Septic Neutrophils Research Articles

LncRNA-mRNA co-expression analysis reveals aquaporin-9-promoted neutrophil extracellular trap formation and inflammatory activation in sepsis

Sepsis is a life-threatening condition caused by an excessive inflammatory response to an infection. However, the precise regulatory mechanism of sepsis remains unclear. Using a strand-specific RNA-sequencing, we identified 115 hub differentially expressed long noncoding RNAs (lncRNAs) and 443 mRNAs in septic patients, primarily participated in crucial pathways including neutrophil extracellular trap (NET) formation and toll-like receptor signaling. Notably, NETs related gene aquaporin-9 (AQP9) and its associated lncRNAs exhibited significant upregulation in septic neutrophils. Functional experiments revealed AQP9 interacts with its lncRNAs to augment the formation of neutrophil NETs. In murine sepsis models, AQP9 inhibition with phloretin reduced proinflammatory cytokine production and lung damage. These findings provide crucial insights into the regulatory role of AQP9 in sepsis, unraveling its interaction with associated lncRNAs in transmitting downstream signals, holding promise in informing the development of novel therapeutic strategies aimed at ameliorating the debilitating effects of sepsis.

INHIBITION OF INTEGRIN VLA-3 AND TETRASPANIN CD151 PROTECTS AGAINST NEUTROPHIL-MEDIATED ENDOTHELIAL DAMAGE.

Background: The recruitment of neutrophils to sites of localized injury or infection is initiated by changes on the surface of endothelial cells located in proximity to tissue damage. Inflammatory mediators, such as TNF-α, increase surface expression of adhesive ligands and receptors on the endothelial surface to which neutrophils tether and adhere. Neutrophils then transit through the activated endothelium to reach sites of tissue injury with little lasting vascular injury. However, in cases of sepsis, the interaction of endothelial cells with highly activated neutrophils can cause damage vascular damage. The identification of molecules that are essential for neutrophil diapedesis may reveal targets of therapeutic opportunity for preservation of endothelial function in the presence of critical illness. We tested the hypothesis that inhibition of neutrophil β1 integrin very late antigen-3 (VLA-3; α3β1) and/or inhibition of the tetraspanin (TM4) family member CD151 would protect against neutrophil-mediated loss of endothelial function. Methods: Blood was obtained from septic patients or healthy donors. Neutrophils were purified, and aliquots were treated with/without proinflammatory molecules. Confluent human umbilical vascular endothelial cells were activated with TNF-α. Electric cell impedance sensing was used to determine monolayer resistance over time after the addition of neutrophils that were treated with blocking antibodies against VLA-3 and/or CD151 or isotype controls. Groups (depending on relevancy) were analyzed by Mann-Whitney U test, Wilcoxon test, or repeated-measures one-way ANOVA. Results: Neutrophils from septic patients and neutrophils activated ex vivo reduced endothelial monolayer resistance to a greater extent than neutrophils from healthy donors. Antibody blockade of VLA-3 and/or CD151 significantly reduced activation-associated endothelial damage. Similar findings were demonstrated on fibronectin, collagen I, collagen IV, and laminin, suggesting that neutrophil surface VLA-3 and CD151 are responsible for endothelial damage regardless of substrata and are likely to be operative in all bodily tissues. Conclusion: This report identifies VLA-3 and CD151 on the activated human neutrophil, which are responsible for damage to endothelial function. Targeting these molecules in vivo may demonstrate preservation of organ function during critical illness.

PKM2/STAT1-mediated PD-L1 upregulation on neutrophils during sepsis promotes neutrophil organ accumulation by serving an anti-apoptotic role

