In Vitro Studies of Flemish, Dutch, and Wild-Type β-Amyloid Provide Evidence for Two-Staged Neurotoxicity

Rempel

Proc. Natl. Acad. Sci. U.S.A.

et al. 2013

113

179

Probing the conformational changes of amyloid beta (Aβ) peptide aggregation is challenging owing to the vast heterogeneity of the resulting soluble aggregates. To investigate the formation of these aggregates in solution, we designed an MS-based biophysical approach and applied it to the formation of soluble aggregates of the Aβ 42 peptide, the proposed causative agent in Alzheimer's disease. The approach incorporates pulsed hydrogen-deuterium exchange coupled with MS analysis. The combined approach provides evidence for a self-catalyzed aggregation with a lag phase, as observed previously by fluorescence methods. Unlike those approaches, pulsed hydrogen-deuterium exchange does not require modified Aβ 42 (e.g., labeling with a fluorophore). Furthermore, the approach reveals that the center region of Aβ 42 is first to aggregate, followed by the C and N termini. We also found that the lag phase in the aggregation of soluble species is affected by temperature and Cu 2+ ions. This MS approach has sufficient structural resolution to allow interrogation of Aβ aggregation in physiologically relevant environments. This platform should be generally useful for investigating the aggregation of other amyloid-forming proteins and neurotoxic soluble peptide aggregates.soluble Aβ oligomers | amyloid beta peptide | copper | electrospray ionization | Finke-Watsky mode P rotein aggregation is one of the immediate causes of Alzheimer's, Parkinson, and Huntington diseases, motivating biophysical studies of the responsible proteins. More than 20 small proteins undergo amyloidosis in humans. In Alzheimer's disease (AD), the aggregation of the 40-or 42-aa-long amyloid beta (Aβ) peptide, generally called Aβ 40 or Aβ 42 , respectively, is proposed to be involved in the onset of the disease (1, 2). Aβ 42 is more amyloidogenic and more neurotoxic than Aβ 40 . Although the amyloid-cascade hypothesis suggests that the Aβ-containing amyloid plaques are responsible for neurodegeneration (3-7), other studies suggest that soluble aggregates of Aβ 42 are more neurotoxic than the amyloid plaques (8-13).The amyloid plaques in AD-affected brains contain high levels of copper, zinc, and iron (14-20). Among these, Cu has drawn the most attention because the Aβ precursor protein is likely a Cuchaperone protein (21). Several studies of Cu 2+ -Aβ 40 interactions show that Cu 2+ can promote Aβ 40 aggregation (14,18,19).The structure of Aβ 42 and its aggregates, although studied extensively, remains of high interest. Studies of amyloid fibrils invoke X-ray crystallography (22-24), EM (19,25,26), and thioflavin T fluorescence (19, 27), revealing the polypeptide's global behavior, whereas NMR studies provide residue-level information for the fibrils (28-30). Nevertheless, we know little about soluble Aβ aggregates owing to their intrinsically high heterogeneity.MS should offer an opportunity for investigating soluble aggregates of Aβ 42 . Thus far, there are no MS-based, timedependent studies of the formation of soluble aggregates. Moreover, there are no ot...

Section: Application Of Finke-watzky Modeling and Statistical Evaluatmentioning

confidence: 99%

Pulsed hydrogen–deuterium exchange mass spectrometry probes conformational changes in amyloid beta (Aβ) peptide aggregation

Rempel

Proc. Natl. Acad. Sci. U.S.A.

et al. 2013

113

179

“…However, it can become a harmful factor for mature brains under injured or diseased circumstances, such as brain neuronal loss in neurodegenerative diseases (Anderson et al, 1996;Estus et al, 1997;Troy et al, 2000;Yuan and Yankner, 2000;Jellinger and Stadelmann, 2001;Morishima et al, 2001;Cregan et al, 2002;Galvan et al, 2002;Kumar-Singh et al, 2002;Wei et al, 2002;Eberhardt and Schulz, 2003;Koriyama et al, 2003;Lopez Salon et al, 2003;Tamagno et al, 2003). Alzheimer disease (AD) is a neurodegenerative disease and is characterized by significant neuronal loss.…”

Section: Introductionmentioning

confidence: 99%

Tumor Necrosis Factor Death Receptor Signaling Cascade Is Required for Amyloid-β Protein-Induced Neuron Death

et al. 2004

Tumor necrosis factor type I receptor (TNFRI), a death receptor, mediates apoptosis and plays a crucial role in the interaction between the nervous and immune systems. A direct link between death receptor activation and signal cascade-mediated neuron death in brains with neurodegenerative disorders remains inconclusive. Here, we show that amyloid-␤ protein (A␤), a major component of plaques in the Alzheimer's diseased brain, induces neuronal apoptosis through TNFRI by using primary neurons overexpressing TNFRI by viral infection or neurons from TNFRI knock-out mice. This was mediated via alteration of apoptotic protease-activating factor (Apaf-1) expression that in turn induced activation of nuclear factor B (NF-B). A␤-induced neuronal apoptosis was reduced with lower Apaf-1 expression, and little NF-B activation was found in the neurons with mutated Apaf-1 or a deletion of TNFRI compared with the cells from wild-type (WT) mice. Our studies suggest a novel neuronal response of A␤, which occurs through a TNF receptor signaling cascade and a caspase-dependent death pathway.

“…A similar result was obtained when the neurotoxic effects of the Dutch Aβ were compared with those of the Flemish mutant and WT Aβ in a neuroblastoma cell line incubated in the presence of these peptides for a very short time (2 h). In contrast, E22Q Aβ was more toxic than the Flemish mutant and WT Aβ with longer incubation times (24 h) (39). The enhanced survival observed in E22Q Aβ-treated hippocampal neurons could be due to the reduced rate of calpain-mediated tau cleavage leading to the generation of the 17-kDa tau fragment.…”

Section: Discussionmentioning

confidence: 91%

β-Amyloid Carrying the Dutch Mutation Has Diverse Effects on Calpain-Mediated Toxicity in Hippocampal Neurons

Nicholson

Wold

Walsh

et al. 2011

Mol Med

Hereditary cerebral hemorrhage with amyloidosis-Dutch type is a disorder associated with a missense mutation (E693Q) in the β-amyloid (Aβ)-coding region of the amyloid precursor protein (APP). This familial disease is characterized by cognitive deficits secondary to intracerebral hemorrhage and, in some cases, progressive Alzheimer's disease (AD)-like dementia. Although this mutation was the first ever reported in the human APP gene, little is known about the molecular mechanisms underlying the direct toxic effects of this mutated Aβ on central neurons. In the present study, we assessed the role of calpain-mediated toxicity in such effects using an AD primary culture model system. Our results showed that Dutch mutant Aβ (E22Q) induced calpain-mediated cleavage of dynamin 1 and a significant decrease in synaptic contacts in mature hippocampal cultures. These synaptic deficits were similar to those induced by wild-type (WT) Aβ. In contrast, calpain-mediated tau cleavage leading to the generation of a 17-kDa neurotoxic fragment, as well as neuronal death, were significantly reduced in E22Q Aβ-treated neurons when compared with WT Aβ-treated ones. This complex regulation of the calpain-mediated toxicity pathway by E22Q Aβ could have some bearing in the pathobiology of this familial AD form.