BCNU decreases Aβ and CTF levels in CHO cells stably expressing APP751WT (7WD10 cells). A) An initial screening of a library of oncology drugs used at 10.0 μM identified carmustine (BCNU) as a potential anti-Aβ drug, while drugs such as 1 and 4 increased Aβ levels. The cells were exposed to the drugs for 48 hours and the conditioned media were immunoprecipitated for Aβ and detected by immunoblots. B) 7WD10 cells were incubated with different concentrations of BCNU for 48 hours and the conditioned media were immunoprecipitated for Aβ and detected by immunoblots. C) Quantitation of Aβ levels by ImageJ revealed decreased Aβ levels to 61% (P < 0.05) at 5 μM, 49% (P < 0.01) at 10 μM and 37% (P < 0.01) at 20 μM compared to controls. D) The lysates were used to detect c-terminal fragments (CTFs) and APP as well as actin as a loading control. E) CTF levels were also significantly decreased to 44% (P < 0.05) at 5 μM, 30% (P < 0.01) at 10 μM and 43% (P < 0.05) at 20 μM compared to controls. F) The conditioned media from B was subjected to immunoblotting to detect different species of sAPPs. G) sAPPα levels were increased to 167% (P < 0.01) at 0.5 uM, 186% (P < 0.05) at 1 uM, 204% (P < 0.01) at 5 uM and 152% (P < 0.05) at 10 uM compared to untreated cells. H) sAPPtotal levels were increased to 159% (P < 0.05) at 5 μM, 172% (P < 0.01) at 10 μM and 170% (P < 0.01) at 20 μM in BCNU treated cells compared to controls. For all samples, n = 4 ± SEM. *, P < 0.05, **, P < 0.01 versus control by analysis of variance (ANOVA) followed by post-hoc test by Dunnett multiple comparisons. Aβ, amyloid β; APP, Alzheimer's precursor protein; BCNU, 1, 3 bis (2-chloroethyl)-1-nitrosourea; CHO, Chinese hamster ovary; SEM, standard error of the mean.
Hayes et al. BMC Medicine 2013 11:81 doi:10.1186/1741-7015-11-81