BackgroundDelayed neutrophil apoptosis during sepsis may impact neutrophil organ accumulation and tissue immune homeostasis. Elucidating the mechanisms underlying neutrophil apoptosis may help identify potential therapeutic targets. Glycolysis is critical to neutrophil activities during sepsis. However, the precise mechanisms through which glycolysis regulates neutrophil physiology remain under-explored, especially those involving the non-metabolic functions of glycolytic enzymes. In the present study, the impact of programmed death ligand-1 (PD-L1) on neutrophil apoptosis was explored. The regulatory effect of the glycolytic enzyme, pyruvate kinase M2 (PKM2), whose role in septic neutrophils remains unaddressed, on neutrophil PD-L1 expression was also explored.MethodsPeripheral blood neutrophils were isolated from patients with sepsis and healthy controls. PD-L1 and PKM2 levels were determined by flow cytometry and Western blotting, respectively. Dimethyl sulfoxide (DMSO)-differentiated HL-60 cells were stimulated with lipopolysaccharide (LPS) as an in vitro simulation of septic neutrophils. Cell apoptosis was assessed by annexin V/propidium iodide (annexin V/PI) staining, as well as determination of protein levels of cleaved caspase-3 and myeloid cell leukemia-1 (Mcl-1) by Western blotting. An in vivo model of sepsis was constructed by intraperitoneal injection of LPS (5 mg/kg) for 16 h. Pulmonary and hepatic neutrophil infiltration was assessed by flow cytometry or immunohistochemistry.ResultsPD-L1 level was elevated on neutrophils under septic conditions. Administration of neutralizing antibodies against PD-L1 partially reversed the inhibitory effect of LPS on neutrophil apoptosis. Neutrophil infiltration into the lung and liver was also reduced in PD-L1−/− mice 16 h after sepsis induction. PKM2 was upregulated in septic neutrophils and promoted neutrophil PD-L1 expression both in vitro and in vivo. In addition, PKM2 nuclear translocation was increased after LPS stimulation, which promoted PD-L1 expression by directly interacting with and activating signal transducer and activator of transcription 1 (STAT1). Inhibition of PKM2 activity or STAT1 activation also led to increased neutrophil apoptosis.ConclusionIn this study, a PKM2/STAT1-mediated upregulation of PD-L1 on neutrophils and the anti-apoptotic effect of upregulated PD-L1 on neutrophils during sepsis were identified, which may result in increased pulmonary and hepatic neutrophil accumulation. These findings suggest that PKM2 and PD-L1 could serve as potential therapeutic targets.

PD-L1 promotes GSDMD-mediated NET release by maintaining the transcriptional activity of Stat3 in sepsis-associated encephalopathy.

Sepsis-associated encephalopathy (SAE), as shown as acute and long-term cognitive impairment, is associated with increased mortality of sepsis. The causative factors of SAE are diverse and the underlying pathological mechanisms of SAE remain to be fully elucidated. Multiple studies have demonstrated a crucial role of microglia in the development of SAE, but the role of neutrophils and neutrophil extracellular traps (NETs) in SAE is still unclear. Here, we firstly show that in murine sepsis model, neutrophils and NETs promote blood-brain barrier (BBB) disruption, neuronal apoptosis and microglia activation in hippocampus and induce hippocampus-dependent memory impairment. Anti-Gr-1 antibody or DNase I treatment attenuates these sepsis-induced changes. Then, we find that genetic deletion of neutrophil GSDMD or PD-L1 reduces NET release and improves SAE in murine sepsis model. Finally, in human septic neutrophils, p-Y705-Stat3 binds to PD-L1, promotes PD-L1 nuclear translocation and enhances transcription of the gasdermin D (GSDMD) gene. In summary, our findings firstly identify a novel function of PD-L1 in maintaining transcriptional activity of p-Y705-Stat3 to promote GSDMD-dependent NET release in septic neutrophils, which plays a critical role in the development of SAE.

A2A adenosine receptor activation prevents neutrophil aging and promotes polarization from N1 towards N2 phenotype.

Extracellular adenosine is a biologically active signaling molecule that accumulates at sites of metabolic stress in sepsis. Extracellular adenosine has potent immunosuppressive effects by binding to and activating G protein-coupled A2A adenosine receptors (A2AARs) on the surface of neutrophils. A2AAR signaling reproduces many of the phenotypic changes in neutrophils that are characteristic of sepsis, including decreased degranulation, impaired chemotaxis, and diminished ability to ingest and kill bacteria. We hypothesized that A2AARs also suppress neutrophil aging, which precedes cell death, and N1 to N2 polarization. Using human neutrophils isolated from healthy subjects, we demonstrate that A2AAR stimulation slows neutrophil aging, suppresses cell death, and promotes the polarization of neutrophils from an N1 to N2 phenotype. Using genetic knockout and pharmacological blockade, we confirmed that A2AARs decrease neutrophil aging in murine sepsis induced by cecal ligation and puncture. A2AARs expression is increased in neutrophils from septic patients compared to healthy subject but A2AAR expression fails to correlate with aging or N1/N2 polarization. Our data reveals that A2AARs regulate neutrophil aging in healthy but not septic neutrophils.

