title: Opsonization of malaria-infected erythrocytes activates the inflammasome and enhances inflammatory cytokine secretion by human macrophages.
abstract: BACKGROUND
  Antibody opsonization of Plasmodium falciparum-infected erythrocytes (IE) plays a crucial role in anti-malarial immunity by promoting clearance of blood-stage infection by monocytes and macrophages. The effects of phagocytosis of opsonized IE on macrophage pro-inflammatory cytokine responses are poorly understood.
  
  
  METHODS
  Phagocytic clearance, cytokine response and intracellular signalling were measured using IFN-γ-primed human monocyte-derived macrophages (MDM) incubated with opsonized and unopsonized trophozoite-stage CS2 IE, a chondroitin sulphate-binding malaria strain. Cytokine secretion was measured by bead array or ELISA, mRNA using quantitative PCR, and activation of NF-κB by Western blot and electrophoretic mobility shift assay. Data were analysed using the Mann-Whitney U test or the Wilcoxon signed rank test as appropriate.
  
  
  RESULTS
  Unopsonized CS2 IE were not phagocytosed whereas IE opsonized with pooled patient immune serum (PPS) were (Phagocytic index (PI)=18.4, [SE 0.38] n=3). Unopsonized and opsonized IE induced expression of TNF, IL-1β and IL-6 mRNA by MDM and activated NF-κB to a similar extent. Unopsonized IE induced secretion of IL-6 (median= 622 pg/ml [IQR=1,250-240], n=9) but no IL-1β or TNF, whereas PPS-opsonized IE induced secretion of IL-1β (18.6 pg/mL [34.2-14.4]) and TNF (113 pg/ml [421-17.0]) and increased IL-6 secretion (2,195 pg/ml [4,658-1,095]). Opsonized, but not unopsonized, CS2 IE activated caspase-1 cleavage and enzymatic activity in MDM showing that Fc receptor-mediated phagocytosis activates the inflammasome. MDM attached to IgG-coated surfaces however secreted IL-1β in response to unopsonized IE, suggesting that internalization of IE is not absolutely required to activate the inflammasome and stimulate IL-1β secretion.
  
  
  CONCLUSIONS
  It is concluded that IL-6 secretion from MDM in response to CS2 IE does not require phagocytosis, whereas secretion of TNF and IL-1β is dependent on Fcγ receptor-mediated phagocytosis; for IL-1β, this occurs by activation of the inflammasome. The data presented in this paper show that generating antibody responses to blood-stage malaria parasites is potentially beneficial both in reducing parasitaemia via Fcγ receptor-dependent macrophage phagocytosis and in generating a robust pro-inflammatory response.
authors.affiliation: Centre for Virology, Burnet Institute, PO Box 2284, Melbourne, Victoria, 3001, Australia.
authors.forename: Jingling
authors.initials: J
authors.lastname: Zhou
authors.forename: Louise E
authors.initials: LE
authors.lastname: Ludlow
authors.forename: Wina
authors.initials: W
authors.lastname: Hasang
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Rogerson
authors.forename: Anthony
authors.initials: A
authors.lastname: Jaworowski
mesh_terms: D015153:Blotting, Western; D016207:Cytokines / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000469:parasitology*; D006801:Humans; D058847:Inflammasomes / Q000378:metabolism*; D008264:Macrophages / Q000276:immunology*; D016778:Malaria, Falciparum / Q000276:immunology* / Q000469:parasitology; D009895:Opsonin Proteins / Q000276:immunology*; D010963:Plasmodium falciparum / Q000276:immunology*; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D016207:Cytokines; D058847:Inflammasomes; D009895:Opsonin Proteins