Escherichia coli hemolysin is a potent inductor of phosphoinositide hydrolysis and related metabolic responses in human neutrophils.

Grimminger, Friedrich; Sibelius, Ulf; Bhakdi, Sucharit; Suttorp, Norbert; Seeger, Werner

doi:10.1172/jci115463

Cited by 77 publications

(52 citation statements)

References 61 publications

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“…The THP-1 monocytes also express P2Y 2 receptors, which is likely responsible for the HlyA-induced IP 3 degradation observed in leukocytes (79). Here, we show that stimulation of P2Y receptors by UTP significantly potentiated cell lysis in THP-1 monocytes.…”

Section: Discussionsupporting

confidence: 51%

Inhibition of P2X Receptors Protects Human Monocytes against Damage by Leukotoxin from Aggregatibacter actinomycetemcomitans and α-Hemolysin from Escherichia coli

et al. 2016

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␣-Hemolysin (HlyA) from Escherichia coli and leukotoxin A (LtxA) from Aggregatibacter actinomycetemcomitans are important virulence factors in ascending urinary tract infections and aggressive periodontitis, respectively. The extracellular signaling molecule ATP is released immediately after insertion of the toxins into plasma membranes and, via P2X receptors, is essential for the erythrocyte damage inflicted by these toxins. Moreover, ATP signaling is required for the ensuing recognition and phagocytosis of damaged erythrocytes by the monocytic cell line THP-1. Here, we investigate how these toxins affect THP-1 monocyte function. We demonstrate that both toxins trigger early ATP release and a following increase in the intracellular receptors markedly reduces toxin-induced cytolysis. This pattern is paralleled in freshly isolated human monocytes from healthy volunteers. Interestingly, only a minor fraction of the toxin-damaged THP-1 monocytes eventually lyse. P2X 7 receptor inhibition generally prevents cell damage, except from a distinct cell shrinkage that prevails in response to the toxins. Moreover, we find that preexposure to HlyA preserves the capacity of THP-1 monocytes to phagocytose damaged erythrocytes and may induce readiness to discriminate between damaged and healthy erythrocytes. These findings suggest a new pharmacological target for protecting monocytes during exposure to poreforming cytolysins during infection or injury.␣ -Hemolysin (HlyA) is an important virulence factor frequently produced by strains of pathogenic Escherichia coli (1-3). The frequency with which HlyA-producing E. coli strains are isolated from patients increases with severity of the disease (for a review, see reference 2). HlyA is a pore-forming repeat in toxin (RTX) family member which inserts itself receptor independently into cell membranes (1). The cytotoxic effect of HlyA is massively amplified by ATP release, presumably through the HlyA pore (4) and following P2X receptor activation (5, 6). In erythrocytes, P2X 1 and P2X 7 receptors have been implicated in HlyA-induced hemolysis, and blocking of either of these receptors substantially reduces the hemolysis (5, 6). Interestingly, insertion of a HlyA pore does not cause immediate cell swelling and rupture but initially triggers a significant volume reduction that results from an increase in the intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ) followed by activation of the Ca 2ϩ -sensitive K ϩ and Cl Ϫ channels K Ca 3.1 and TMEM16A (7). During erythrocyte shrinkage, cells expose phosphatidyl serine (PS) in the outer plasma membrane leaflet (7). Recently, we discovered that THP-1 monocytes are more likely to recognize and phagocytose erythrocytes that have been exposed to HlyA (8). This phagocytosis is prevented if HlyA-induced cell damage is diminished by P2X receptor antagonists or if cell shrinkage and PS exposure are blocked (8).Leukotoxin A (LtxA) is a virulence factor often released from Aggregatibacter actinomycetemcomitans in the periodontal connective tissue (9-11). ...

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Section: Discussionsupporting

confidence: 51%

Inhibition of P2X Receptors Protects Human Monocytes against Damage by Leukotoxin from Aggregatibacter actinomycetemcomitans and α-Hemolysin from Escherichia coli

et al. 2016

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“…Bauldry [ 191 also reported no cytochrome-c-specific reduction by atoxin in the presence of 100 pM NADPH. As already cited above, Grimminger [20] showed that a-toxin induced in human neutrophils no cellular responses. All together, these results indicate this is a reproducible method for eosinophil and neutrophil peimeabilization, leaving the cells in a permeable, well functioning and intact state.…”

Section: Discussionmentioning

confidence: 71%

Nanomolar arachidonic acid influences the respiratory burst in eosinophils and neutrophils induced by GTP‐binding protein

