Adenosine Triphosphate Neutralizes Pneumolysin-Induced Neutrophil Activation

Cuypers, Fabian; Klabunde, Björn; Salazar, Manuela Gesell; Surabhi, Surabhi; Skorka, Sebastian B; Burchhardt, Gerhard; Michalik, Stephan; Thiele, Thomas; Rohde, Manfred; Völker, Uwe; Hammerschmidt, Sven; Siemens, Nikolai

doi:10.1093/infdis/jiaa277

Cited by 9 publications

(11 citation statements)

References 43 publications

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“…CD11b regulates leukocyte adhesion and migration to mediate inflammatory responses, while CD62L is an adhesion molecule that mediates neutrophil rolling and sheds upon neutrophil activation. In addition, and consistent with previous studies [ 57 , 58 , 59 , 60 ], we observed that PLY caused an increased release of elastase, suggestive of degranulation and enhanced ROS production ( Figure 1 C,D). A recent study demonstrated that PLY activates NET formation [ 42 ].…”

Section: Discussionsupporting

confidence: 92%

Neutrophil-Derived Extracellular Vesicles Activate Platelets after Pneumolysin Exposure

Letsiou

Alves

Felten

et al. 2021

Cells

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Pneumolysin (PLY) is a pore-forming toxin of Streptococcus pneumoniae that contributes substantially to the inflammatory processes underlying pneumococcal pneumonia and lung injury. Host responses against S. pneumoniae are regulated in part by neutrophils and platelets, both individually and in cooperative interaction. Previous studies have shown that PLY can target both neutrophils and platelets, however, the mechanisms by which PLY directly affects these cells and alters their interactions are not completely understood. In this study, we characterize the effects of PLY on neutrophils and platelets and explore the mechanisms by which PLY may induce neutrophil–platelet interactions. In vitro studies demonstrated that PLY causes the formation of neutrophil extracellular traps (NETs) and the release of extracellular vesicles (EVs) from both human and murine neutrophils. In vivo, neutrophil EV (nEV) levels were increased in mice infected with S. pneumoniae. In platelets, treatment with PLY induced the cell surface expression of P-selectin (CD62P) and binding to annexin V and caused a significant release of platelet EVs (pl-EVs). Moreover, PLY-induced nEVs but not NETs promoted platelet activation. The pretreatment of nEVs with proteinase K inhibited platelet activation, indicating that the surface proteins of nEVs play a role in this process. Our findings demonstrate that PLY activates neutrophils and platelets to release EVs and support an important role for neutrophil EVs in modulating platelet functions in pneumococcal infections.

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Section: Discussionsupporting

confidence: 92%

Neutrophil-Derived Extracellular Vesicles Activate Platelets after Pneumolysin Exposure

Letsiou

Alves

Felten

et al. 2021

Cells

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“…13 Briefly, we incubated washed human platelets in Tyrode buffer containing Ca 21 and Mg 21 with phosphate-buffered saline (PBS), 20 mM thrombin receptor activator peptide 6 (TRAP-6), pneumolysin, or pneumococci (D39, TIGR4, or their pneumolysin-free mutants D39Dply and TIGR4Dply). 14,15 We did grow bacteria to the mid-log exponential phase before incubating 1.8 3 10 6 bacteria with 9 3 10 6 platelets (ratio bacteria platelets 1:5) for 2 or 3 hours. We incubated platelets with pneumolysin at different concentrations or the pneumolysin mutants for 10 minutes.…”

Section: Flow Cytometry-based Platelet-activation Assaymentioning

confidence: 99%

“…To test the cytolytic activity of pneumolysin proteins, the hemolysis assay was performed as described recently. 15 In a subset of experiments, a pneumolysin-inhibiting monoclonal mouse antibody (7.5 mg/mL), polyclonal rabbit anti-pneumolysin antibodies (10 mg/mL), or a pharmaceutical human IgG preparation (1 mg/mL; IgG-enriched Privigen; CSL-Behring) were added.…”

Section: Hemolysis Assaymentioning

confidence: 99%

“…Generation of the pneumolysin mutants has been described recently. 15 After 2 hours and 3 hours of incubation in PBS/Tyrode buffer at 37°C, pneumococci and supernatants were collected for immunoblotting. A total of 1 3 10 8 bacteria and the respective trichloroacetic acid precipitated supernatant of 1 3 10 8 bacteria were run on a 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).…”

