 2C3 oligomeric conformers displayed non-fibrilar, relatively spherical structure. Of note, such AβOs were taken up by neuroblastoma (SH-SY5Y) cell, resulted in neuronal death. In humans, immunohistochemical analysis employing 1A9 or 2C3 revealed that 1A9 and 2C3 stain intraneuronal granules accumulated in the perikaryon of pyramidal neurons and some diffuse plaques. Fluoro Jade-B binding assay also revealed 1A9- or 2C3-stained neurons, indicating their impending degeneration. In a long-term low-dose prophylactic trial using active 1A9 or 2C3 antibody, we found that passive immunization protected a mouse model of Alzheimer's disease (AD) from memory deficits, synaptic degeneration, promotion of intraneuronal AβOs, and neuronal degeneration. Because the primary antitoxic action of 1A9 and 2C3 occurs outside neurons, our results suggest that extracellular AβOs initiate the AD toxic process and intraneuronal AβOs may worsen neuronal degeneration and memory loss.ConclusionNow, we have evidence that HMW-AβOs are among the earliest manifestation of the AD toxic process in mice and humans. We are certain that our studies move us closer to our goal of finding a therapeutic target and/or confirming the relevance of our therapeutic strategy.

Monoclonal 2C3 specific to β-amyloid (Aβ) oligomers (AβOs) enabled us to test our hypothesis that the alteration of lipoprotein-Aβ interaction in the central nervous system (CNS) initiates and/or accelerates the cascade favoring Aβ assembly. Immunoprecipitation of frontal cortex employing 2C3 unequivocally detected soluble 4-, 8-, and 12-mers in Alzheimer's disease (AD) brains. Immunoblot analysis of the entorhinal cortex employing 2C3 revealed that the accumulation of soluble 12-mers precedes the appearance of neuronal loss or cognitive impairment and is enhanced as the Braak neurofibrially tangle (NFT) stages progress. The dissociation of soluble Aβ from lipoprotein particles occurs in cerebrospinal fluid (CSF), and the presence of lipoprotein-free oligomeric 2C3 conformers (4- to 35-mers) was evident, which mimic CNS environments. Such CNS environments may strongly affect conformation of soluble Aβ peptides, resulting in the conversion of soluble Aβ(42) monomers into soluble Aβ(42) assembly. The findings suggest that functionally declined lipoproteins may accelerate the generation of metabolic conditions leading to higher levels of soluble Aβ(42) assembly in the CNS.