Figure 2

Both mTOR and p70S6 kinase suppress OPN-induced NF-κB and AP-1 mediated ICAM-1 expression. A. MCF-7 cells were transfected with wild type or rapamycin resistant mTOR or pretreated with 20 nM rapamycin for 1 h and then treated with 0.5 μM OPN for 24 h. B. In another experiments, MCF-7 cells were transfected with wild type or rapamycin resistant p70S6 kinase or pretreated with 20 nM rapamycin and then treated with OPN under the same condition as mentioned above. Cell lysates were analyzed by western blot using anti-ICAM-1 antibody. Actin was used as loading control. C.MCF-7 cells were transfected with ICAM-1 luciferase reporter construct along with Renilla luciferase and then treated with 0.5 μM OPN for 24 h or pretreated with rapamycin (20 nM) for 1 h and then treated with OPN. The transfection efficiency was normalized and fold changes in luciferase activity with respect to control were calculated, and mean ± S.D. of triplicate determinations are plotted. The values were also analyzed by One Way ANOVA (*, p > 0.05; **, p > 0.05). D. MCF-7 cells were individually transfected with IκBα sup. rep., wild type and dominant negative c-Jun or A-Fos and then treated with 0.5 μM OPN for 24 h. Cell lysates were analyzed by western blot using anti-ICAM-1 antibody. Actin was used as loading control.