Serum-independent binding of lipopolysaccharide to human monocytes is trypsin sensitive and does not involve CD14.
The nature of the binding sites for lipopolysaccharide (LPS) on human monocytes was investigated using fluorescein isothiocyanate (FITC)-labelled LPS from Salmonella minnesota R595 (ReLPS). In the absence of serum, ReLPS bound to monocytes and this interaction was trypsin sensitive. A concentration of 0.1 mg/ml resulted in a 90% loss of LPS binding, while low concentrations increased this binding. Trypsin-treated monocytes recovered FITC-ReLPS binding after 20 hr culture, which was abrogated in the presence of cycloheximide and actinomycin D. This showed that de novo protein and mRNA synthesis were essential. A number of different proteins have been implicated in cellular binding of LPS to monocytes. In this paper we show that CD14 is not involved in direct binding of FITC-ReLPS to monocytes, since anti-CD14 monoclonal antibody (mAb) (3C10) and removal of most of cell-surface CD14 by phosphatidylinositol-specific phospholipase C did not prevent FITC-ReLPS binding. Furthermore, LPS also bound to CD14-deficient cells from a patient with paroxysmal nocturnal haemoglobinuria (PNH). FITC-ReLPS binding was not mediated by the CD11/CD18 complex since mAb to the alpha and beta chains of the CD11/CD18 complex did not alter the binding of FITC-ReLPS to cells. These observations indicate that ReLPS may interact with monocyte membrane protein(s) in the absence of serum. This binding site(s) for LPS might be different from those previously described by others.
Corrales
I
I
Eijkman-Winkler Laboratory for Medical and Clinical Microbiology, Utrecht University, The Netherlands.
Weersink
A J
AJ
Verhoef
J
J
van Kessel
K P
KP
D015703:Antigens, CD / Q000276:immunology*; D015214:Antigens, Differentiation, Myelomonocytic / Q000276:immunology*; D001665:Binding Sites / Q000276:immunology; D018845:CD11 Antigens; D018821:CD18 Antigens; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D006801:Humans; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides / Q000276:immunology*; D009000:Monocytes / Q000276:immunology*; D012475:Salmonella / Q000276:immunology; D014357:Trypsin / Q000276:immunology*
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D018845:CD11 Antigens; D018821:CD18 Antigens; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D014357:Trypsin; D016650:Fluorescein-5-isothiocyanate