Figure 6
Luteolin increases the mTORC1 activation. HT-22 cells were pre-treated with luteolin at 5–25 µM for 24 h, followed by 5 mM glutamate for 18 h. (A) The dose-dependent mTORC1 protein expression. The protein expression level of p-mTOR S2448 (mTOR activation) and Raptor (mTORC1 complex) were analyzed by Western blot, and β-actin served as the loading control. (B) Relative protein levels of p-mTOR S2448 (mTOR activation) and Raptor (mTORC1 complex) were quantified by densitometry and the mean data from at least three independent experiments were normalized to the results (n = 3). (C) The time dependent mTORC1 protein expression. The protein expression level of p-mTOR S2448 (mTOR activation) and Raptor (mTORC1 complex) were analyzed at 3 h or 6 h of glutamate induction, and β-actin served as the loading control. (D) Relative protein levels of p-mTOR S2448 (mTOR activation) and Raptor (mTORC1 complex) (n = 3). (E) Representative immunoblotting of mTORC1 downstream target p-S6 (S235/236), p-4E-BP1 (Thr37/46), and p-ULK1 (S757). (F) Relative protein levels of p-mTOR S2448, Raptor, p-S6 (S235/236), p-4E-BP1 (Thr37/46), and p-ULK1 (S757) after rapamycin treatment (n = 3). The Western blot was quantified using NIH Imaging J. The data represent the means ± SEM collected from at least three independent experiments and. *p value < 0.05, **p value < 0.01, ****p value < 0.001 compared with the glutamate treatment group. C:control, Glu:glutamate, L:luteolin, Q:quercetin.
