Abstract
BackgroundMulti-organ failure occurs during critical illness and is mediated in part by destructive neutrophil-to-endothelial interactions. The β2 integrin receptor, CR3 (complement receptor 3; Mac-1; CD11b/CD18), which binds endothelial intercellular adhesion molecule-1 (ICAM-1), plays a key role in promoting the adhesion of activated neutrophils to inflamed endothelia which, when prolonged and excessive, can cause vascular damage. Leukadherin-1 (LA-1) is a small molecule allosteric activator of CR3 and has been shown to promote adhesion of blood neutrophils to inflamed endothelium and restrict tissue infiltration. Therefore, LA-1 offers a novel mechanism of anti-inflammatory action by activation, rather than inhibition, of the neutrophil CR3 integrin. However, whether promotion of neutrophil-to-endothelial interaction by this novel therapeutic is of benefit or detriment to endothelial barrier function is not known.MethodsCritically ill septic and trauma patients were prospectively enrolled from the surgical and the trauma ICU. Blood was collected from these patients and healthy volunteers. Neutrophils were isolated by dextran sedimentation and adhered to TNF-α (tumor necrosis factor-α)-activated human umbilical vein endothelial (HUVEC) monolayers in the presence or absence of fMLP (formylmethionine-leucine-phenylalanine) and/or LA-1. Electric cell-substrate impedance sensing (ECIS) and exposure of underlying collagen were used to quantify endothelial barrier function and permeability.ResultsNeutrophils from critically ill trauma and septic patients caused similar degrees of endothelial barrier disruption which exceeded that caused by cells obtained from healthy controls both kinetically and quantitatively. LA-1 protected barrier function in the absence and presence of fMLP which served as a secondary stimulant to cause maximal loss of barrier function. LA-1 protection was also observed by quantifying collagen exposure underlying endothelial cells challenged with fMLP-stimulated neutrophils. LA-1 treatment resulted in decreased migration dynamics of neutrophils crawling on an endothelial monolayer with reduced speed (μm/s = 0.25 ± 0.01 vs. 0.06 ± 0.01, p < 0.05), path length (μm = 199.5 ± 14.3 vs. 42.1 ± 13.0, p < 0.05), and displacement (μm = 65.2 ± 4.7 vs. 10.4 ± 1.3; p < 0.05).ConclusionNeutrophils from patients with trauma or sepsis cause endothelial barrier disruption to a similar extent relative to each other. The CR3 agonist LA-1 protects endothelial barrier function from damage caused by neutrophils obtained from both populations of critically ill patients even when exposed to secondary stimulation.
Highlights
Multi-organ failure occurs during critical illness and is mediated in part by destructive neutrophil-toendothelial interactions
Using Electric cellsubstrate impedance sensing (ECIS), we previously reported that neutrophils from septic patients with ARDS have a more deleterious effect on endothelial barrier function than neutrophils from patients with sepsis but without ARDS or ex vivo activated neutrophils isolated from healthy donors [12]
TNF-αactivated Human umbilical vein endothelial cells (HUVEC) were used throughout the work shown below to mimic the systemic cytokine response during injury and/or sepsis, and thrombin served as a positive control for loss of barrier function
Summary
Multi-organ failure occurs during critical illness and is mediated in part by destructive neutrophil-toendothelial interactions. The β2 integrin receptor, CR3 (complement receptor 3; Mac-1; CD11b/CD18), which binds endothelial intercellular adhesion molecule-1 (ICAM-1), plays a key role in promoting the adhesion of activated neutrophils to inflamed endothelia which, when prolonged and excessive, can cause vascular damage. The neutrophilic response to a localized injury or infection includes a transient interaction with vascular endothelial cells as peripheral blood neutrophils cross from the circulation into an afflicted tissue [1]. When an injury or infection is severe, such as in the critically ill post-trauma or septic patient, systemic hyperactivation of both endothelial cells and neutrophils results in leukocyte-mediated vascular damage which in turn is a contributing factor to multi-system organ failure [3, 4]. There is significant clinical evidence for a mechanistic role of endothelial damage resulting to organ dysfunction
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