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nan12032-sup-0001-fs1.tif9175K

Figure S1. DAB images of ageing associated dystrophic neurites. Coronal sections of the hippocampus from 15-month-old mice were immunostained with the labelled antibodies, which shows the same pattern as the fluorescence images. Arrowheads indicate clusters of dystrophic neurites. High-power images of clusters are shown next to the low-power images. Scale bar = 50 μm (Low power). Scale bar = 5 μm (High power).

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Figure S2. (A) High-power confocal images of clusters of dystrophic neurites. Coronal sections of the hippocampus from 15-month-old mice were immunostained with the labelled antibodies and confocal images were taken. Scale bar = 5 μm. (B) Coronal sections of the hippocampus from 15-month-old mice were immunostained with the APP-NT antibody. Higher-power image of clusters in the square box is shown in the lower panel. Scale bar = 5 μm. (C) Transmission electron microscopy images of clusters of dystrophic neurites. High magnification image of a dystrophic neurite (arrowhead) filled with translucent or amorphous electrodense materials. Scale bar = 2 μm (upper panel). Scale bar = 500 nm (lower panel).

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Figure S3. Accumulation of cathepsin-D and γ-tubulin in the dystrophic neurites of lactacystin-treated neurones. Treatment of primary cortical neurones with the proteasome inhibitor lactacystin (10 μM) for 24 h induced accumulation of cathepsin-D and γ-tubulin (arrowheads) in dystrophic neurites, indicating the accumulation of lysosomes and aggresomes. Scale bar = 20 μm.

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Figure S4. Accumulation of α-secretase-cleaved APP peptides in the dystrophic neurites of lactacystin-treated neurones. (A) Immunofluorescence of various APP epitopes in control neurones. (B) After treating with 10 μM lactacystin for 24 h, dystrophic neurites that accumulated cathepsin-D were also immunopositive for 6E10 staining, which detects Aβ1–16 (arrowheads); however, immunostaining for the 4G8 antibody, which detects Aβ17–24, was less intense in dystrophic neurites and was not colocalized with cathepsin-D (arrows). APP-NT immunostaining was prominent in dystrophic neurites, but APP-CT695 immunostaining was light and did not colocalize with APP-NT (arrowheads), indicating the predominant accumulation of α-secretase-cleaved APP peptides in the dystrophic neurites of lactacystin-treated neurones. Scale bar = 20 μm.

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