Journal: Journal of Alzheimer's Disease
Article Title: Impact of the Cellular Prion Protein on Amyloid-β and 3PO-Tau Processing
doi: 10.3233/jad-130566
Figure Lengend Snippet: Fig. 2. Effect of PrPC on the expression of APP, BACE1, A40, sAPP, and sAPP. A) PrPC overexpressing HEK293 cells were analyzed 24 and 48 h after transfection by western blotting in comparison to control cells (1). Quantification of the band-intensity showed that PrPC was approximately 2-fold upregulated 24 h- and approximately 3.5 fold upregulated 48 h after transfection (1). Treatment with PNGase resulted in a deglycosylation of PrPC to prove the specificity of the PrPC detection (2). Concentration of PrPC was determined 48 h after transfection using a PrP ELISA-assay (3). B) Expression of APP and BACE1 was analyzed 24 and 48 h after transfection by western blotting. Both proteins were not influenced by the expression of PrPC. C) Amount of APP-cleavage fragments were measured via ELISA. Intra- and extracellular levels of A40, sAPP, and sAPP were decreased significantly in PrPC over-expressing cells (1–3). All in vitro experiments were carried out at least in triplicates. D) In mice, analysis of APP and BACE1 expression revealed no differences between Prnp0/0 and WT mice. E) ELISA measurement of A40 generation in mice brains revealed a marked, age-independent increase in Prnp0/0 mice compared to WT animals (1). PrPC overexpressing Tg35 mice revealed a decreased level of A40 (2) whereas concentration of -secretase remained unchanged in Prnp0/0 mice (3). All values were calculated in % of WT. Beta actin was used as an internal standard. For comparison between groups we used the student’s t-test (n = 5 per group in three independent experiments). Error bars indicate standard errors of the mean (SEM).
Article Snippet: We used a commercial Human -Secretase ELISA Kit (Cusabio, Wuhan, China) to analyze brain homogenates derived from Prnp0/0 and WT mice.
Techniques: Expressing, Transfection, Western Blot, Comparison, Control, Concentration Assay, Enzyme-linked Immunosorbent Assay, In Vitro
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