Substoichiometric Levels of Cu2+ Ions Accelerate the Kinetics of Fiber Formation and Promote Cell Toxicity of Amyloid-β from Alzheimer Disease

Abstract

A role for Cu 2؉ ions in Alzheimer disease is often disputed, as it is believed that Cu 2؉ ions only promote nontoxic amorphous aggregates of amyloid-␤ (A␤). In contrast with currently held opinion, we show that the presence of substoichiometric levels of Cu 2؉ ions in fact doubles the rate of production of amyloid fibers, accelerating both the nucleation and elongation of fiber formation. We suggest that binding of Cu 2؉ ions at a physiological pH causes A␤ to approach its isoelectric point, thus inducing self-association and fiber formation. We further show that Cu 2؉ ions bound to A␤ are consistently more toxic to neuronal cells than A␤ in the absence of Cu 2؉ ions, whereas Cu 2؉ ions in the absence of A␤ are not cytotoxic. The degree of Cu-A␤ cytotoxicity correlates with the levels of Cu 2؉ ions that accelerate fiber formation. We note the effect appears to be specific for Cu 2؉ ions as Zn 2؉ ions inhibit the formation of fibers. An active role for Cu 2؉ ions in accelerating fiber formation and promoting cell death suggests impaired copper homeostasis may be a risk factor in Alzheimer disease. Alzheimer disease (AD)2 is characterized by extracellular amyloid plaques, composed predominantly of fibrillar amyloid-␤ peptide (A␤), a 39 -43-residue peptide. Genetic alterations underlying familial AD are associated with mutations or increased production of A␤, indicating that A␤ plays a central role in the disease (1).A notable characteristic of AD is altered metal ion concentrations in the brain and disrupted metal ion homeostasis (2). Cu 2ϩ ions are found concentrated within senile plaques of AD patients directly bound to A␤ (3-5). Recent in vivo studies using a Drosophila model of AD have shown that impaired copper homeostasis enhances the toxic effects of A␤ (6). Furthermore, copper in a cholesterol high diet induces amyloid plaques and learning deficits in a rabbit model of AD (7). Other in vivo studies have shown that copper homeostasis can influence AD pathology. In contrast to the Drosophila model, transgenic mice have shown a reduced AD pathology with increased intracellular copper levels (8 -10).Although studies of A␤ neurotoxicity suggest that small diffusible oligomers, rather than mature amyloid fibers, are the more toxic form (11, 12), there remains strong evidence suggesting that amyloid plaques, or possibly intermediates of the fibrils, are critical in neuronal toxicity (13,14). A␤ oligomers may be precursors of fiber formation and may also arise from fiber fragmentation. Alternatively, oligomers may be in competition with fiber formation. Both possibilities require the self-association of monomeric A␤, and thus factors that affect fibrillization will also influence oligomer generation.The mechanism by which A␤ is toxic is hotly debated (11, 15). It has been proposed that A␤ can form ion channels or pores or can thin the membrane, all of which will cause membrane leakage and loss of cellular Ca 2ϩ ion homeostasis. One popular hypothesis is that the membrane integrity is compromised by lipid peroxida...

Cu(II) Mediates Kinetically Distinct, Non-amyloidogenic Aggregation of Amyloid-β Peptides

“…Thus, compounds that can modulate or shift the equilibrium of the A␤-Cu(II) complex to a less stable conformation or toward a non-toxic aggregated state could be a new therapeutic strategy for treating AD. Here it should be mentioned that a recent study showed, contrary to the previous reports, that it is the substoichiometric Cu(II) concentrations that are most cytotoxic, whereas an equimolar Cu(II) concentration does not increase A␤ cytotoxicity, and supra-stoichiometric Cu(II) concentrations reduce cytotoxicity (24).…”

“…In all events elucidating the underlying molecular mechanisms of the A␤-Cu, interactions are vital for understanding the role of Cu(II) in the pathology of AD, and hence in developing new therapeutic strategies for the treatment of AD (7,22). Despite extensive studies, however, these mechanisms remain largely unsettled and conflicting effects have been reported; in some studies Cu(II) appears to be involved in amyloid oligomerization (4,23,24), other studies report that Cu(II) does enhance aggregation but only along non-amyloidogenic pathways (17,25), or even that Cu(II) inhibits the aggregation of A␤ (26). We recently showed that Cu(II) could induce both oligomerization and non-oligomerization pathways of A␤ on the millisecond-second time scale (27).…”

Truncated Amyloid-β(11–40/42) from Alzheimer Disease Binds Cu2+ with a Femtomolar Affinity and Influences Fiber Assembly

“…A␤ forms fibers much faster than A␤ and is also more toxic in vivo (61)(62)(63)(64). The heightened rate of fiber nucleation and elongation observed for A␤ (11-40/42) may be caused by the loss of negatively charged side chains, as it is known that the pI and charge of A␤ have a profound influence on its rate of fiber formation (22,65). The absence of a number of charged residues at the N terminus increases the theoretical pI from 5.1 for A␤ (1-40/42) to 6.0 for A␤ (11-40/42) .…”

Human Serum Albumin Can Regulate Amyloid-β Peptide Fiber Growth in the Brain Interstitium

“…Albumin is also responsible for the transport of the labile pool of Cu 2ϩ ions in blood plasma (21). Of note, the A␤-Cu 2ϩ interaction and its role in AD have received significant interest (22)(23)(24), and Cu 2ϩ may be transferred from A␤ to HSA (25), meaning that albumin could have two mechanisms by which it inhibits fiber formation.…”