Lps of e coli o111 b4

ELISA assay was used to check the level of inflammatory cytokine. Raw264.7 cells plated at a density of 1 × 10⁵ cells/well were stimulated with 20 ng/mL LPS of E. coli O111:B4 (Sigma-Aldrich, St. Louis, MO, USA) with or without peptide for 16 h. The inhibition effect of peptide on mTNF-α, mIL-6 and mIL-1α expression, which belong to inflammatory cytokines, was measured by enzyme-linked immunosorbent assays (ELISA; R&D Systems, Minneapolis, MN, USA), as previously reported [54 (link)]. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used control in the RT-PCR. Relative intensity of the DNA bands was quantified by ImageJ software (Version 1.52q, National Institutes of Health, Bethesda, MD, USA).

Hypoxia green reagent; pHrodo Green E. coli; DAF-FM DA; JSH-23; QNZ; ABT-263; and ABT-737 were purchased from Thermo Fisher Scientific (Waltham, MA, USA). The fetal bovine serum was from Gibco (Gibco, Sigma Aldrich Company Ltd., Waltham, MA, USA). Intracellular staining permeabilization wash buffer; Human TruStain FcX (Fc Receptor Blocking Solution); PE anti-human Ki-67; isotype control antibodies PE Mouse IgG1 k isotype Ctrl; APC Mouse IgG1 k isotype Ctrl; FITC Mouse IgG1 k isotype Ctrl; PE Mouse IgG2a k isotype; PE Mouse IgG2b k isotype and antibodies APC anti-human CD11b; PE anti-human CD284; PE anti-human CD36; PE anti-human CD33; PE anti-human HLA-DR; PE anti-human HIF-1α were from BioLegend (San Diego, CA, USA). MycoFluor™ Mycoplasma Detection Kit was from Molecular Probes Inc. (Eugene, Oregon, USA). FITC anti-human CD68 was from BD Bioscience (Franklin Lakes, NJ, USA). Culture medium RPMI 1640; 2-mercaptoethanol; PBS; Calcein AM; Bisbenzimide Hoechst 33342 (H33342); propidium iodide (PI), resazurin; LPS of E. coli O111: B4; antibodies FITC anti-human CD11c; FITC anti-human CD14; FITC anti-human CD54; NF-kB activation inhibitor IV, and other chemicals were purchased from Sigma-Aldrich (St. Louis, MO, USA).