High sensitivity analysis of amyloid-beta peptide composition in amyloid deposits from human and PS2APP mouse brain

Abstract

Cortical amyloid-beta (Aβ) deposition is considered essential in Alzheimer’s disease (AD) and is also detectable in nondemented individuals with pathologic aging (PA). The present work presents a detailed analysis of the Aβ composition in various plaque types from human AD and PA cases, compared with plaque Aβ isolated from PS2APP mice. To determine minute amounts of Aβ from 30 to 50 laser-dissected amyloid deposits, we used a highly sensitive mass spectrometry procedure after restriction protease lysyl endopeptidase (Lys-C) digestion. This approach allowed the analysis of the amino-terminus and, including a novel ionization modifier, for the first time the carboxy-terminus of Aβ at a detection limit of ∼200 fmol. In addition, full length Aβ 40/42 and pyroglutamate 3-42 were analyzed using a highly sensitive urea-based Western blot procedure. Generally, Aβ fragments were less accessible in human deposits, indicative of more posttranslational modifications. Thioflavine S positive cored plaques in AD were found to contain predominantly Aβ 42, whereas thioflavine S positive compact plaques and vascular amyloid consist mostly of Aβ 40. Diffuse plaques from AD and PA, as well as from PS2APP mice are composed predominantly of Aβ 1-42. Despite biochemical similarities in human and PS2APP mice, immuno-electron microscopy revealed an extensive extracellular matrix associated with Aβ fibrils in AD, specifically in diffuse plaques. Amino-terminal truncations of Aβ, especially pyroglutamate 3-40/42, are more frequently found in human plaques. In cored plaques we measured an increase of N-terminal truncations of ∼20% between Braak stages IV to VI. In contrast, diffuse plaques of AD and PA cases, show consistently only low levels of amino-terminal truncations. Our data support the concept that diffuse plaques represent initial Aβ deposits but indicate a structural difference for Aβ depositions in human AD compared with PS2APP mice already at the stage of diffuse plaque formation.