title: Neutrophil IL-1β processing induced by pneumolysin is mediated by the NLRP3/ASC inflammasome and caspase-1 activation and is dependent on K+ efflux.
abstract: Although neutrophils are the most abundant cells in acute infection and inflammation, relatively little attention has been paid to their role in inflammasome formation and IL-1β processing. In the present study, we investigated the mechanism by which neutrophils process IL-1β in response to Streptococcus pneumoniae. Using a murine model of S. pneumoniae corneal infection, we demonstrated a requirement for IL-1β in bacterial clearance, and we showed that Nod-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC), and caspase-1 are essential for IL-1β production and bacterial killing in the cornea. Neutrophils in infected corneas had multiple specks with enzymatically active caspase-1 (YVAD-FLICA 660), and bone marrow neutrophils stimulated with heat-killed S. pneumoniae (signal 1) and pneumolysin (signal 2) exhibited multiple specks when stained for NLRP3, ASC, or Caspase-1. High-molecular mass ASC complexes were also detected, consistent with oligomer formation. Pneumolysin induced K(+) efflux in neutrophils, and blocking K(+) efflux inhibited caspase-1 activation and IL-1β processing; however, neutrophils did not undergo pyroptosis, indicating that K(+) efflux and IL-1β processing is not a consequence of cell death. There was also no role for lysosomal destabilization or neutrophil elastase in pneumolysin-mediated IL-1β processing in neutrophils. Taken together, these findings demonstrate an essential role for neutrophil-derived IL-1β in S. pneumoniae infection, and they elucidate the role of the NLRP3 inflammasome in cleavage and secretion of IL-1β in neutrophils. Given the ubiquitous presence of neutrophils in acute bacterial and fungal infections, these findings will have implications for other microbial diseases.
authors.affiliation: Department of Ophthalmology and Visual Sciences, Case Western Reserve University, Cleveland, OH 44106;
authors.forename: Mausita
authors.initials: M
authors.lastname: Karmakar
authors.affiliation: Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, OH 44106;
authors.forename: Michael
authors.initials: M
authors.lastname: Katsnelson
authors.affiliation: Department of Clinical Infection, Microbiology and Immunology, Institute of Infection and Global Health, University of Liverpool, Liverpool L69 7BE, United Kingdom;
authors.forename: Hesham A
authors.initials: HA
authors.lastname: Malak
authors.affiliation: Graduate Program in Molecular Microbiology, Department of Molecular Biology and Microbiology and Howard Hughes Medical Institute, Sackler School of Graduate Biomedical Sciences, Tufts University School of Medicine, Boston, MA 02111; and.
authors.forename: Neil G
authors.initials: NG
authors.lastname: Greene
authors.affiliation: Department of Ophthalmology and Visual Sciences, Case Western Reserve University, Cleveland, OH 44106;
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Howell
authors.affiliation: Department of Medicine, Louis Stokes Cleveland Veterans Affairs Medical Center, Cleveland, OH 44106.
authors.forename: Amy G
authors.initials: AG
authors.lastname: Hise
authors.affiliation: Graduate Program in Molecular Microbiology, Department of Molecular Biology and Microbiology and Howard Hughes Medical Institute, Sackler School of Graduate Biomedical Sciences, Tufts University School of Medicine, Boston, MA 02111; and.
authors.forename: Andrew
authors.initials: A
authors.lastname: Camilli
authors.affiliation: Department of Clinical Infection, Microbiology and Immunology, Institute of Infection and Global Health, University of Liverpool, Liverpool L69 7BE, United Kingdom;
authors.forename: Aras
authors.initials: A
authors.lastname: Kadioglu
authors.affiliation: Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, OH 44106;
authors.forename: George R
authors.initials: GR
authors.lastname: Dubyak
authors.affiliation: Department of Ophthalmology and Visual Sciences, Case Western Reserve University, Cleveland, OH 44106; eric.pearlman@case.edu.
authors.forename: Eric
authors.initials: E
authors.lastname: Pearlman
mesh_terms: D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000276:immunology; D001426:Bacterial Proteins / Q000276:immunology; D015153:Blotting, Western; D053468:CARD Signaling Adaptor Proteins; D002352:Carrier Proteins / Q000276:immunology; D020170:Caspase 1 / Q000276:immunology* / Q000378:metabolism; D004195:Disease Models, Animal; D004789:Enzyme Activation / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D015818:Eye Infections, Bacterial / Q000276:immunology / Q000378:metabolism; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D058847:Inflammasomes / Q000276:immunology*; D053583:Interleukin-1beta / Q000276:immunology* / Q000378:metabolism; D007634:Keratitis / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; D009504:Neutrophils / Q000276:immunology* / Q000378:metabolism; D011008:Pneumococcal Infections; D011188:Potassium / Q000378:metabolism*; D015398:Signal Transduction / Q000276:immunology; D013054:Spectrophotometry, Atomic; D013301:Streptolysins / Q000276:immunology
chemical_list: D051017:Apoptosis Regulatory Proteins; D001426:Bacterial Proteins; D053468:CARD Signaling Adaptor Proteins; D002352:Carrier Proteins; D058847:Inflammasomes; D053583:Interleukin-1beta; D000071199:NLR Family, Pyrin Domain-Containing 3 Protein; C492716:Nlrp3 protein, mouse; C498534:Pycard protein, mouse; D013301:Streptolysins; C032290:plY protein, Streptococcus pneumoniae; D020170:Caspase 1; D011188:Potassium