Dataset Information

Plasma ATP is required for neutrophil activation in a mouse sepsis model.

ABSTRACT: Our previous work has shown that polymorphonuclear neutrophils (PMNs) require cellular adenosine triphosphate (ATP) release and autocrine purinergic signaling for their activation. Here we studied in a mouse model of cecal ligation and puncture (CLP) whether sepsis affects this purinergic signaling process and thereby alters PMN responses after sepsis. Using high-performance liquid chromatography, we found that plasma ATP, adenosine diphosphate (ADP), and adenosine monophosphate (AMP) concentrations increased up to 6-fold during the first 8 h after CLP, reaching top levels that were significantly higher than those in sham control animals without CLP. Although leukocyte and PMN counts in sham animals increased significantly after 4 h, these blood cell counts decreased in sepsis animals. CD11b expression on the cell surface of PMNs of septic animals was significantly higher compared with sham and untreated control animals. These findings suggest increased PMN activation and sequestration of PMN from the circulation after sepsis. Plasma ATP levels correlated with CD11b expression, suggesting that increased ATP concentrations in plasma contribute to PMN activation. We found that treatment of septic mice with the ATP receptor antagonist suramin diminished CD11b expression, indicating that plasma ATP contributes to PMN activation by stimulating P2 receptors of PMNs. Increased PMN activation can protect the host from invading microorganisms. However, increased PMN activation can also be detrimental by promoting secondary organ damage. We conclude that pharmacological targeting of P2 receptors may allow modulation of PMN responses in sepsis.

SUBMITTER: Sumi Y

PROVIDER: S-EPMC4107005 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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Plasma ATP is required for neutrophil activation in a mouse sepsis model.

Sumi Yuka Y Woehrle Tobias T Chen Yu Y Bao Yi Y Li Xiaoou X Yao Yongli Y Inoue Yoshiaki Y Tanaka Hiroshi H Junger Wolfgang G WG

Shock (Augusta, Ga.) 20140801 2

Our previous work has shown that polymorphonuclear neutrophils (PMNs) require cellular adenosine triphosphate (ATP) release and autocrine purinergic signaling for their activation. Here we studied in a mouse model of cecal ligation and puncture (CLP) whether sepsis affects this purinergic signaling process and thereby alters PMN responses after sepsis. Using high-performance liquid chromatography, we found that plasma ATP, adenosine diphosphate (ADP), and adenosine monophosphate (AMP) concentrat ...[more]

PMID: 24675414

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Project description:Neutrophils play a central role in eliminating bacterial pathogens, but may also contribute to end-organ damage in sepsis. Interleukin-8 (IL-8), a key modulator of neutrophil function, signals through neutrophil specific surface receptors CXCR-1 and CXCR-2. In this study a mechanistic computational model was used to evaluate and deploy an extracorporeal sepsis treatment which modulates CXCR-1/2 levels. First, a simplified mechanistic computational model of IL-8 mediated activation of CXCR-1/2 receptors was developed, containing 16 ODEs and 43 parameters. Receptor level dynamics and systemic parameters were coupled with multiple neutrophil phenotypes to generate dynamic populations of activated neutrophils which reduce pathogen load, and/or primed neutrophils which cause adverse tissue damage when misdirected. The mathematical model was calibrated using experimental data from baboons administered a two-hour infusion of E coli and followed for a maximum of 28 days. Ensembles of parameters were generated using a Bayesian parallel tempering approach to produce model fits that could recreate experimental outcomes. Stepwise logistic regression identified seven model parameters as key determinants of mortality. Sensitivity analysis showed that parameters controlling the level of killer cell neutrophils affected the overall systemic damage of individuals. To evaluate rescue strategies and provide probabilistic predictions of their impact on mortality, time of onset, duration, and capture efficacy of an extracorporeal device that modulated neutrophil phenotype were explored. Our findings suggest that interventions aiming to modulate phenotypic composition are time sensitive. When introduced between 3-6 hours of infection for a 72 hour duration, the survivor population increased from 31% to 40-80%. Treatment efficacy quickly diminishes if not introduced within 15 hours of infection. Significant harm is possible with treatment durations ranging from 5-24 hours, which may reduce survival to 13%. In severe sepsis, an extracorporeal treatment which modulates CXCR-1/2 levels has therapeutic potential, but also potential for harm. Further development of the computational model will help guide optimal device development and determine which patient populations should be targeted by treatment.

Project description:Platelet-activating factor (PAF) is an important mediator of the systemic inflammatory response. In the case of sepsis, proper activation and function of neutrophils as the first line of cellular defense are based on a well-balanced physiological response. However, little is known about the role of PAF in cellular changes of neutrophils during sepsis. Therefore, this study investigates the reaction patterns of neutrophils induced by PAF with a focus on membrane potential (MP), intracellular pH, and cellular swelling under physiological and pathophysiological conditions and hypothesizes that the PAF-mediated response of granulocytes is altered during sepsis. The cellular response of granulocytes including MP, intracellular pH, cellular swelling, and other activation markers were analyzed by multiparametric flow cytometry. In addition, the chemotactic activity and the formation of platelet-neutrophil complexes after exposure to PAF were investigated. The changes of the (electro-)physiological response features were translationally verified in a human ex vivo whole blood model of endotoxemia as well as during polymicrobial porcine sepsis. In neutrophils from healthy human donors, PAF elicited a rapid depolarization, an intracellular alkalization, and an increase in cell size in a time- and dose-dependent manner. Mechanistically, the alkalization was dependent on sodium-proton exchanger 1 (NHE1) activity, while the change in cellular shape was sodium flux- but only partially NHE1-dependent. In a pathophysiological altered environment, the PAF-induced response of neutrophils was modulated. Acidifying the extracellular pH in vitro enhanced PAF-mediated depolarization, whereas the increases in cell size and intracellular pH were largely unaffected. Ex vivo exposure of human whole blood to lipopolysaccharide diminished the PAF-induced intracellular alkalization and the change in neutrophil size. During experimental porcine sepsis, depolarization of the MP was significantly impaired. Additionally, there was a trend for increased cellular swelling, whereas intracellular alkalization remained stable. Overall, an impaired (electro-)physiological response of neutrophils to PAF stimulation represents a cellular hallmark of those cells challenged during systemic inflammation. Furthermore, this altered response may be indicative of and causative for the development of neutrophil dysfunction during sepsis.

Dataset Information

Plasma ATP is required for neutrophil activation in a mouse sepsis model.

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Plasma ATP is required for neutrophil activation in a mouse sepsis model.

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