Rpmi 1640 medium without glucose

After 24 h in the presence of the unloaded and antibacterial loaded dressing materials, the glucose uptake by macrophages was evaluated. After washing the wells of the 24-well plates with PBS, RPMI 1640 medium without glucose (Gibco^TM) supplemented with 100 μM of fluorescent D-glucose analog 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose (2-NBDG) (Cayman Chemical, Biomol GmbH, Germany) was added. The plates were incubated at 37°C for 60 min in the dark. The supernatant was removed and the wells were washed once with PBS. Accutase (PAN Biotech) was added to induce the detachment of the cells and the content of the wells was transferred to a black 96-well plate to read fluorescence (excitation 465/emission 540 nm) in a Tecan microplate reader Infinite^® 200 PRO.

For in vitro differentiation, naïve CD8⁺ T cells (1.5 x 10⁶ cells/mL and 1 x 10⁶ cells/mL) were distributed into four wells of a 24-well plate and activated for 8 days by adding prewashed anti-CD3/CD28 Dynabeads at a 1:1 bead-to-cell ratio (Invitrogen/Thermo Fisher Scientific, Massachusetts, USA) and human rIL-2 (30 U/mL) (Roche/Merck, Darmstadt, Germany) to the culture. To generate effector memory T cells, 1 x 10⁶ cells/mL were restimulated on day 7 by adding prewashed anti-CD3/CD28 Dynabeads at a 1:1 bead-to-cell ratio and human rIL-2 (30 U/mL) a second time to the culture. The cells were cultured in RPMI 1640 medium without glucose (Gibco/Life Technologies, Carlsbad, USA) where 11 mM D-glucose was added to 10% FBS, 2 mM L-glutamine and 1% penicillin-streptomycin (all from Sigma Aldrich, St.Louis, USA). 24 h before collection, culture medium was replaced with RPMI 1640 medium containing 11 mM of ¹³C labeled-glucose (Cambridge Isotope Laboratories, Massachusetts, USA). At indicated time points, naïve T cells (T_N, day 1), stem cell memory T cells (T_SCM, day 2), central memory T cells (T_CM, day 5) and effector memory T cells (T_EM, day 8) were collected and anti-CD3/CD28 beads were removed by using a DynaMag-2 magnet (Thermo Fisher Scientific, Massachusetts, USA) before further processing for flow cytometry, RNA sequencing and metabolomics analyses.

THP-1 was purchased from Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences (Shanghai, China). Panax notoginseng saponins were purchased from Kunming Pharmaceutical Company (KPC) (Yunnan Province, China), which mainly contained notoginsenoside R1 9%, ginsenoside Rg1 32%, ginsenoside Re 5%, ginsenoside Rb1 36%, and ginsenoside Rd 8%. The chemical purity of PNS was about 90%. RPMI-1640 medium without glucose was bought from Gibco (Carlsbad, CA). PMA, D-glucose, and NF-κB inhibitor of BAY 11-7082 were from Sigma-Aldrich (St. Louis, MO, USA). M-MLV Reverse Transcriptase was from GeneCopoeia (Maryland, USA). SYBP Premix Ex Taq TM was from Takara (Shiga, Japan). TRIzol, RIPA, and BCA protein assay kit were from Thermo Fisher Scientific (Waltham, USA). Real-time PCR detection system was from Bio-Rad (Hercules, CA, USA). Flow cytometry was purchased from BD Biosciences (CA, USA). Rabbit polyclonal p50 and p65 antibodies were from Proteintech (Chicago, USA). Mouse monoclonal antibodies to CD16 and CD206 and rabbit monoclonal antibodies to phospho-IκB alpha and arginase 1 were from Abcam (Cambridge, UK). Mouse monoclonal antibody to β-actin and rabbit polyclonal antibody to β-tubulin were purchased from Boster Biological Technology Co., Ltd. (Wuhan, China) and Cell Signaling Technology (Danvers, MA). ECL kit was purchased from Pierce Biotechnology (Waltham, USA).

Cell culture reagents (RPMI-1640 medium without glucose and fetal bovine serum) were purchased from Gibco, California, USA. The [9,10-³H]-palmitic acid, [³H]-H₂O, 2-[1,2-³H]-deoxy-D-glucose, [1-¹⁴C]-mannitol, and liquid scintillation counting cocktail were from Perkin Elmer, Massachusetts, USA. Hydrocortisone sodium phosphate, as GC treatment used in this study, was purchased from Nycomed Pharma (Zurich, Switzerland). Lipoprotein lipase (LPL) was purchased from Sigma-Aldrich (St. Louis, USA). MTT for the colorimetric assay for the nonradioactive quantification of cellular metabolic activity was purchased from Roche (Mannheim, Germany). Apoptosis in the placental explants was analyzed by the Dead-End TM Fluorometric TUNEL System Kit (Promega, Madison, WI, USA) and the Caspase-GLO 3/7 Assay Kit (Promega, Madison, USA).