title: Response of Neutrophils to Extracellular Haemoglobin and LTA in Human Blood System.
abstract: BACKGROUND
  Haemolytic infection lyses red blood cells, releasing haemoglobin (Hb) into the plasma. Although recent studies showed that immune cells recognize redox-active cytotoxic extracellular Hb (metHb) bound to pathogen-associated molecular patterns (PAMPs), currently available information is limited to experiments performed in defined conditions using single cell lines. Therefore, a systemic approach targeting primary whole blood cells is required to better understand the cellular immune defence against metHb and PAMPs, when under a haemolytic infection.
  
  
  METHODS
  We investigated how human white blood cells, including neutrophils, respond to metHb and lipoteichoic acid (LTA) by measuring reactive oxygen species (ROS), signalling mediators (ERK and p38), NF-κB, cytokines, elastase secretion and cell activation markers.
  
  
  RESULTS
  metHb activates NF-κB in TLR2-expressing HEK293 cells but not in normal or TLR9-expressing HEK293 cells. Treatment of isolated neutrophils with metHb increased production of ROS and expressions of IL-8, TNFα, and CD11b, which were further enhanced by metHb + LTA complex. While LTA stimulated the survival of neutrophils, it caused apoptotic cell death when complexed with metHb. The activation of neutrophils by metHb + LTA was subdued by the presence of other types of white blood cells.
  
  
  CONCLUSIONS
  metHb and metHb + LTA complex are ligands of TLR2, inducing an unconventional TLR signalling pathway. Neutrophils are a highly sensitive cell type to metHb + LTA complex. During a haemolytic infection, white blood cells in the vicinity crosstalk to modulate neutrophil TLR-signalling induced by metHb and LTA.
authors.affiliation: Department of Biological Sciences, NUS Graduate School for Integrative Sciences and Engineering (NGS), National University of Singapore, 14 Science Drive 4, Singapore 117543, Singapore.
authors.forename: Sae-Kyung
authors.initials: SK
authors.lastname: Lee
authors.affiliation: Department of Biological Sciences, NUS Graduate School for Integrative Sciences and Engineering (NGS), National University of Singapore, 14 Science Drive 4, Singapore 117543, Singapore.
authors.forename: Suh Yee
authors.initials: SY
authors.lastname: Goh
authors.affiliation: Department of Biological Sciences, NUS Graduate School for Integrative Sciences and Engineering (NGS), National University of Singapore, 14 Science Drive 4, Singapore 117543, Singapore.
authors.forename: Yuan Qi
authors.initials: YQ
authors.lastname: Wong
authors.affiliation: Department of Biological Sciences, NUS Graduate School for Integrative Sciences and Engineering (NGS), National University of Singapore, 14 Science Drive 4, Singapore 117543, Singapore.
authors.forename: Jeak Ling
authors.initials: JL
authors.lastname: Ding
mesh_terms: D016923:Cell Death / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D016207:Cytokines / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007249:Inflammation / Q000276:immunology / Q000473:pathology; D007962:Leukocytes / Q000187:drug effects / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism / Q000494:pharmacology*; D008706:Methemoglobin / Q000378:metabolism / Q000494:pharmacology*; D016328:NF-kappa B / Q000378:metabolism; D018375:Neutrophil Activation / Q000187:drug effects / Q000502:physiology*; D010196:Pancreatic Elastase / Q000378:metabolism; D000069452:Pathogen-Associated Molecular Pattern Molecules / Q000378:metabolism; D015398:Signal Transduction; D013682:Teichoic Acids / Q000378:metabolism / Q000494:pharmacology*; D051195:Toll-Like Receptor 2 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: D016207:Cytokines; D008070:Lipopolysaccharides; D016328:NF-kappa B; D000069452:Pathogen-Associated Molecular Pattern Molecules; C495342:TLR2 protein, human; D013682:Teichoic Acids; D051195:Toll-Like Receptor 2; C009900:lipoteichoic acid; D008706:Methemoglobin; D010196:Pancreatic Elastase
keywords: Extracellular haemoglobin; Haemolytic infection; Leukocytes; Lipoteichoic acid; Reactive oxygen species; Staphylococcus aureus; Toll like receptor