TY - JOUR
T1 - Effect of extracellular vesicles from S. aureus-challenged human neutrophils on macrophages
AU - Allen, Edwina R.
AU - Lempke, Samantha L.
AU - Miller, Michaela M.
AU - Bush, Delaney M.
AU - Braswell, Brandyn G.
AU - Estes, Casey L.
AU - Benedict, Everett L.
AU - Mahon, Andrew R.
AU - Sabo, Shasta L.
AU - Greenlee-Wacker, Mallary C.
N1 - Funding Information:
The authors thank Justin Camiller, Maham Khan, Jenna Owens, Nicholas DeMattei, and Phil Oshel (Central Michigan University) for their excellent technical assistance. The authors also thank Dr. Jeffery Schorey and Li Li (University of Notre Dame) for their help with NTA analysis, Dr. Frank R. DeLeo (Rocky Mountain Laboratories, NIAID) and Dr. Alex Horswill (University of Colorado, Anschutz Medical Campus) for bacteria, and Dr. William Nauseef (University of Iowa) for his advice in the creation of the manuscript. Research in this publication was supported by funds (M.C.G.-W.) awarded by Central Michigan University, as well as the National Institute of General Medical Sciences of the National Institutions of Health under award number R15GM132992. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Funding Information:
The authors thank Justin Camiller, Maham Khan, Jenna Owens, Nicholas DeMattei, and Phil Oshel (Central Michigan University) for their excellent technical assistance. The authors also thank Dr. Jeffery Schorey and Li Li (University of Notre Dame) for their help with NTA analysis, Dr. Frank R. DeLeo (Rocky Mountain Laboratories, NIAID) and Dr. Alex Horswill (University of Colorado, Anschutz Medical Campus) for bacteria, and Dr. William Nauseef (University of Iowa) for his advice in the creation of the manuscript. Research in this publication was supported by funds (M.C.G.‐W.) awarded by Central Michigan University, as well as the National Institute of General Medical Sciences of the National Institutions of Health under award number R15GM132992. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Publisher Copyright:
©2020 Society for Leukocyte Biology
PY - 2020/12
Y1 - 2020/12
N2 - Staphylococcus aureus enhances neutrophil extracellular vesicle (EV) production. To investigate whether S. aureus viability influences EV biogenesis, EVs were isolated from human neutrophils incubated with viable bacteria (bEVs) or heat-killed bacteria (heat-killed EVs). Protein analysis, nanoparticle tracking and transmission electron microscopy showed comparable EV production between subsets, and both viable and nonviable bacteria were also detected in respective EV subsets. As anticipated, S. aureus, as well as bEVs with viable bacteria, were proinflammatory, and killing bacteria with gentamicin reduced cytokine production to baseline levels. Although heat-killed bacteria induced macrophage IL-6 production, heat-killed EVs did not. Additionally, we found that human and bacterial DNA associated with bEVs, but not heat-killed EVs, and that the DNA association could be partially decreased by disrupting electrostatic interactions. We investigated the potential for DNA isolated from EVs (EV-DNA) or EVs to cause inflammation. Although liposomal encapsulation of EV-DNA increased IL-6 production from baseline by 7.5-fold, treatment of bEVs with DNase I had no effect on IL-6 and IL-1β production, suggesting that the DNA did not contribute to the inflammatory response. Filtered EVs, which lacked DNA and associated bacteria, exhibited less proinflammatory activity relative to bEVs, and enhanced macrophage expression of CD86 and HLA-DR. Ultimately, we show that bEVs isolated by differential centrifugation co-purify with bacteria and DNA, and studying their concerted activity and relative contribution to immune response is important to the study of host-pathogen interactions.
AB - Staphylococcus aureus enhances neutrophil extracellular vesicle (EV) production. To investigate whether S. aureus viability influences EV biogenesis, EVs were isolated from human neutrophils incubated with viable bacteria (bEVs) or heat-killed bacteria (heat-killed EVs). Protein analysis, nanoparticle tracking and transmission electron microscopy showed comparable EV production between subsets, and both viable and nonviable bacteria were also detected in respective EV subsets. As anticipated, S. aureus, as well as bEVs with viable bacteria, were proinflammatory, and killing bacteria with gentamicin reduced cytokine production to baseline levels. Although heat-killed bacteria induced macrophage IL-6 production, heat-killed EVs did not. Additionally, we found that human and bacterial DNA associated with bEVs, but not heat-killed EVs, and that the DNA association could be partially decreased by disrupting electrostatic interactions. We investigated the potential for DNA isolated from EVs (EV-DNA) or EVs to cause inflammation. Although liposomal encapsulation of EV-DNA increased IL-6 production from baseline by 7.5-fold, treatment of bEVs with DNase I had no effect on IL-6 and IL-1β production, suggesting that the DNA did not contribute to the inflammatory response. Filtered EVs, which lacked DNA and associated bacteria, exhibited less proinflammatory activity relative to bEVs, and enhanced macrophage expression of CD86 and HLA-DR. Ultimately, we show that bEVs isolated by differential centrifugation co-purify with bacteria and DNA, and studying their concerted activity and relative contribution to immune response is important to the study of host-pathogen interactions.
KW - CD86 and HLA-DR
KW - MRSA
KW - ectosomes
KW - exosomes
KW - extracellular DNA
KW - microparticles
UR - http://www.scopus.com/inward/record.url?scp=85085474494&partnerID=8YFLogxK
U2 - 10.1002/JLB.3AB0320-156R
DO - 10.1002/JLB.3AB0320-156R
M3 - Article
C2 - 32450612
AN - SCOPUS:85085474494
SN - 0741-5400
VL - 108
SP - 1841
EP - 1850
JO - Journal of Leukocyte Biology
JF - Journal of Leukocyte Biology
IS - 6
ER -