Activation of the PI3K/mTOR Pathway Is Involved in Cystic Proliferation of Cholangiocytes of the PCK Rat
Figure 2
In vitro effects of PI3K and mTOR inhibitors on cholangiocytes.
Normal and PCK cholangiocytes were treated with the PI3K and/or mTOR inhibitors, and the cell proliferative activity was determined using the WST-1 assay as described in the Materials and Methods. Rapamycin, everolimus, NVP-BEZ235 and LY294002 significantly inhibited the cell proliferative activity of normal (A) and PCK (B) cholangiocytes in a dose-dependent fashion, where the most prominent inhibitory effect was observed in both cholangiocytes treated with NVP-BEZ235 for 120 hours. Rictor siRNA significantly reduced the cell proliferative activity of the PCK cholangiocytes when it was used in combination with rapamycin and LY 294002 (C). Western blot analysis showed that the expression of p-Akt (Se473) and p-S6 was reduced following 24 hour-treatment with rapamycin, everolimus and NVP-BEZ235 in a dose-dependent fashion in PCK cholangiocytes (D). The results of the semiquantitative analysis of Western blotting are shown in E. Note that the expression of p-Akt (Ser473) and p-4E-BP1 was markedly decreased in PCK cholangiocytes following the treatment with 500 nM NVP-BEZ235 (D and E). *, p<0.01; **, p<0.05 (vs. untreated control).