July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Involvement of Neutrophils in Human Ocular Toxoplasmosis
Author Affiliations & Notes
  • Liam M. Ashander
    Eye and Vision Health, Flinders University of South Australia, Bedford Park, South Australia, Australia
  • Shervi Lie
    Eye and Vision Health, Flinders University of South Australia, Bedford Park, South Australia, Australia
    Flinders Centre for Innovation in Cancer, Flinders University of South Australia, Bedford Park, South Australia, Australia
  • Yuefang Ma
    Eye and Vision Health, Flinders University of South Australia, Bedford Park, South Australia, Australia
  • Elise Rochet
    Eye and Vision Health, Flinders University of South Australia, Bedford Park, South Australia, Australia
  • Joao M Furtado
    Ophthalmology, Ribeirão Preto Medical School, University of São Paulo, Brazil
  • Binoy Appukuttan
    Eye and Vision Health, Flinders University of South Australia, Bedford Park, South Australia, Australia
    Flinders Centre for Innovation in Cancer, Flinders University of South Australia, Bedford Park, South Australia, Australia
  • Justine R Smith
    Eye and Vision Health, Flinders University of South Australia, Bedford Park, South Australia, Australia
    Flinders Centre for Innovation in Cancer, Flinders University of South Australia, Bedford Park, South Australia, Australia
  • Footnotes
    Commercial Relationships   Liam Ashander, None; Shervi Lie, None; Yuefang Ma, None; Elise Rochet, None; Joao Furtado, None; Binoy Appukuttan, None; Justine Smith, None
  • Footnotes
    Support  NHMRC (Australia) GNT 1066235; ARC (Australia) FT 130101648; ORIA (Australia)
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 4647. doi:https://doi.org/
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Liam M. Ashander, Shervi Lie, Yuefang Ma, Elise Rochet, Joao M Furtado, Binoy Appukuttan, Justine R Smith; Involvement of Neutrophils in Human Ocular Toxoplasmosis. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4647. doi: https://doi.org/.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

Purpose : Ocular toxoplasmosis is a parasitic retinal disease in which tissue destruction is due to T. gondii-induced cell lysis and reactive inflammation. Retinal histopathology demonstrates neutrophils, but involvements have not been investigated. We conducted in vitro experiments to evaluate roles in taxiing T. gondii into the retina and contributing to local inflammation.

Methods : Human neutrophils were isolated from peripheral blood of healthy adults by density gradient centrifugation. Migration assays were performed in transwells seeded with human retinal endothelial cells, using neutrophils infected with GT-1 strain T. gondii tachyzoites (gift of JP Dubey, USDA; multiplicity of infection, MOI=5). Integrity of endothelial monolayers was determined by dextran permeability assay. Infected neutrophils also were migrated in transwells without endothelial cells to chemokines expressed during ocular toxoplasmosis. In other experiments, neutrophils were co-cultured with ARPE19 human retinal pigment epithelial cells previously infected with T. gondii. Neutrophil production of reactive oxygen species (ROS) was estimated by dihydroethidium reaction, and relative expression of inflammatory cytokine transcripts – TNF-α and IL-1β – was measured by RT-qPCR. Effect of tachyzoite lysate on neutrophil activity was also quantified. ARPE19 cell protein expression during infection was profiled by R&D Systems Proteome Profiler Human Cytokine Array.

Results : Neutrophil migration across retinal endothelium was reduced by infection with T. gondii (≥90.0%, p≤0.002). Migration to CXCL1, CXCL2 and CXCL8 – chemokines produced in ocular toxoplasmosis – was also significantly inhibited in infected neutrophils (≥95.5%, p≤0.006). Neutrophils produced higher levels of ROS and inflammatory cytokine transcripts when co-cultured with infected versus uninfected ARPE19 cells (ROS: ≥3.4-fold; TNF-α: ≥19.8-fold; IL-1β: ≥85.7-fold, p≤0.048). Infected ARPE19 cells significantly increased expression of 7 proteins known to activate neutrophils: G-CSF, GM-CSF, IL-1α, IL-18, IL-21, CXCL1, CXCL8 (≥2.6-fold, p≤0.003). Tachyzoite lysate also induced neutrophil activation.

Conclusions : Our findings suggest that in human ocular toxoplasmosis, neutrophils are unlikely to act as parasite taxis. However, they may be attracted by locally produced chemokines into the retina, where they may promote inflammation following activation by T. gondii and retinal pigment epithelial proteins.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×