Intracellular Aβ is increased by okadaic acid exposure in transfected neuronal and non-neuronal cell lines

Intracellular Aβ was examined in both a neuronal cell line (B103) expressing human APP with Swedish mutation and a non-neuronal cell line (Chinese hamster ovary, CHO) expressing wild human APP. Exposure of the APP695sw-transfected B103 cells to okadaic acid for 3 h, Aβ immunostaining was enhanced, as demonstrated by two independent anti-Aβ antibodies. The confocal microscopic study revealed that the immunoreactivity of Aβ was mainly colocalized with a Golgi marker and partially with an ER marker. Quantitative analyses, using Aβ sandwich ELISA, showed significantly increased intracellular Aβ. False positive detection of Aβ by antibody cross-reaction with APP was ruled out by extracting the fraction with formic acid and making it alkaline before subjecting it to ELISA. This procedure resulted in a fraction that contained little APP. Using CHO cells, OA treatment was also shown to be effective in increasing Aβ, as demonstrated by Western blot. The increased full-length APP and decreased APPC99 were also observed. This is the first study to demonstrate that OA treatment significantly increases intracellular Aβ.