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Trans-epithelial migration is essential for neutrophil activation during RSV infection.

ABSTRACT: The recruitment of neutrophils to the infected airway occurs early following respiratory syncytial virus (RSV) infection, and high numbers of activated neutrophils in the airway and blood are associated with the development of severe disease. The aim of this study was to investigate whether trans-epithelial migration is sufficient and necessary for neutrophil activation during RSV infection. Here, we used flow cytometry and novel live-cell fluorescent microscopy to track neutrophil movement during trans-epithelial migration and measure the expression of key activation markers in a human model of RSV infection. We found that when migration occurred, neutrophil expression of CD11b, CD62L, CD64, NE, and MPO increased. However, the same increase did not occur on basolateral neutrophils when neutrophils were prevented from migrating, suggesting that activated neutrophils reverse migrate from the airway to the bloodstream side, as has been suggested by clinical observations. We then combined our findings with the temporal and spatial profiling and suggest 3 initial phases of neutrophil recruitment and behavior in the airways during RSV infection; (1) initial chemotaxis; (2) neutrophil activation and reverse migration; and (3) amplified chemotaxis and clustering, all of which occur within 20 min. This work and the novel outputs could be used to develop therapeutics and provide new insight into how neutrophil activation and a dysregulated neutrophil response to RSV mediates disease severity.

SUBMITTER: Robinson E

PROVIDER: S-EPMC11334017 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Trans-epithelial migration is essential for neutrophil activation during RSV infection.

Robinson Elisabeth E Herbert Jenny Amanda JA Palor Machaela M Ren Luo L Larken Isobel I Patel Alisha A Moulding Dale D Cortina-Borja Mario M Smyth Rosalind Louise RL Smith Claire Mary CM

Journal of leukocyte biology 20230301 4

The recruitment of neutrophils to the infected airway occurs early following respiratory syncytial virus (RSV) infection, and high numbers of activated neutrophils in the airway and blood are associated with the development of severe disease. The aim of this study was to investigate whether trans-epithelial migration is sufficient and necessary for neutrophil activation during RSV infection. Here, we used flow cytometry and novel live-cell fluorescent microscopy to track neutrophil movement duri ...[more]

PMID: 36807711

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Project description:BackgroundFollicular helper T cells (Tfh) are pivotal in B cell responses. Activation of the purinergic receptor P2X7 on Tfh cells regulates their activity. We investigated the ATP-P2X7R axis in circulating Tfh (cTfh) cells during Respiratory Syncytial Virus (RSV) infection.MethodsWe analyzed two cohorts: children with RSV infection (moderate, n=30; severe, n=21) and healthy children (n=23). We utilized ELISA to quantify the levels of PreF RSV protein-specific IgG antibodies, IL-21 cytokine, and soluble P2X7R (sP2X7R) in both plasma and nasopharyngeal aspirates (NPA). Additionally, luminometry was employed to determine ATP levels in plasma, NPA and supernatant culture. The frequency of cTfh cells, P2X7R expression, and plasmablasts were assessed by flow cytometry. To evaluate apoptosis, proliferation, and IL-21 production by cTfh cells, we cultured PBMCs in the presence of Bz-ATP and/or P2X7R antagonist (KN-62) and a flow cytometry analysis was performed.ResultsIn children with severe RSV disease, we observed diminished titers of neutralizing anti-PreF IgG antibodies. Additionally, severe infections, compared to moderate cases, were associated with fewer cTfh cells and reduced plasma levels of IL-21. Our investigation revealed dysregulation in the ATP-P2X7R pathway during RSV infection. This was characterized by elevated ATP levels in both plasma and NPA samples, increased expression of P2X7R on cTfh cells, lower levels of sP2X7R, and heightened ATP release from PBMCs upon stimulation, particularly evident in severe cases. Importantly, ATP exposure decreased cTfh proliferative response and IL-21 production, while promoting their apoptosis. The P2X7R antagonist KN-62 mitigated these effects. Furthermore, disease severity positively correlated with ATP levels in plasma and NPA samples and inversely correlated with cTfh frequency.ConclusionOur findings indicate that activation of the ATP-P2X7R pathway during RSV infection may contribute to limiting the cTfh cell compartment by promoting cell death and dysfunction, ultimately leading to increased disease severity.

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Trans-epithelial migration is essential for neutrophil activation during RSV infection.

Publications

Trans-epithelial migration is essential for neutrophil activation during RSV infection.

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