Lipopolysaccharide modulation of a CD14-like molecule on porcine alveolar macrophages.
Cluster of differentiation antigen 14 (CD14) functions as a receptor for lipopolysaccharide (LPS) LPS-binding protein (LBP) complexes. Because LPS has varying effects on CD14 expression in vitro, we evaluated CD14 expression in response to LPS with a fully differentiated macrophage phenotype, the alveolar macrophage. By using flow microfluorometric analysis and a radioimmunoassay with an anti-human CD14 monoclonal antibody (My4) that cross-reacts with porcine CD14, we found that macrophages stimulated with LPS for 24 h exhibited a two- to fivefold increase in CD14-like antigen compared with unstimulated cells. At low concentrations of LPS, up-regulation of the CD14-like antigen was dependent on serum; at higher concentrations of LPS, serum was not required. In the absence of serum a 10-fold higher dose of LPS (10 ng/ml) was required to increase CD14-like expression. In addition, LPS-induced CD14-like up-regulation correlated with secretion of tumor necrosis factor-alpha, regardless of serum concentration. Blockade with My4 antibody significantly inhibited LPS-induced tumor necrosis factor-alpha secretion at 1 ng/ml of LPS. However, inhibition decreased as we increased the LPS concentration, suggesting the existence of CD14-independent pathways of macrophage activation in response to LPS. Alternatively, My4 may have a lower affinity for the porcine CD14 antigen than LPS, which may have only partially blocked the LPS-LBP binding site at high concentrations of LPS. Therefore, these data suggest that LPS activation of porcine alveolar macrophages for 24 h increased CD14-like receptor expression. The degree of CD14-like up-regulation was related to LPS concentration, however, activation did not require the presence of serum at high concentrations of LPS.
Kielian
T L
TL
Department of Anatomy, Kansas State University, Manhattan 66506-5602, USA.
Ross
C R
CR
McVey
D S
DS
Chapes
S K
SK
Blecha
F
F
D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D015703:Antigens, CD / Q000378:metabolism / Q000502:physiology*; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism / Q000502:physiology*; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D007700:Kinetics; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides / Q000494:pharmacology*; D016676:Macrophages, Alveolar / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology*; D011869:Radioligand Assay; D013268:Stimulation, Chemical; D013552:Swine; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D016895:Culture Media, Serum-Free; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha