title: Neither CD14 nor serum is absolutely necessary for activation of mononuclear phagocytes by bacterial lipopolysaccharide.
abstract: The stimulation of mononuclear phagocytes by lipopolysaccharide (LPS) is facilitated by the binding of complexes of LPS and LPS-binding protein to CD14. Although it is clear that CD14 is involved in LPS-induced signaling, other investigators have hypothesized the existence of additional signaling pathways in macrophages. We sought to determine whether CD14-independent pathways of monocyte activation might exist. Washed human mononuclear cells responded with reduced sensitivity to LPS in the absence of serum. Anti-CD14 monoclonal antibody (MAb) inhibited the response to LPS in serum-free conditions, but this was easily reversed at higher concentrations of LPS. We established a human monocytic cell line, designated SFM (derived from THP-1), in serum-free medium to examine LPS responses under defined conditions. Differentiation of SFM cells with 1,25-dihydroxycholecalciferol promoted the expression of abundant cell surface CD14. Differentiated SFM cells responded to LPS despite the complete absence of serum proteins for > 20 generations of growth. LPS stimulation of differentiated SFM cells was inhibited by anti-CD14 MAbs only when serum was present. In contrast to anti-CD14 MAb, the LPS antagonists lipid IVa and Rhodobacter sphaeroides lipid A inhibited monocyte activation under serum-free conditions, implying that these compounds compete with LPS at a site distinct from CD14. Undifferentiated SFM cells (expressing minimal CD14) still responded to LPS in serum-free conditions, and anti-CD14 MAb had little inhibitory effect. The addition of purified LPS-binding protein or human serum promoted a CD14-dependent pathway of monocyte activation by LPS in these cells. We conclude that monocytes do not absolutely require serum proteins to be stimulated by LPS and that CD14-independent LPS signaling pathways exist which are inhibitable by lipid IVa and R. sphaeroides lipid A.
authors.affiliation: Department of Internal Medicine, Maxwell Finland Laboratory for Infectious Diseases, Boston City Hospital, Boston University School of Medicine, Massachusetts 02118.
authors.forename: W A
authors.initials: WA
authors.lastname: Lynn
authors.forename: Y
authors.initials: Y
authors.lastname: Liu
authors.forename: D T
authors.initials: DT
authors.lastname: Golenbock
mesh_terms: D000209:Acute-Phase Proteins; D015703:Antigens, CD / Q000502:physiology*; D015214:Antigens, Differentiation, Myelomonocytic / Q000502:physiology*; D001798:Blood Proteins / Q000502:physiology; D002117:Calcitriol / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D003470:Culture Media; D006801:Humans; D066298:In Vitro Techniques; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D008262:Macrophage Activation / Q000187:drug effects; D008562:Membrane Glycoproteins; D009000:Monocytes / Q000502:physiology*; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D000099231:Lipopolysaccharide-Binding Protein
chemical_list: D000209:Acute-Phase Proteins; D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D001798:Blood Proteins; D002117:Calcitriol; D002352:Carrier Proteins; D003470:Culture Media; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D013755:Tetradecanoylphorbol Acetate; D014409:Tumor Necrosis Factor-alpha; D000099231:Lipopolysaccharide-Binding Protein