Propofol Ameliorates Exaggerated Human Neutrophil Activation in a LPS Sepsis Model

Abstract

Background Sepsis is a leading cause of morbidity and mortality worldwide. Many patients suffering from sepsis are treated on intensive care units and many of them require mechanical ventilation under sedation or general anesthesia. Propofol, a drug used for these purposes, is known to interact with polymorphonuclear granulocytes (PMNs). Therefore, the aim of this study was to investigate the influence of propofol on PMN functions after experimental Gram-negative induced sepsis using lipopolysaccharide (LPS) stimulation. Methods A total of 34 granulocyte-enriched samples were collected from healthy subjects. PMNs were isolated by density gradient centrifugation and incubated simultaneously with either 6 µg/mL or 60 µg/mL propofol, or none (control). Additionally, the experimental sepsis samples were incubated with either 40 pg/mL or 400 pg/mL LPS. Live cell imaging was conducted in order to observe granulocyte chemotactic migration, ROS production, and NETosis. Flow cytometry was used to analyze viability and antigen expression. Results Propofol led to significantly reduced PMN track length (p < 0.001) and track speed (p < 0.014) after LPS-induced sepsis in a dose-dependent manner. NETosis (p = 0.018) and ROS production (p = 0.039) were accelerated by propofol without LPS incubation, indicating improved immune function. Propofol also ameliorated LPS-induced increased NETosis and ROS-production. Antigen expression for CD11b, CD62l and CD66b was unaffected by propofol. Conclusion Propofol improves LPS-induced exaggerated PMN activation in an ex vivo model. Beneficial effects due to restored immune function in septic patients might be possible, but needs further investigation.

Is the In Vitro Observed NETosis the Favored Physiological Death of Neutrophils or Mainly Induced by an Isolation Bias?

“…NETosis was observed within 8 h when PMNs were isolated by density gradient separation [ 17 , 27 , 28 ]. The DAPI staining was seen within minutes and only in non-migrating cells, due to NETotic processes.…”

“…This process is promoted by reactive oxygen species (ROS) leading to a violent disruption of the nuclear and cellular membrane and release of chromatin, DNA, citrullinated histones and contents of intracellular granules [ 14 ]. In previous publications of our laboratory, by default, NETosis was observed within 8 h when PMNs were isolated by density gradient separation (DGS) [ 15 , 16 , 17 ].…”

“…Standard procedures to isolate neutrophils are, for instance, the density gradient separation [ 15 , 16 , 17 ], magnetic-activated [ 24 ] or fluorescence-activated cell sorting [ 25 ]. These methods have in common that they involve centrifugation steps.…”

A GABA-receptor agonist reduces pneumonitis severity, viral load, and death rate in SARS-CoV-2-infected mice

“…GABA decreases the secretion of inflammatory factors by human bronchial epithelial cells in vitro ( 37 ) and GABA A -R PAMs reduce macrophage infiltration and inflammatory cytokine levels in bronchoalveolar lavage fluid and reduce rodent and human macrophage inflammatory responses ( 10 , 13 , 33 , 42 , 47 – 49 ). Moreover, GABA A -R ligands inhibit human neutrophil activation ( 50 ) and platelet aggregation ( 51 ), which are potentially important because pulmonary thrombosis often occurs in critically ill COVID-19 patients. Finally, lower levels of plasma GABA are detected in hospitalized COVID-19 patients and associated with the pathogenesis of COVID-19 ( 52 , 53 ), although the basis for this observation is not understood.…”

GABA-receptors are a new druggable target for limiting disease severity, lung viral load, and death in SARS-CoV-2 infected mice

“…GABA reduces human bronchial epithelial cell secretion of inflammatory factors in vitro (33) and GABAA-R PAMs reduce macrophage infiltrates and inflammatory cytokine levels in bronchoalveolar lavage fluid and reduce rodent and human macrophage inflammatory responses (8,11,29,38,(43)(44)(45). Moreover, GABA inhibits human neutrophil activation (46) and platelet aggregation (47), which is potentially important because pulmonary thrombosis often occurs in critically ill COVID-19 patients. Finally, lower levels of plasma GABA are detected in hospitalized COVID-19 patients and associated with the pathogenesis of COVID-19 (48), although the basis for this observation is not understood.…”