Bacterial Lipopolysaccharide Induces Release of Tumor Necrosis Factor-α From Bovine Peripheral Blood Monocytes and Alveolar Macrophages In Vitro

Abstract

In this study, we demonstrate that freshly adherent bovine monocytes release tumor necrosis factor-α (TNF-α) in response to stimulation with bacterial lipopolysaccharide (LPS). TNF-α was detected using actinomycin D-treated WEHI-164 murine fibrosarcoma cells as targets in an 18 hr cytotoxicity assay. Doses of LPS from 20 ng/ml to 20 μg/ml were capable of inducing bovine TNF-α. The kinetics of TNF-α release from bovine monocytes demonstrated peak levels of cytotoxic activity at 1–3 hr post-LPS treatment, with a subsequent decline to background levels by 18 hr post-LPS treatment. A monoclonal antibody that neutralizes recombinant human TNF-α (rHuTNF-α) significantly reduced the cytotoxicity of LPS-stimulated bovine monocyte culture supematants. Size exclusion high-performance liquid chromatography (HPLC) analysis of LPS-stimulated monocyte and alveolar macrophage culture supematants resulted in a molecular weight elution profile similar to that of recombinant human TNF-α. These elution profiles are consistent with the presence of multimers of TNF-α. This is believed to be the first report of the in vitro production of bovine TNF-α.