Figure 1.
Pore formation by Aβ1–40 and Aβ1–42 in planar lipid bilayers.
A) Cartoon of the experimental setup. B) Example of transmembrane ion flux induced by 15 µM Aβ1–40 prepared by one day incubation in water (method A, Table S1). C) Example of transmembrane current induced by 15 µM Aβ1–42 prepared by three day incubation (method A). Addition of 10 mM Zn2+ (arrows) inhibited Aβ-induced ion flux.
Figure 2.
Pore formation, cytotoxicity, ThT fluorescence and β-sheet content as a function of aggregation time of Aβ samples in water (method A).
Red and blue curves in each panel are the two component predictions from PLS regression for Aβ1–40 or Aβ1–42, respectively. A) Percentage of experiments that showed pore formation in planar lipid bilayers. Each point represents 10–15 experiments in the presence of 15–25 µM Aβ1–40 or Aβ1–42; error bars represent the error of proportion. B) Cell death of human neuroblastoma SH-SY5Y cells 24 h after exposure to serum-free media containing 20 µM Aβ prepared by method A. Each point represents 5–15 independent experiments. C) Degree of fibril formation as determined by ThT fluorescence. Each point represents an average from 5–20 experiments, error bars represent standard errors of the mean. D) Beta-sheet formation as determined by CD spectroscopy. Each point represents an average from 3–5 experiments, error bars represent standard errors of the mean.
Figure 3.
Separation of Aβ samples by SDS-PAGE followed by Western blot and densitometry analysis.
A) Example of a Western blot of after SDS-PAGE of Aβ1–40 (left) and Aβ1–42 (right) preparations that had been incubated in water for 0, 1, 2, 3, 10, and 20 days (method A). All samples were cross-linked with 12 mM glutaraldehyde before electrophoresis through a 16.5% Tris-Tricine gel. Grouping of oligomers is indicated on the right. B) Average relative abundance of aggregated Aβ1–40 (red) and Aβ1–42 (blue) species of different size as a function of aggregation time. Each point represents the mean value of the relative abundance of each species from 4 to 6 gels; error bars represent the standard error of the mean. Red and blue curves are best curve fits of equation S1 to the data (see Supporting Information).
Figure 4.
Dependence of pore formation, cytotoxicity, β-sheet content and ThT fluorescence on the oligomer levels of Aβ1–40 (top) and Aβ1–42 (bottom).
The sign of the regression coefficients indicates whether a particular species contributes positively or negatively to a particular observation. The error bars are the standard deviation of the coefficient estimated through a jackknife procedure [90]. Coefficients less than 0.25 were considered insignificant and values that were at least one or two standard deviations from zero are marked as significant (*) or highly significant (**) respectively.