title: CD14 and TLR4 mediate cytokine release promoted by electronegative LDL in monocytes.
abstract: OBJECTIVE
  Electronegative LDL (LDL(-)), a minor modified LDL present in the circulation, induces cytokine release in monocytes. We aimed to determine the role of the receptor CD14 and toll-like receptors 2 and 4 (TLR2, TLR4) in the inflammatory action promoted by LDL(-) in human monocytes.
  
  
  RESULTS
  Monocytes were preincubated with antibodies to neutralize CD14, TLR2 and TLR4. The release of monocyte chemoattractant protein 1 (MCP1), and interleukin 6 and 10 (IL6 and IL10) promoted by LDL(-) was inhibited 70-80% by antiCD14 and antiTLR4, and 15-25% by antiTLR2. The involvement of CD14 and TLR4 was confirmed by gene silencing experiments. The human monocytic THP1 cell line overexpressing CD14 released more cytokines in response to LDL(-) than the same THP1 cell line without expressing CD14. VIPER, a specific inhibitor of the TLR4 signaling pathway, blocked 75-90% the cytokine release promoted by LDL(-). Cell binding experiments showed that monocytes preincubated with neutralizing antibodies presented lesser LDL(-) binding than non-preincubated monocytes The inhibitory capacity was antiCD14>antiTLR4>>antiTLR2. Cell-free experiments performed in CD14-coated microtiter wells confirmed that CD14 was involved in LDL(-) binding. When LDL(-) and lipopolysaccharide (LPS) were added simultaneously to monocytes, cytokine release was similar to that promoted by LDL(-) alone. Binding experiments showed that LDL(-) and LPS competed for binding to monocytes and to CD14 coated-wells.
  
  
  CONCLUSIONS
  CD14 and TLR4 mediate cytokine release induced by LDL(-) in human monocytes. The cross-competition between LPS and LDL(-) for the same receptors could be a counteracting action of LDL(-) in inflammatory situations.
authors.affiliation: Instituto de Investigaciones Biomédicas Sant Pau (IIB SantPau), Barcelona, Spain. mestruch@santpau.cat
authors.forename: Montserrat
authors.initials: M
authors.lastname: Estruch
authors.forename: Cristina
authors.initials: C
authors.lastname: Bancells
authors.forename: Lorea
authors.initials: L
authors.lastname: Beloki
authors.forename: Jose Luis
authors.initials: JL
authors.lastname: Sanchez-Quesada
authors.forename: Jordi
authors.initials: J
authors.lastname: Ordóñez-Llanos
authors.forename: Sonia
authors.initials: S
authors.lastname: Benitez
mesh_terms: D057134:Antibodies, Neutralizing / Q000494:pharmacology; D001667:Binding, Competitive / Q000187:drug effects / Q000276:immunology; D016207:Cytokines / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D018950:Lipopolysaccharide Receptors / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology; D008077:Lipoproteins, LDL / Q000276:immunology / Q000378:metabolism* / Q000494:pharmacology; D009000:Monocytes / Q000276:immunology / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects / Q000276:immunology; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism
chemical_list: D057134:Antibodies, Neutralizing; D016207:Cytokines; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008077:Lipoproteins, LDL; D016328:NF-kappa B; D034741:RNA, Small Interfering; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D018808:Transcription Factor AP-1; C099252:oxidized low density lipoprotein
keywords: CD14; Electronegative LDL; Lipopolysaccharide; Monocytes; Toll-like receptors