title: Pathways regulating lipopolysaccharide-induced neutrophil survival revealed by lentiviral transduction of primary human neutrophils.
abstract: Human neutrophils express Toll-like receptor 4 (TLR4) at low levels, and the role of this receptor in neutrophil responses to microbial stimuli has been questioned. Genetic manipulation of these cells to enable the study of the role of proteins such as TLR4 in their function is challenging. Here, we show that primary human neutrophils rapidly express novel proteins such as enhanced green fluorescent protein (eGFP) after transduction with lentivirus. Stimulation of transduced neutrophils with lipopolysaccharide (LPS) resulted in increased cell survival, which was inhibited when neutrophils were transduced with a lentivirus encoding a dominant negative (dn) TLR4 protein. LPS-induced survival was also inhibited by lentiviruses encoding dnMyD88 or a truncated TRIF (Toll/interleukin-1R homologous domain-containing adapter protein inducing interferon-beta) molecule, whilst, in contrast, neutrophil survival was enhanced by overexpression of kinase-mutated interleukin-1 receptor-associated kinase 1 (kmIRAK-1), which activated nuclear factor (NF)-kappaB. These studies provide proof of the role of TLR4 in human neutrophil biology, have begun to elucidate TLR-dependent pathways regulating neutrophil survival, and demonstrate that neutrophils can be genetically manipulated to enhance or inhibit survival.
authors.affiliation: University of Sheffield, UK.
authors.forename: Emily P
authors.initials: EP
authors.lastname: Dick
authors.forename: Lynne R
authors.initials: LR
authors.lastname: Prince
authors.forename: Elizabeth C
authors.initials: EC
authors.lastname: Prestwich
authors.forename: Stephen A
authors.initials: SA
authors.lastname: Renshaw
authors.forename: Moira K B
authors.initials: MK
authors.lastname: Whyte
authors.forename: Ian
authors.initials: I
authors.lastname: Sabroe
mesh_terms: D002470:Cell Survival / Q000276:immunology; D002478:Cells, Cultured; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D008070:Lipopolysaccharides / Q000276:immunology*; D018375:Neutrophil Activation / Q000276:immunology; D009504:Neutrophils / Q000276:immunology*; D015398:Signal Transduction / Q000276:immunology; D051197:Toll-Like Receptor 4 / Q000276:immunology*; D014161:Transduction, Genetic
chemical_list: D008070:Lipopolysaccharides; C495345:TLR4 protein, human; D051197:Toll-Like Receptor 4; D049452:Green Fluorescent Proteins