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Pneumolysin mediates platelet activation in vitro

Jan-Gert Nel, Chrisna Durnadt, Timothy Mitchell, Charles Feldman, Ronald Anderson, Gregory Tintinger
European Respiratory Journal 2016 48: PA2593; DOI: 10.1183/13993003.congress-2016.PA2593
Jan-Gert Nel
1Haematology, University of Pretoria, Pretoria, GautengSouth Africa
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Chrisna Durnadt
2Institute for Cellular and Molecular Medicine, University of Pretoria, Pretoria, GautengSouth Africa
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Timothy Mitchell
3Institute of Microbiology and Infection, University of Birmingham, Birmingham, United Kingdom
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Charles Feldman
4Internal Medicine, University of the Witwatersrand, Johannesburg, GautengSouth Africa
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Ronald Anderson
2Institute for Cellular and Molecular Medicine, University of Pretoria, Pretoria, GautengSouth Africa
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Gregory Tintinger
5Internal Medicine, University of Pretoria, Pretoria, GautengSouth Africa
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Abstract

INTRODUCTION: Platelets are key players in inflammatory responses, with platelet activation contributing to the pathogenesis of acute lung injury (ALI) and acute coronary events (ACE) in community-acquired pneumonia.

AIM: To explore mechanisms underpinning platelet activation in severe pneumococcal disease, by investigating the effects of exposure of human platelets to the pneumococcal cytotoxin, pneumolysin (Ply) in vitro.

METHODS: Recombinant Ply was used at pathologically relevant concentrations (10–80 nanograms/mL), while platelet activation was measured flow cytometrically according to the level of expression of P-selectin (CD62P). The influx of extracellular calcium, an event which precedes and is a prerequisite for platelet activation, including upregulation of CD62P, was measured using a Fura-2AM-6-based spectrofluorimetric procedure.

RESULTS: Exposure of platelets to Ply resulted in a dose-related increase in the expression of CD62P which was significant at concentrations of >20 ng/ml (p<0.008–p<0.0001), reaching levels comparable to those following receptor-mediated activation with platelet-activating factor (PAF, 400nM). Ply-mediated upregulation of expression of CD62P on platelets was associated with increases in the concentrations of cytosolic calcium and attenuated by depletion of calcium from the cell-suspending medium, as well as by exposure of the cells to a pneumolysoid devoid of pore-forming activity. These observations are consistent with Ply-mediated platelet activation involving sub-lytic pore formation, calcium influx, and discharge of CD62P-expressing α-granules.

CONCLUSION: If operative in the setting of severe pneumococcal disease, Ply-mediated platelet activation may contribute to the pathogenesis of ALI and ACE.

  • Pneumonia
  • Infections
  • Inflammation
  • Copyright ©the authors 2016
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Pneumolysin mediates platelet activation in vitro
Jan-Gert Nel, Chrisna Durnadt, Timothy Mitchell, Charles Feldman, Ronald Anderson, Gregory Tintinger
European Respiratory Journal Sep 2016, 48 (suppl 60) PA2593; DOI: 10.1183/13993003.congress-2016.PA2593

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Pneumolysin mediates platelet activation in vitro
Jan-Gert Nel, Chrisna Durnadt, Timothy Mitchell, Charles Feldman, Ronald Anderson, Gregory Tintinger
European Respiratory Journal Sep 2016, 48 (suppl 60) PA2593; DOI: 10.1183/13993003.congress-2016.PA2593
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