SERUM FACTORS, CELL-MEMBRANE CD14, AND BETA(2) INTEGRINS ARE NOT REQUIRED FOR ACTIVATION OF BOVINE MACROPHAGES BY LIPOPOLYSACCHARIDE

The role of serum factors such as lipopolysaccharide (LPS)-binding pro tein (LBP) and of macrophage-expressed CD14 and beta(2) integrins in t he activation of bovine macrophages by LPS was investigated, Macrophag e activation was determined by measuring tumor necrosis factor product ion, NO generation, and upregulation of procoagulant activity by LPS ( Escherichia coli O55:B5) at concentrations of 100 pg/ml to 100 ng/ml, The 50% effective dose for LPS was 1 order of magnitude higher than th at for activating human macrophages, Macrophages were activated by LPS in the presence of serum or in the presence of albumin demonstrated t o be free of LBP. The capacity to react to LPS in the absence of LBP w as not due to the acquisition of LBP during a previous culture in seru m, It was then established which CD14-specific antibodies block LPS bi nding to monocytes. Among the CD14-specific antibodies recognizing bov ine mononuclear phagocytes (60bca, 3C10, My4, CAM36, VPM65, CMRF31, an d TUK4), the first four blocked the binding of LPS-fluorescein isothio cyanate to bovine monocytes at low concentrations. Anti-CD14 antibodie s did not block LPS-mediated activation of bovine bone marrow-derived macrophages, monocyte-derived macrophages, and alveolar macrophages, T his was observed in experiments in which anti-CD14 concentrations exce eded the 50% inhibitory dose by >30-fold (3C10 and My4) or >300-fold ( 60bca), as defined in the binding assay described above, Monocyte-deri ved macrophages from an animal deficient in beta(2) integrins and cont rol macrophages were activated by similar concentrations of LPS, sugge sting that beta(2) integrins are not important bovine LPS receptors, T hus, in bovine macrophages, LPS recognition pathways which are indepen dent of exogenous LBP, of membrane-expressed CD14, and of beta(2) inte grins may exist.