Effect of inhibitors of secretase activity on Aβ42 levels in our engineered cell lines assayed by High Content analysis.

<p>Cell lines w5c1 and s12c8 cells were stained for nuclei with Hoechst 33342 (blue), and GFP expression (green) was used to identify cells and APP location. Aβ42 expression was identified by tetramethylrhodamineisothiocyanate (TRITC, red). Images were acquired with the ArrayScan HCS reader with a 10x objective. Quantitation was performed with the Cell Health Profiling BioApplication (Cellomics, Pittsburgh, PA, USA). Cells were treated with ibuprofen, donepezil, galantamin, aspirin and chlorpromazine. PBS was used as a control. Data were obtained from three independent experiments, each performed in triplicate, and expressed as mean ± SD. Error bars represent SD. (A) Activity of drugs in cell line w5c1 measured by High Content. (B) Activity of drugs in cell line s12c8 measured by High Content. (C) Quantification of changes in APP and Aβ42 levels between drugs treatment groups. APP was detected in High Content using the anti-human APP Thr668 antibody as primary antibody, CST #2452, and a secondary antibody with TRITC as the reporter (data are not shown).Aβ42 expression in cells is significantly inhibited by ibuprofen, donepezil and galantamin in both the w5c1 and s12c8 cell lines. No significant change in the abundance of APP was seen in either cell lines after drug treatment. Results were obtained from three independent experiments, each performed in triplicate, and expressed as mean ± SD. Error bars represent SD. Asterisk indicates a statistically significant difference between two groups (*p<0.05, **p<0.01, ***p<0.001).