Objective To investigate the effects of phosphoinositide 3-kinase (PI3K) inhibitor LY294002 on the activity of PI3K/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway, as well as relevant biological behavioural alteration of gastric cancer MGC-803 cells.
Methods Human gastric cancer MGC-803 cells were treated with LY294002 at 12.5, 25, and 50 μmol/L (experimental groups). The mRNA expressions of key signal molecules in PI3K/AKT/mTOR signaling pathway including PTEN, PI3K, AKT, mTOR, P70S6K, and 4EBP, as well as related protein expressions, were detected by quantitative real-time PCR and Western blot, respectively. The growth-inhibiting effects of LY294002 on MGC-803 cells were detected by MTT assay. The effects on cell cycle and cell apoptosis were detected by flow cytometry.
Results Compared with the control (DMSO) group, LY294002 significantly regulated the mRNA expressions of PI3K, PTEN, AKT, mTOR, 4EBP, and P70S6K (P < 0.05), and the mRNA expression levels of AKT, mTOR, 4EBP, and P70S6K were declined with LY294002 concentration increasing. LY294002 significantly down-regulated the protein expression of AKT, p-mTOR, p-70S6K, and p-AKT, which showed a declining trend with LY294002 concentration increasing. The MTT assay results showed significant inhibiting effects of LY294002 on MGC-803 cells' proliferation as the proliferation rate increased with LY294002 concentration reducing (F=26.15, P < 0.01). The results of flow cytometry showed that the proportions of G0/G1 cells (F=358.594, P < 0.001) and the apoptosis rates (F=4.929, P < 0.05) of the LY294002 groups were higher than those of the control group in a dose-dependent manner.
Conclusion LY294002 may inhibit the key signaling molecules of PI3K/AKT/mTOR signaling pathway in MGC-803 cells to varied degrees. The mRNA expressions of AKT, mTOR, 4EBP, and P70S6K and related phosphorylated proteins show a declining pattern with LY294002 concentration increasing. Besides, LY294002 could decrease cell proliferation rate, change the proportion of cell cycle, and increase apoptosis rate.
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