PI3K activation in InsR silenced cells was dependent on IGF-1R signaling. (A) Cells were incubated with AG538, an IGF-1R specific antagonist, or vehicle (DMSO) for 12 hours, and the cell lysates were subject to in vitro lipid kinase assay. PIP (phosphoinositide 3-phosphate), the phosphorylated end product. Ori, origin of migration in TLC. The bar graph shows the densitometric scanning results (means ± SEM) from four independent experiments. *, P<0.01 vs. SC transfected cells. (B) Cells were treated with vehicle (H2O) or recombinant IGF-1 (rIGF-1, 10ng/mL) for 5 minutes, and the cell lysates were subject to in vitro lipid kinase assay (upper panel). Cell sensitivity to extrinsic IGF-1 is expressed as the ratio of PI3K activity in IGF-1 treated cells to that of vehicle treated cells. Data are means ± SEM from four independent experiments. *, P<0.05 vs. SC transfected cells.