Identification of differentially expressed genes, transcription factors, microRNAs and pathways in neutrophils of sepsis patients through bioinformatics analysis.

Sepsis has been recognized to be a life-threatening organ dysfunction caused by the dysregulation of the host response to infections. Our work aims to screen key biomarkers related to neutrophils in sepsis using bioinformatics analysis. For this purpose, the microarray datasets related to neutrophils in sepsis patients were downloaded from the Gene Expression Omnibus (GEO) database. According to the Bayesian test, the Limma package in R was used to screen differentially expressed genes (DEGs). Then, DEGs were uploaded to the DAVID online diagnostic tool for subsequent Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment on the selected DEGs. Next, protein-protein interaction (PPI) network was established based on the selected DEGs using the STRING website and the Cytoscape software. Furthermore, according to the function of the iRegulon plug-in in Cytoscape, our study further predicts and established regulatory networks related to transcription factors and regulatory genes. In addition, the miRWalk2.0 database was used to search for miRNA-DEG pairs, associated with the conduction of intersections of miRNAs predicted by TargetScan, Miranda, miRDB and RNA22 databases. Then, these miRNA-DEG pairs were also displayed in the form of a regulatory network through Cytoscape. Finally, two datasets were selected to verify the screened genes, regulatory factors, and miRNAs, to plot receiver operating characteristics (ROC) curves and compute the area under the curve (AUC) values. The results showed that AKT1, MMP9, ARG1, ETS1 targeting AKT1, and has-miR-124-3p targeting RPS6KA5 may have diagnostic value for patients with sepsis and septic shock. While further experimental studies are required to confirm their role in septic neutrophils.

TICAM2-related pathway mediates neutrophil exhaustion

Pathogenic inflammation and immune suppression are the cardinal features that underlie the pathogenesis of severe systemic inflammatory syndrome and sepsis. Neutrophil exhaustion may play a key role during the establishment of pathogenic inflammation and immune suppression through elevated expression of inflammatory adhesion molecules such as ICAM1 and CD11b as well as immune-suppressors such as PD-L1. However, the mechanism of neutrophil exhaustion is not well understood. We demonstrated that murine primary neutrophils cultured in vitro with the prolonged lipopolysaccharides (LPS) stimulation can effectively develop an exhaustive phenotype resembling human septic neutrophils with elevated expression of ICAM1, CD11b, PD-L1 as well as enhanced swarming and aggregation. Mechanistically, we observed that TICAM2 is involved in the generation of neutrophil exhaustion, as TICAM2 deficient neutrophils have the decreased expression of ICAM1, CD11b, PD-L1, and the reduced aggregation following the prolonged LPS challenge as compared to wild type (WT) neutrophils. LPS drives neutrophil exhaustion through TICAM2 mediated activation of Src family kinases (SFK) and STAT1, as the application of SFK inhibitor Dasatinib blocks neutrophil exhaustion triggered by the prolonged LPS challenge. Functionally, TICAM2 deficient mice were protected from developing severe systemic inflammation and multi-organ injury following the chemical-induced mucosal damage. Together, our data defined a key role of TICAM2 in facilitating neutrophil exhaustion and that targeting TICAM2 may be a potential approach to treating the severe systemic inflammation.

Bioinformatics Analysis for Multiple Gene Expression Profiles in Sepsis.

