Differential toxicity, conformation and morphology of typical initial aggregation states of Aβ1-42 and Aβpy3-42 beta-amyloids

Description

Among the different species of water-soluble beta-peptides (Abeta1-42, Abeta1-40 and N-terminal truncated Abeta-peptides), Abetapy3-42 is thought to play a relevant role in Alzheimer's pathogenesis due to its abundance, resistance to proteolysis, fast aggregation kinetics, dynamic structure and high neurotoxicity. To evaluate the specific structural characteristics and neurotoxicity of Abetapy3-42, we separated different aggregation states of Abeta1-42 and Abetapy3-42 using fast protein liquid chromatography, isolating in both cases three peaks that corresponded to sa (small), ma (medium) and la (large) aggregates. Conformational analysis, by circular dichroism showed a prevailing random coil conformation for sa and ma, and typical beta-sheet conformation for la. AFM and TEM show differential structural features between the three aggregates of a given beta-peptide and among the aggregate of the two beta-peptides. The potential toxic effects of the different aggregates were evaluated using human neuroblastoma SH-SY5Y cells in the MTT reduction experiments, using the xCELLigence System, and the annexin V binding to measure cell apoptosis. In the case of Abeta1-42 the most toxic aggregate is la, while in the case of Abetapy3-42 both sa and la are equally toxic. Abeta aggregates were found to be internalized in the cells, as estimated by confocal immunofluorescence microscopy, with a higher effect observed for Abetapy3-42, showing a good correlation with the toxic effects. Together these experiments allowed the discrimination of the intermediate states more responsible of oligomer toxicity, providing new insights on the correlation between the aggregation process and the toxicity and confirming the peculiar role in the pathogenesis of Alzheimer disease of Abetapy3-42 peptide.

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