Fig. 4

Increased p-AMPKα contributed to enhanced autophagy in TSC1 KO BMMϕ. TSC1 WT and KO peritoneal macrophages (a) or BMMϕ (b) were left uninfected or infected with BCG at a MOI of 10 for 24 h and Western blot analysis was performed against the indicated antibodies. c Quantification of AMP level was performed by LC-MC analysis in TSC1 WT and KO BMMϕ. d Compound C was added into TSC1 KO BMMϕ at the concentration of 0, 2.5, or 25 μM for 2 h. Cells were subjected to Western blot analysis. e TSC1 KO BMMϕ were treated with rapamycin at 0.1, 10, 100 ng/ml overnight. f Amino acids were added to TSC1 KO BMMϕ for 0, 30, 60 and 120 min. g, h, and i TSC1 KO BMMϕ were left uninfected or infected with BCG or BCG plus amino acids for 4 h and then lysed with Trizol reagent for qRT-PCR. IL-1β (g), IL-1α (h), and TNFα (i) mRNA levels in TSC1 KO BMMϕ are shown. Each experiment was repeated at least twice. Densitometric quantification of LC3-II (n = 3) was normalized to total Actin. The arrow indicates LC3B-II. AA, amino acids; BCG, Bacillus Calmette-Guérin; Comp C, compound C; KO, knockout; Rapa, rapamycin; WT, Wild type. *, p < 0.05; **, p < 0.01