BackgroundThis work aimed to screen key biomarkers related to sepsis progression by bioinformatics analyses.Material/MethodsThe microarray datasets of blood and neutrophils from patients with sepsis or septic shock were downloaded from Gene Expression Omnibus database. Then, differentially expressed genes (DEGs) from 4 groups (sepsis versus normal blood samples; septic shock versus normal blood samples; sepsis neutrophils versus normal controls and septic shock neutrophils versus controls) were respectively identified followed by functional analyses. Subsequently, protein–protein network was constructed, and key functional sub-modules were extracted. Finally, receiver operating characteristic analysis was conducted to evaluate diagnostic values of key genes.ResultsThere were 2082 DEGs between blood samples of sepsis patients and controls, 2079 DEGs between blood samples of septic shock patients and healthy individuals, 6590 DEGs between neutrophils from sepsis and controls, and 1056 DEGs between neutrophils from septic shock patients and normal controls. Functional analysis showed that numerous DEGs were significantly enriched in ribosome-related pathway, cell cycle, and neutrophil activation involved in immune response. In addition, TRIM25 and MYC acted as hub genes in protein–protein interaction (PPI) analyses of DEGs from microarray datasets of blood samples. Moreover, MYC (AUC=0.912) and TRIM25 (AUC=0.843) had great diagnostic values for discriminating septic shock blood samples and normal controls. RNF4 was a hub gene from PPI analyses based on datasets from neutrophils and RNF4 (AUC=0.909) was capable of distinguishing neutrophil samples from septic shock samples and controls.ConclusionsOur findings identified several key genes and pathways related to sepsis development.

Identification of CD177 as the most dysregulated parameter in a microarray study of purified neutrophils from septic shock patients

Sepsis represents the host’s systemic inflammatory response to an infection. In this condition, immune response associates a marked inflammatory response and the delayed development of severe dysfunctions affecting both innate and adaptive responses. As neutrophils are the first line of defense against infection, they are central to the pathophysiology of sepsis in first hours. Nevertheless, their role during immunosuppression phase remains elusive.The main objective of the current work was to perform a transcriptomic study on purified neutrophils from septic shock patients (n=9) so as to identify genes that are differentially expressed during the first week after disease onset both (3–4 and 6–8days) versus healthy donors. Then, 45 septic shock patients were prospectively enrolled to confirm results at the protein level using flow cytometry. Twenty healthy volunteers (HV) were also included for the whole study. By comparing the transcriptome of purified neutrophils, we identified 364 up-regulated and 328 down-regulated genes differentially expressed. Of them, CD177 mRNA, coding for an activation molecule in chemotaxis, had the highest fold change modulation between patients and HV. This increase was then confirmed at the protein level. There was a constant subset of neutrophils that did not express CD177. However, when positive, septic neutrophils presented with significantly increased CD177 expression. Of note, no association between CD177 overexpression and features of immunosuppression has been highlighted. In addition, this up-regulation was negatively correlated with a decreased expression of CD10, a characteristic of immature myeloid cells.In conclusion, in this exploratory work, we shed light on the increased CD177 mRNA and protein expressions in circulating neutrophils after septic shock. Considering the potential dual roles of CD177 neutrophil (i.e., maturation/chemotaxis), negatively correlated in this study, its participation in septic shock pathophysiology deserves further investigation. Furthermore, its potential as a biomarker for sepsis would deserve to be investigated in large cohorts of patients.

Measurement of functional and morphodynamic neutrophil phenotypes in systemic inflammation and sepsis

We read with great interest the review by Leliefeld et al. [1] published in Critical Care on the role of neutrophils in immune paralysis during systemic inflammation (SI). This is of high clinical importance since immune paralysis potentially increases susceptibility to new infections or may cause inability to clear existing infections, leading to detrimental outcome. One mechanism proposed for immune paralysis is the release of neutrophil populations with decreased microbicidal properties [1]. During SI, heterogeneous subsets of neutrophils exist with different priming states and functions [2]. In particular, large numbers of immature neutrophils appear in the circulation with diminished expression of receptors, important for pathogen killing [3]. We recently reviewed the literature on morphodynamic changes in neutrophils during sepsis [4]. Sepsis increases neutrophil circulating numbers (and percentages of immature neutrophils) and their cell size and stiffness, and decreases migration/chemotaxis, compared with nondiseased conditions or mild infection. Septic neutrophils are also prone to produce neutrophil extracellular traps (NETs).

Sepsis-induced impairment of neutrophil chemotaxis on a microfluidic chip

This study aimed to design a microfluidic chip to measure neutrophil chemotaxis, which is a convenient assay to assess the severity and prognosis of sepsis, and to study the mechanisms involved in the variation of neutrophil chemotaxis. Neutrophil chemotaxis was investigated in this microfluidic device by measuring the migration speed of neutrophils following the LPS concentration gradient stimulus. Neutrophils of 32 sepsis patients were divided into three groups according to the seriousness of physician-diagnosed sepsis, and 12 healthy individuals served as controls. Statistical significance was set at an alpha value of P<0.05. Neutrophil chemotaxis was significantly decreased following the seriousness of sepsis. By contrast, in septic neutrophils, the expression of TLR2 was significantly increased, whereas the expression of CXCR2 was significantly decreased. Neutrophil chemotaxis in sepsis was significantly reduced as compared to healthy individuals. We speculated that impaired neutrophil chemotaxis in sepsis was probably mediated by the TLR2-CXCR2 pathway.

