2.6 Cell culture and treatment
The mouse neuroblastoma cells overexpressing the human APP695 Swedish
mutation (K595N/M596L) (N2a-sw cells) was a gift from professor Huaxi Xu
in Xiamen University. The N2a-sw cells (passage 8-15) were used for the
in vitro studies. Cells were cultured in high glucose DMEM (Gibco,
Carlsbad, CA) containing 10% FBS (Gibco, Carlsbad, CA), 100 U/ml
penicillin (Sigma), 100 ug/ml streptomycin (Sigma) at 37 °C in a
humidified atmosphere of 5% CO2. Cells were seeded onto
a six-well plate or slides for 36 h and subsequently starved in FBS free
medium for 12 h. Cells were subsequently treated with Lf (0.05 mg/ml and
0.1 mg/ml) for 24 h, or co-treated with Lf (0.1 mg/ml) and p38 inhibitor
SB203580 (MCE, 2 μM) or PP2A inhibitor calyculin A (CA, MCE, 0.5 nM) for
24 h, or co-treated with p38 inhibitor SB 203580 (MCE, 2 μM) and PP2A
agonist DT-061 (MCE, 1 nM) or for 24 h. Cells were then harvested and
analyzed.