Section: Quantification Of Pneumolysin In Pneumococci and Culture Supmentioning

confidence: 99%

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Pneumolysin induces platelet destruction, not platelet activation, which can be prevented by immunoglobulin preparations in vitro

et al. 2020

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Community-acquired pneumonia by primary or superinfections with Streptococcus pneumoniae can lead to acute respiratory distress requiring mechanical ventilation. The pore-forming toxin pneumolysin alters the alveolar-capillary barrier and causes extravasation of protein-rich fluid into the interstitial pulmonary tissue, which impairs gas exchange. Platelets usually prevent endothelial leakage in inflamed pulmonary tissue by sealing inflammation-induced endothelial gaps. We not only confirm that S pneumoniae induces CD62P expression in platelets, but we also show that, in the presence of pneumolysin, CD62P expression is not associated with platelet activation. Pneumolysin induces pores in the platelet membrane, which allow anti-CD62P antibodies to stain the intracellular CD62P without platelet activation. Pneumolysin treatment also results in calcium efflux, increase in light transmission by platelet lysis (not aggregation), loss of platelet thrombus formation in the flow chamber, and loss of pore-sealing capacity of platelets in the Boyden chamber. Specific anti-pneumolysin monoclonal and polyclonal antibodies inhibit these effects of pneumolysin on platelets as do polyvalent human immunoglobulins. In a post hoc analysis of the prospective randomized phase 2 CIGMA trial, we show that administration of a polyvalent immunoglobulin preparation was associated with a nominally higher platelet count and nominally improved survival in patients with severe S pneumoniae–related community-acquired pneumonia. Although, due to the low number of patients, no definitive conclusion can be made, our findings provide a rationale for investigation of pharmacologic immunoglobulin preparations to target pneumolysin by polyvalent immunoglobulin preparations in severe community-acquired pneumococcal pneumonia, to counteract the risk of these patients becoming ventilation dependent. This trial was registered at www.clinicaltrials.gov as #NCT01420744.

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“…In many cases, initial mild manifestations may result in severe invasive diseases such as pneumonia, meningitis, and sepsis [2, 3]. One of the most important and well-studied secreted virulence factors of pneumococci is the pore-forming cytolysin pneumolysin (Ply) [4]. Moreover, pneumococci secrete high amounts of hydrogen peroxide (H 2 O 2 ) as a product of the pneumococcal carbohydrate-metabolizing enzyme pyruvate oxidase SpxB [5].…”

Section: Introductionmentioning

confidence: 99%

Hydrogen Peroxide Is Crucial for NLRP3 Inflammasome-Mediated IL-1β Production and Cell Death in Pneumococcal Infections of Bronchial Epithelial Cells

et al. 2021

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Epithelial cells play a crucial role in detection of the pathogens as well as in initiation of the host immune response. Streptococcus pneumoniae (pneumococcus) is a typical colonizer of the human nasopharynx, which can disseminate to the lower respiratory tract and subsequently cause severe invasive diseases such as pneumonia, sepsis, and meningitis. Hydrogen peroxide (H2O2) is produced by pneumococci as a product of the pyruvate oxidase SpxB. However, its role as a virulence determinant in pneumococcal infections of the lower respiratory tract is not well understood. In this study, we investigated the role of pneumococcal-derived H2O2 in initiating epithelial cell death by analyzing the interplay between 2 key cell death pathways, namely, apoptosis and pyroptosis. We demonstrate that H2O2 primes as well as activates the NLRP3 inflammasome and thereby mediates IL-1β production and release. Furthermore, we show that pneumococcal H2O2 causes cell death via the activation of both apoptotic as well as pyroptotic pathways which are mediated by the activation of caspase-3/7 and caspase-1, respectively. However, H2O2-mediated IL-1β release itself occurs mainly via apoptosis.

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Adenosine Triphosphate Neutralizes Pneumolysin-Induced Neutrophil Activation

Cited by 9 publications

References 43 publications

Neutrophil-Derived Extracellular Vesicles Activate Platelets after Pneumolysin Exposure

Neutrophil-Derived Extracellular Vesicles Activate Platelets after Pneumolysin Exposure

Pneumolysin induces platelet destruction, not platelet activation, which can be prevented by immunoglobulin preparations in vitro

Hydrogen Peroxide Is Crucial for NLRP3 Inflammasome-Mediated IL-1β Production and Cell Death in Pneumococcal Infections of Bronchial Epithelial Cells

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