Connexin-43-dependent ATP release mediates macrophage activation during sepsis
- University of Bern, Switzerland
Abstract
Bacterial spillage into a sterile environment following intestinal hollow-organ perforation leads to peritonitis and fulminant sepsis. Outcome of sepsis critically depends on macrophage activation by extracellular ATP-release and associated autocrine signaling via purinergic receptors. ATP-release mechanisms, however, are poorly understood. Here we show that TLR-2 and -4 agonists trigger ATP-release via Connexin-43 hemichannels in macrophages leading to poor sepsis survival. In humans, Connexin-43 was upregulated on macrophages isolated from the peritoneal cavity in patients with peritonitis but not in healthy controls. Using a murine peritonitis/sepsis model, we identified increased Connexin-43 expression in peritoneal and hepatic macrophages. onditional Lyz2cre/creGja1flox/flox mice were developed to specifically assess Connexin-43 impact in macrophages. Both macrophage-specific Connexin-43 deletion and pharmacological Connexin-43 blockade were associated with reduced cytokine secretion by macrophages in response to LPS and CLP, ultimately resulting in increased survival. In conclusion, inhibition of autocrine Connexin-43-dependent ATP signaling on macrophages improves sepsis outcome.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files.
Article and author information
Author details
Funding
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (323530_158117)
- Michel Dosch
Novartis Stiftung für Medizinisch-Biologische Forschung (14C160)
- Michel Dosch
- Guido Beldi
University of Bern (Interdisciplinary Grant)
- Guido Beldi
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (166594)
- Guido Beldi
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (146986)
- Guido Beldi
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Jos WM van der Meer, Radboud University Medical Centre, Netherlands
Ethics
Animal experimentation: Animal experiments were planned, carried out and reported in agreement with current 3R and ARRIVE guidelines (Kilkenny et al., 2010) and approved according to Swiss animal protection laws by the Veterinary Authorities of the Canton Bern, Switzerland (license no. BE 4/15).
Human subjects: All human studies were approved by the Ethical Commission of the Canton Bern and written informed consent was obtained from all subjects. Peritoneal fluid collection at the beginning of an operation was included in a larger clinical trial, whose protocol is published on ClinicalTrials.gov (NCT03554148, Study ID Number: 2017-00573).
Version history
- Received: October 10, 2018
- Accepted: February 5, 2019
- Accepted Manuscript published: February 8, 2019 (version 1)
- Version of Record published: March 13, 2019 (version 2)
Copyright
© 2019, Dosch et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Connexin-43-dependent ATP release mediates macrophage activation during sepsis
eLife 8:e42670.
https://doi.org/10.7554/eLife.42670
Further reading
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- Immunology and Inflammation
- Medicine
Background:
Prinflammatory extracellular chromatin from neutrophil extracellular traps (NETs) and other cellular sources is found in COVID-19 patients and may promote pathology. We determined whether pulmonary administration of the endonuclease dornase alfa reduced systemic inflammation by clearing extracellular chromatin.
Methods:
Eligible patients were randomized (3:1) to the best available care including dexamethasone (R-BAC) or to BAC with twice-daily nebulized dornase alfa (R-BAC + DA) for seven days or until discharge. A 2:1 ratio of matched contemporary controls (CC-BAC) provided additional comparators. The primary endpoint was the improvement in C-reactive protein (CRP) over time, analyzed using a repeated-measures mixed model, adjusted for baseline factors.
Results:
We recruited 39 evaluable participants: 30 randomized to dornase alfa (R-BAC +DA), 9 randomized to BAC (R-BAC), and included 60 CC-BAC participants. Dornase alfa was well tolerated and reduced CRP by 33% compared to the combined BAC groups (T-BAC). Least squares (LS) mean post-dexamethasone CRP fell from 101.9 mg/L to 23.23 mg/L in R-BAC +DA participants versus a 99.5 mg/L to 34.82 mg/L reduction in the T-BAC group at 7 days; p=0.01. The anti-inflammatory effect of dornase alfa was further confirmed with subgroup and sensitivity analyses on randomised participants only, mitigating potential biases associated with the use of CC-BAC participants. Dornase alfa increased live discharge rates by 63% (HR 1.63, 95% CI 1.01–2.61, p=0.03), increased lymphocyte counts (LS mean: 1.08 vs 0.87, p=0.02) and reduced circulating cf-DNA and the coagulopathy marker D-dimer (LS mean: 570.78 vs 1656.96 μg/mL, p=0.004).
Conclusions:
Dornase alfa reduces pathogenic inflammation in COVID-19 pneumonia, demonstrating the benefit of cost-effective therapies that target extracellular chromatin.
Funding:
LifeArc, Breathing Matters, The Francis Crick Institute (CRUK, Medical Research Council, Wellcome Trust).
Clinical trial number:
-
- Immunology and Inflammation
Trained immunity is the long-term functional reprogramming of innate immune cells, which results in altered responses toward a secondary challenge. Despite indoxyl sulfate (IS) being a potent stimulus associated with chronic kidney disease (CKD)-related inflammation, its impact on trained immunity has not been explored. Here, we demonstrate that IS induces trained immunity in monocytes via epigenetic and metabolic reprogramming, resulting in augmented cytokine production. Mechanistically, the aryl hydrocarbon receptor (AhR) contributes to IS-trained immunity by enhancing the expression of arachidonic acid (AA) metabolism-related genes such as arachidonate 5-lipoxygenase (ALOX5) and ALOX5 activating protein (ALOX5AP). Inhibition of AhR during IS training suppresses the induction of IS-trained immunity. Monocytes from end-stage renal disease (ESRD) patients have increased ALOX5 expression and after 6 days training, they exhibit enhanced TNF-α and IL-6 production to lipopolysaccharide (LPS). Furthermore, healthy control-derived monocytes trained with uremic sera from ESRD patients exhibit increased production of TNF-α and IL-6. Consistently, IS-trained mice and their splenic myeloid cells had increased production of TNF-α after in vivo and ex vivo LPS stimulation compared to that of control mice. These results provide insight into the role of IS in the induction of trained immunity, which is critical during inflammatory immune responses in CKD patients.