IMMUNE CELL PHENOTYPE AND FUNCTION IN SEPSIS.

Cells of the innate and adaptive immune systems play a critical role in the host response to sepsis. Moreover, their accessibility for sampling and their capacity to respond dynamically to an acute threat increases the possibility that leukocytes might serve as a measure of a systemic state of altered responsiveness in sepsis.The working group of the 14th Acute Dialysis Quality Initiative (ADQI) conference sought to obtain consensus on the characteristic functional and phenotypic changes in cells of the innate and adaptive immune system in the setting of sepsis. Techniques for the study of circulating leukocytes were also reviewed and the impact on cellular phenotypes and leukocyte function of nonextracorporeal treatments and extracorporeal blood purification therapies proposed for sepsis was analyzed.A large number of alterations in the expression of distinct neutrophil and monocyte surface markers have been reported in septic patients. The most consistent alteration seen in septic neutrophils is their activation of a survival program that resists apoptotic death. Reduced expression of HLA-DR is a characteristic finding on septic monocytes, but monocyte antimicrobial function does not appear to be significantly altered in sepsis. Regarding adaptive immunity, sepsis-induced apoptosis leads to lymphopenia in patients with septic shock and it involves all types of T cells (CD4, CD8, and Natural Killer) except T regulatory cells, thus favoring immunosuppression. Finally, numerous promising therapies targeting the host immune response to sepsis are under investigation. These potential treatments can have an effect on the number of immune cells, the proportion of cell subtypes, and the cell function.

The uncovering of ESE-1 in human neutrophils: implication of its role in neutrophil function and survival.

Epithelium-specific Ets transcription factor 1 (ESE-1) is a member of the E26 transformation-specific family of transcription factors that has an epithelial-restricted constitutive expression but is induced by inflammatory stimuli in non-epithelial cells. Here we report that ESE-1 is constitutively expressed in human, but not in murine, neutrophils and that ESE-1 is modestly upregulated in septic patient neutrophils. In normal human neutrophils, ESE-1 was detected at both RNA and protein levels but was found to be an unstable nuclear protein ex vivo. ESE-1 transcription was also induced during all-trans retinoic acid-mediated HL-60 differentiation, a human promyelocytic cell line often used as an in vitro model of human neutrophils. Elf3-/- mice had normal neutrophils but a reduced number of circulating B-lymphocytes. These findings indicate a potential role of ESE-1 in regulating human neutrophil differentiation and function, and that it has different roles in the immune system of different species.

Up-regulation of programmed cell death 1 ligand 1 on neutrophils may be involved in sepsis-induced immunosuppression: an animal study and a prospective case-control study.

Recent studies have shown that neutrophils may display an antigen-presenting function and inhibit lymphocyte proliferation by expressing programmed cell death 1 ligand 1 (PD-L1). The current study was performed to investigate the effect of neutrophils and their pathophysiological significance during sepsis. Neutrophil PD-L1 expression was determined in both septic mice (n = 6) and patients (n = 41). Neutrophils from septic mice were subtyped into PD-L1 and PD-L1 populations to determine their phenotypes and functions. Septic neutrophils were cocultured with lymphocytes to observe the effect of septic neutrophils on lymphocyte apoptosis. The PD-L1 level on neutrophils from septic mice was significantly up-regulated (21.41 ± 4.76%). This level increased with the progression of sepsis and the migration of neutrophils from the bone marrow to the blood and peritoneal cavity. The percentages of CD11a, CD62L, and C-C chemokine receptor type 2 were lower, whereas the percentages of CD16 and CD64 were higher on PD-L1 neutrophils than on PD-L1 neutrophils. The migratory capacity of PD-L1 neutrophils was compromised. Septic neutrophils induced lymphocyte apoptosis via a contact mechanism, and this process could be reversed by anti-PD-L1 antibody. PD-L1 was also up-regulated on neutrophils from patients with severe sepsis (14.6% [3.75%, 42.1%]). The levels were negatively correlated with the monocyte human leukocyte antigen-DR level and positively correlated with the severity of septic patients. Neutrophil PD-L1 was a predictor for the prognosis of severe sepsis, with an area of 0.74 under the receiver operating curve. PD-L1 is up-regulated on neutrophils during sepsis, which may be related to sepsis-induced immunosuppression.

