Oligomeric and Fibrillar Species of Amyloid-β Peptides Differentially Affect Neuronal Viability

Abstract

Genetic evidence predicts a causative role for amyloid-␤ (A␤) in Alzheimer's disease. Recent debate has focused on whether fibrils (amyloid) or soluble oligomers of A␤ are the active species that contribute to neurodegeneration and dementia. We developed two aggregation protocols for the consistent production of stable oligomeric or fibrillar preparations of A␤-(1-42). Here we report that oligomers inhibit neuronal viability 10-fold more than fibrils and ϳ40-fold more than unaggregated peptide, with oligomeric A␤-(1-42)-induced inhibition significant at 10 nM. Under A␤-(1-42) oligomer-and fibril-forming conditions, A␤-(1-40) remains predominantly as unassembled monomer and had significantly less effect on neuronal viability than preparations of A␤-(1-42). We applied the aggregation protocols developed for wild type A␤-(1-42) to A␤-(1-42) with the Dutch (E22Q) or Arctic (E22G) mutations. Oligomeric preparations of the mutations exhibited extensive protofibril and fibril formation, respectively, but were not consistently different from wild type A␤-(1-42) in terms of inhibition of neuronal viability. However, fibrillar preparations of the mutants appeared larger and induced significantly more inhibition of neuronal viability than wild type A␤-(1-42) fibril preparations. These data demonstrate that protocols developed to produce oligomeric and fibrillar A␤-(1-42) are useful in distinguishing the structural and functional differences between A␤-(1-42) and A␤-(1-40) and genetic mutations of A␤-(1-42). Amyloid-␤ (A␤)1 is derived by the proteolytic processing of amyloid precursor protein (APP), resulting in a peptide predominantly 40 or 42 amino acids in length. Mutations in APP and the presenilins that increase the amount of A␤-(1-42) cause AD (for review, see Ref. 1). Historically, the "amyloid cascade" hypothesis has defined the fibrillization of A␤ into amyloid deposits, a pathologic hallmark of AD, as a toxic gain of function (2). That A␤-(1-42) is more fibrillogenic than A␤-(1-40) fits well with this hypothesis. However, amyloid plaques do not always correlate in number, tempo, or distribution with neurodegeneration or clinical dementia. Thus, recent debate within the AD community has focused on whether fibrillar (amyloid) or soluble oligomers of A␤ are the active species of the peptide that ultimately cause the synaptic loss and dementia associated with AD (3-7). In vivo, small, stable oligomers of A␤-(1-42) have been isolated from brain, plasma, and cerebrospinal fluid (8 -10) and correlate with the severity of neurodegeneration in AD (11, 12). Thus, although genetic evidence predicts that A␤ is a causative factor in AD, the role of fibrillar and oligomeric A␤ in the pathogenesis of AD remains unclear.Initial in vitro studies suggested that A␤-induced neurotoxicity required the peptide to adopt a fibrillar aggregation state, with unaggregated peptide at low doses actually exhibiting neurotrophic effects (13)(14)(15)(16)(17)(18)). Recent studies demonstrate that non-fibrillar structures, including oligomers and ...

Prostaglandin I 2 upregulates the expression of anterior pharynx‐defective‐1α and anterior pharynx‐defective‐1β in amyloid precursor protein/presenilin 1 transgenic mice

“…The methods for preparing Aβ oligomers or fibrils had been described previously (Dahlgren et al ., 2002; Moore et al ., 2002; Pan et al ., 2011). The detailed information was provided in the supportive information.…”

Aminopyridazines inhibit β-amyloid-induced glial activation and neuronal damage in vivo

“…Briefly, C57/Bl6 mice weighing ∼25 g were anesthetized with vaporized halothane, and a micro-osmotic pump (Alzet #1002) was attached to a cannula (Plastics One) stereotaxically implanted into the right lateral cerebral ventricle (at coordinates −1.0 mm mediolateral and −0.5 mm anterioposterior from Bregma; −1.5 mm dorsal-ventral from skull). Pumps contained either oligomeric [4] A␤1-42 (45 g; American Peptide) in vehicle (4 mM Hepes + 250 g/ml human high-density lipoprotein, HDL) or vehicle alone. HDL, which normally carries A␤ in plasma, was used in the pump to reduce A␤ aggregation and act as an A␤ chaperone for better neuropil delivery [6,7].…”

Amyloid‐beta 1‐40 is associated with alterations in NG2+ pericyte population ex vivo and in vitro

“…This finding goes well with the previously mentioned study demonstrating degenerated pericytes adjacent to fibril Aβ1‐40‐formed CAA (Maia et al., 2007). The finding is also in line with several cell cultures studies demonstrating that the fibril form of Aβ1‐40 is toxic to neurons (Dahlgren et al., 2002; Okada, Ikeda, Wakabayashi, Ogawa & Matsuzaki, 2008; Walsh, Hartley, Condron, Selkoe & Teplow, 2001), endothelial cells (Hernandez‐Guillamon et al., 2010; Qosa, LeVine, Keller & Kaddoumi, 2014) and smooth muscle cells (Fossati et al., 2010). However, fibril preparations most likely also contain small portion of protofibrils (the first elongated unit appearing in fibril formation; Walsh, Lomakin, Benedek, Condron & Teplow, 1997), and since protofibrils of Aβ have some neurotoxic features (although the Aβ1‐42 protofibrils are substantially less toxic compared to Aβ1‐42 oligomers (Ahmed et al., 2010)), it is important to reflect upon the possibility that also our fibril preparations contain toxic protofibrils.…”

“…However, it is important to point out that Aβ can appear in several different species, whereof Aβ1‐42 and Aβ1‐40 are two of the most abundant forms. Alzheimer′s disease is foremost associated with the former specie, as insoluble Aβ1‐42 fibrils are the major component of Aβ plaques and soluble oligomer Aβ1‐42 is strongly neurotoxic (Dahlgren et al., 2002). The impact of Aβ1‐42 on brain pericytes in the human brain is scarcely investigated, but a few in vitro studies show reduced cell survival after prolonged Aβ1‐42 stimulation (Verbeek, de Waal, Schipper & Van Nostrand, 1997; Wilhelmus et al., 2007) and altered shedding of the pericyte adhesion molecule chondroitin sulphate proteoglycan NG2 in the presence of oligomer and fibril Aβ1‐42 (Schultz, Nielsen, Minthon & Wennstrom, 2014).…”