Human neutrophil stimulation by influenza virus: relationship of cytoplasmic pH changes to cell activation

Abstract

We have previously demonstrated that influenza A virus (IAV) stimulates the human neutrophil through phospholipase C activation. With the use of the fluorescent indicator 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF), cytoplasmic acidification and subsequent alkalinization are shown to accompany this activation. These responses are not inhibited by pertussis toxin (PT). The alkalinization is mediated largely *but not entirely) by the Na(+)-H+ antiporter and is not initiated, or modulated, by the IAV-induced cytosolic Ca2+ (Cai2+) rise. Rather, protein kinase C (PKC) is likely the mediator of cell alkalinization, based on studies using the PKC inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7). The acidification can be dissociated from the alkalinization response, which is also independent of Cai2+ fluxes and of PKC. Both pHi responses can be dissociated from the respiratory burst. Cytosolic alkalinization and acidification seem to reflect two independently mediated responses of the activated neutrophil, the former resulting ultimately from phospholipase activation and the latter from other activities that are not yet fully characterized.

Identifying the Critical Domain of LL-37 Involved in Mediating Neutrophil Activation in the Presence of Influenza Virus: Functional and Structural Analysis

“…As shown in Fig 1A , IAV alone causes a modest, but significant increase in neutrophil H 2 O 2 production as previously reported [ 42 ]. Pre-incubation of IAV with GI-20 caused a marked, dose-dependent increase in neutrophil H 2 O 2 response to IAV.…”

Inhibition of Neutrophil Respiratory Burst by Nimesulide

“…The reduction in superoxide anion release produced by nimesulide is an intracellular action that might be expected to involve protein kinase C (PKC), in view of the central role played by this enzyme in the events leading to respiratory burst activation. The 93 current belief that PKC is implicated in the activation of the Na+ /H+ antiport is supported by the finding that PKC inhibitors such as H-7 inhibit late alkalinisation in neutrophils stimulated with tMLP or influenza A virus (Hartshorn et al 1990). Results from studies by our group (Bevilacqua et al 1991) give no convincing evidence that PKC activity is directly altered by nimesulide.…”

Critical role of C-terminal residues of the Alzheimer's associated β-amyloid protein in mediating antiviral activity and modulating viral and bacterial interactions with neutrophils

“…Neutrophils from healthy volunteers were isolated to > 95% purity by using dextran precipitation, followed by Ficoll-Paque gradient separation for the separation of mononuclear cells (layering above the Ficoll-Paque) and neutrophils (below the Ficoll-Paque). The neutrophils were purified further by hypotonic lysis to eliminate any contaminating erythrocytes, as previously described [ 21 ]. Cell viability was determined to be >98% by trypan blue staining.…”