Aebischer

Pasche

Jörg

1993

European Journal of Biochemistry

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To investigate a possible role of phospholipase A, (PLA,) in the respiratory burst in bovine eosinophilic and neutrophilic leukocytes dependent on GTP-binding protein (G-protein), we permeabilized these cells with Staphylococcus aureus a-toxin and induced NADPH oxidase activity with the non-hydrolysable GTP analogue GTP [S] or the aluminium tetrafluoro complex AlF,-. Under same experimental conditions, cells responded with different onset times. The onset time for eosinophils was 50-200 s, for neutrophils it was only a few seconds. GTP[S] stimulated in neutrophils only 5 % of the respiratory burst compared to eosinophils, whereas AlF,--induced comparable responses (neutrophils 120% of eosinophils). GDP inhibited these responses with an IC,, value of 2.4 mM. Arachidonic acid showed, with the exception of AIF,-stimulated neutrophils, on both stimuli and cell types an enhancing effect (150%) that reached its maximum at 0.1-1 pM. The PLA, inhibitor 4-bromophenacylbromide reduced the GTP[S]-and AlF,--induced response almost completely (10 pM) and the inhibition was not significantly different for eosinophils and neutrophils (IC50 1 -3 pM). If the respiratory burst was reduced with 4-bromophenacylbromide to 1 -4% of the original value, 10% of the basal NADPH oxidase activity could be restored by addition of only 20-100 nM arachidonic acid. In addition, the PLA, activator adriamycin enhanced the response in a dose-dependent manner and in the same order as arachidonic acid did. The results presented above suggest that the respiratory burst may be regulated by different low-molecular-mass and/or heterotrimeric G-proteins and an active role for arachidonic acid or its metabolites in the activation and the maintenance of the direct G-protein-stimulated respiratory burst in bovine eosinophils and neutrophils.When stimulated by particulate or soluble stimuli like opsonized zymosan, formyl-methionyl-leucyl-phenylalanine, leuktriene B,, platelet-activating factor or the anaphylatoxine C,a, phagocytes of the immune system generate superoxide (O;-) and other reduced oxygen species by using an NADPH oxidase, for reviews, see [l -31. of Fribourg, me du mussCe 5 , CH-1700 Fribourg, Switzerland cytosolic factors p47ph0" and p67ph"", the small G-protein p21r"'1'2 and an associated GDP dissociation inhibitor that have been identified as components of the activating mechanism at least in the cell-free system [4-61.The phosphorylation of several proteins has been reported. The phosphorylation of the p47phoX occurs in normal cells but has been reported to be absent in patients with autosomal recessive chronic granulomatous disease 171. There have been reports suggesting that phospholipase A, (PLA,) is involved in the activation of the respiratory burst oxidase. An increase in PLA, activity and a release of arachidonic acid has been shown to occur following the stimulation of NADPH oxidase by different stimuli in neutrophils as well as in macrophages [8-101. Quinacrine and 4-bromophenacylbromide, inhibitors of PLA,, have previously be...

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“…Production of Ins(1,4,5)P 3 has also been shown to be triggered by two other pore-forming proteins. Grimminger et al (33,35) have shown that Escherichia coli hemolysin (HlyA) leads to phosphoinositide hydrolysis and production of diacylglycerol but the mechanism by which HlyA triggered a G-proteindependent pathway was not further analyzed (33,35). The C5b-9 membrane attack complex was also shown to trigger mobilization of calcium from intracellular stores secondary to activation of phospholipase C and production of Ins(1,4,5)P 3 (36,37).…”

Section: Discussionmentioning

confidence: 99%

Aerolysin Induces G-protein Activation and Ca2+Release from Intracellular Stores in Human Granulocytes

Krause¹,

Fivaz²,

Monod³

et al. 1998

Journal of Biological Chemistry

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Aerolysin is a pore-forming toxin that plays a key role in the pathogenesis of Aeromonas hydrophila infections. In this study, we have analyzed the effect of aerolysin on human granulocytes (HL-60 cells). Proaerolysin could bind to these cells, was processed into active aerolysin, and led to membrane depolarization, indicating that granulocytes are potential targets for this toxin. Fura-2 measurements were used to analyze the effect of aerolysin on cytosolic [Ca 2؉] homeostasis. As expected for a pore-forming toxin, aerolysin addition led to Ca 2؉ influx across the plasma membrane. In addition, the toxin triggered Ca 2؉ release from agonist and thapsigargin-sensitive intracellular Ca 2؉ stores. This Ca 2؉ release was independent of the aerolysin-induced Ca 2؉ influx and occurred in two kinetically distinct phases: an initial rapid and transient phase and a second, more sustained, phase. The first, but not the second phase was sensitive to pertussis toxin. Activation of pertussis toxin-sensitive G-proteins appeared to be a consequence of pore formation, rather than receptor activation through aerolysinbinding, as it: (i) was not observed with a binding competent, insertion-incompetent aerolysin mutant, (ii) had a marked lag time, and (iii) was also observed in response to other bacterial pore-forming toxins (staphylococcal ␣-toxin, streptolysin O) which are thought to bind to different receptors. G-protein activation through pore-forming toxins stimulated cellular functions, as evidenced by pertussis toxin-sensitive chemotaxis. Our results demonstrate that granulocytes are potential target cells for aerolysin and that in these cells, Ca 2؉ signaling in response to a pore-forming toxin involves G-proteindependent cell activation and Ca 2؉ release from intracellular stores.

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Escherichia coli hemolysin is a potent inductor of phosphoinositide hydrolysis and related metabolic responses in human neutrophils.

Cited by 77 publications

References 61 publications

Inhibition of P2X Receptors Protects Human Monocytes against Damage by Leukotoxin from Aggregatibacter actinomycetemcomitans and α-Hemolysin from Escherichia coli

Inhibition of P2X Receptors Protects Human Monocytes against Damage by Leukotoxin from Aggregatibacter actinomycetemcomitans and α-Hemolysin from Escherichia coli

Nanomolar arachidonic acid influences the respiratory burst in eosinophils and neutrophils induced by GTP‐binding protein

Aerolysin Induces G-protein Activation and Ca2+Release from Intracellular Stores in Human Granulocytes

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