Stimulating NMuMG Cells with LPS and LTA

NMuMG cells were seeded at a density of 4×10⁵ cells/well in 6-well collagen-coated plates (#NCO3506, Corning, Corning, NY, USA) and incubated for 15 ± 1 h before experiments. After incubation, the cells were rinsed with sterile phosphate-buffered saline (PBS) and treated with LPS from Escherichia coli O111:B4 (#115K4029, Sigma-Aldrich, St. Louis, MO, USA) or lipoteichoic acid (LTA) from Bacillus subtilis (L3265, Sigma-Aldrich), which are outer membrane proteins of the Gram-negative and Gram-positive bacteria, respectively. Because our previous studies indicated significantly different responses in cultured mouse colonic, type-II alveolar, and pulmonary epithelial cells [16 (link),17 (link),18 (link)], LPS and LTA were diluted to 10 µg/mL in DMEM without FBS. NMuMG cells treated with LPS or LTA were incubated for 2 h at 37 °C in a 5% CO₂ atmosphere.