Brewer modified thioglycollate medium

C57BL/6 mice were bred and maintained in specific pathogen–free facilities at the University of Edinburgh. Experimental mice were age and sex matched.
Heligmosomoides polygyrus bakeri life cycle was maintained in house and infective third-stage larvae (L3) were obtained as described elsewhere [51 (link)]. Mice were infected with 200 H. polygyrus L3 by oral gavage. Fecal egg burden was determined on day 13 of the infection using a McMaster counting chamber (Hawksley).
The attenuated, aroA deficient Salmonella enterica serovar Typhimurium strain SL3261 [52 (link)] was cultured as stationary overnight culture from frozen stock in Luria-Bertani broth. Unless indicated otherwise animals were injected i.p. with ~3-5x10^4 CFU diluted in PBS. Infectious doses and splenic bacterial burdens were enumerated by plating inocula or tissue homogenates in 10-fold serial dilutions in PBS on LB-Agar plates.
IL-4–anti–IL-4 mAb complex (IL-4c) was prepared as described previously [53 (link)], and mice were injected i.p. with 5 μg of recombinant IL-4 (13.5 kD; PeproTech) complexed to 25 μg 11B11 (Bio X Cell) or 100 μL PBS vehicle control on days 0 and 2, and peritoneal exudate cells were harvested on day 4.
For in vitro experiments mice were injected with 400 μL 4% Brewer modified thioglycollate medium (BD Biosciences) three days prior to necropsy.

BM macrophages were extracted and cultured as previously described [7 (link)] with minor modification. Mouse tibias were collected and the BM was flushed with Dulbecco’s modified Eagle’s medium (DMEM; Gibco, USA) and differentiated into primary BMDMs. Cells were cultured in DMEM containing 10% fetal bovine serum (FBS; Hyclone, USA) and incubated at 37° C in a humidified atmosphere containing 5% CO₂. Macrophages were stimulated with IFN-γ and IL-4 to induce M1 and M2 macrophage polarization, respectively [30 (link)].
The mouse peritoneal macrophages were collected and cultured as previously described [49 (link)] with minor modification. The mouse were injected intraperitoneally with 2 ml of brewer modified thioglycollate medium (BD, USA). Three days later, cells were harvested by peritoneal lavage with 5 ml RPMI-1640 medium (Gibco, USA). Then cells were washed with cold RPMI-1640 medium and seeded on plastic plates. After two hours incubation, the non-adherent cells were washed with RPMI-1640 medium and the adherent cells were monolayer macrophages.

LPS (E. coli 0111:B4), dimethyl sulfoxide (DMSO), Tween-80, Hoechst 33342 and amino acid transporter inhibitor BCH (2-amino-2-norbornanecarboxylic acid) were bought from Sigma-Aldrich (St. Louis, MO, USA). Thioglycollate medium (Brewer modified) was obtained from Becton Dickinson (Sparks, MD, USA). Piperine was purchased from Guangzhou Institute for Drug Control (Guangzhou, China), dissolved in DMSO and stored at −20°C. Rabbit antibodies against phospho(p)-p70S6K, p70S6K, p-4E-BP1, 4E-BP1, p-S6(Ser235/236), GATA6, cleaved caspase-3, SLC7A5, SLC3A2, mTOR, and β-tubulin were purchased from Cell Signaling Technology (Danvers, MA, USA). The mouse antibody against LAMP2 was obtained from Abcam (Cambridge, MA, USA). PE-F4/80, FITC-CD11b, AlexaFluor488-CD11b, and APC-MHCII were obtained from eBioscience (San Diego, CA, USA). DMEM medium, fetal bovine serum (FBS), penicillin and streptomycin were products of Invitrogen (Carlsbad, CA, USA).
Female C57BL/6 mice were bought from the Experimental Animal Center of Southern Medical University (Guangzhou, China). Animal experiments were designed following National Institutes of Health guidelines and were approved by the Committee on the Ethics of Animal Experiments of Jinan University.