Figure 4. Akt activation is induced by Rapamycin treatment, and Akt inhibitors abrogate Rapamycin-induced Akt activation.
A-B. 106 cells/ml of the same four DLBCL cell lines (SUDHL-6, WSU-NHL, SUDHL-4 and OCI-Ly19) were treated with Rapamycin, an Akt inhibitor (either Nelfinavir or MK-2206), and the combination of Rapamycin and Akt inhibitor for 3h and 6h. A flow-cytometry-based multiplex protein assay of phosphorylated Akt, as a proportion of total Akt, was performed in SUDHL-6 (Rapamycin-sensitive) cells treated with Rapamycin and Nelfinavir (A) and SU-DHL 4 (Rapamycin-resistant) cells treated with Rapamycin and MK-2206 (B). Each experiment was performed in triplicate, in independent conditions, with mean and SD displayed. Asterisks indicate statistically significant differences (t-test p value <0.05). C-D. One Rapamycin-resistant cell line (SUDHL-4) and one Rapamycin-sensitive cell line (SUDHL-6) were treated for 6 hours with Rapamycin (25 nM), Nelfinavir (15 uM), and the combination; or Rapamycin (25 nM), MK-2206 (300 nM), and the combination; after which cell lysates were prepared and analyzed by Western blot technique. Each experiment was repeated, with representative results provided. Shown here are results from analysis of phosphorylated Akt (p-Akt), phospho-S-6 ribosomal protein (p-S6RP), and phosphorylated 4-EBP-1 (p-4-EBP-1) in SUDHL-6 cells (C), and SUDHL-4 cells (D) treated with Rapamycin (5 nM) and MK-2206 (100 nM) for 6 hours.