Figure 7. Time-dependent effects of imatinib on PDGF-induced versus insulin-induced PI 3-kinase. Serum-deprived (24 h) RGC-5 cells were pretreated without (control) or with 10 µM imatinib for increasing time intervals (3 h and 24 h). Subsequently, control and imatinib-treated cells were stimulated with either 30 ng/ml PDGF or 30 nM insulin for 6 min. The cell lysates were subjected to immunoprecipitation followed by PI 3-kinase assays or immunoblot analysis (A and B-E) using the indicated primary antibodies (see Methods). The representative thin layer chromatogram and the immunoblots for imatinib regulation of PDGF-induced PI 3-kinase activity (A) and insulin-induced PI 3-kinase activity (B-E) are shown. Note: For data analyses, PDGF- or insulin-induced PI 3-kinase activity in the absence of imatinib was normalized to 100%. The respective bar graphs shown are the mean±SEM values from 3 to 4 experiments. The asterisk indicates a p<0.05 compared with the respective PDGF-induced PI 3-kinase activity.