Figure 3
sPLA 2 -IIA-induced proliferation in microglial cells is dependent on epidermal growth factor receptor (EGFR) transactivation. (A, B) Primary and immortalized BV-2 microglial cells were stimulated with 1 μg/ml of sPLA2-IIA in the absence or presence of AG1478 at 37°C. After 24 h, cell proliferation was investigated and expressed as optical density values ± SD. Values are the average of three separate experiments in quadruplicate (*P < 0.001 vs. control cells, **P < 0.001 vs. sPLA2-IIA-treated cells). (C) After 15 minutes, ERK 1/2, P70S6 and rS6 phosphorylation were identified in the cell lysates of BV-2 microglial cells by western blot. Membranes were stained with Ponceau S as a loading control. (D) BV-2 cells were stimulated with 1 μg/ml of sPLA2-IIA in the absence or presence of 2.5 μM of AG1478 for 15 minutes. Flow cytometry analysis of phospho-EGFR levels in untreated (open black lines), sPLA2-IIA-treated (open dark grey lines) and AG1478 + sPLA2-IIA-treated (open blue lines) cells. Solid light grey curves show negative controls in which the primary antibody was omitted. Results shown are representative of three independent experiments.