Mast cells aggravate sepsis by inhibiting peritoneal macrophage phagocytosis.

Controlling the overwhelming inflammatory reaction associated with polymicrobial sepsis remains a prevalent clinical challenge with few treatment options. In septic peritonitis, blood neutrophils and monocytes are rapidly recruited into the peritoneal cavity to control infection, but the role of resident sentinel cells during the early phase of infection is less clear. In particular, the influence of mast cells on other tissue-resident cells remains poorly understood. Here, we developed a mouse model that allows both visualization and conditional ablation of mast cells and basophils to investigate the role of mast cells in severe septic peritonitis. Specific depletion of mast cells led to increased survival rates in mice with acute sepsis. Furthermore, we determined that mast cells impair the phagocytic action of resident macrophages, thereby allowing local and systemic bacterial proliferation. Mast cells did not influence local recruitment of neutrophils and monocytes or the release of inflammatory cytokines. Phagocytosis inhibition by mast cells involved their ability to release prestored IL-4 within 15 minutes after bacterial encounter, and treatment with an IL-4-neutralizing antibody prevented this inhibitory effect and improved survival of septic mice. Our study uncovers a local crosstalk between mast cells and macrophages during the early phase of sepsis development that aggravates the outcome of severe bacterial infection.

S96 Simvastatin as an adjuvant therapy for infection and sepsis–in-vitro and in-vivo studies suggest pre-emptive / early therapy in the elderly

Ageing is associated with increased episodes of sepsis and poorer outcomes. Statins are associated with improved outcomes during infection. We aimed to characterise the impact of age and acute severe...

Neutrophils from critically ill septic patients mediate profound loss of endothelial barrier integrity

IntroductionSepsis is characterized by systemic immune activation and neutrophil-mediated endothelial barrier integrity compromise, contributing to end-organ dysfunction. Studies evaluating endothelial barrier dysfunction induced by neutrophils from septic patients are lacking, despite its clinical significance. We hypothesized that septic neutrophils would cause characteristic patterns of endothelial barrier dysfunction, distinct from experimental stimulation of normal neutrophils, and that treatment with the immunomodulatory drug β-glucan would attenuate this effect.MethodsBlood was obtained from critically ill septic patients. Patients were either general surgery patients (Primary Sepsis (PS)) or those with sepsis following trauma (Secondary Sepsis (SS)). Those with acute respiratory distress syndrome (ARDS) were identified. Healthy volunteers served as controls. Neutrophils were purified and aliquots were untreated, or treated with fMLP or β-glucan. Endothelial cells were grown to confluence and activated with tissue necrosis factor (TNF)-α . Electric Cell-substrate Impedance Sensing (ECIS) was used to determine monolayer resistance after neutrophils were added. Groups were analyzed by two-way analysis of variance (ANOVA).ResultsNeutrophils from all septic patients, as well as fMLP-normal neutrophils, reduced endothelial barrier integrity to a greater extent than untreated normal neutrophils (normalized resistance of cells from septic patients at 30 mins = 0.90 ± 0.04; at 60 mins = 0.73 ± 0.6 and at 180 mins = 0.56 ± 0.05; p < 0.05 vs normal). Compared to untreated PS neutrophils, fMLP-treated PS neutrophils caused further loss of barrier function at all time points; no additive effect was noted in stimulation of SS neutrophils beyond 30 min. Neutrophils from ARDS patients caused greater loss of barrier integrity than those from non-ARDS patients, despite similarities in age, sex, septic source, and neutrophil count. Neutrophils obtained after resolution of sepsis caused less barrier dysfunction at all time points. β-glucan treatment of septic patients’ neutrophils attenuated barrier compromise, rendering the effect similar to that induced by neutrophils obtained once sepsis had resolved.ConclusionsNeutrophils from septic patients exert dramatic compromise of endothelial barrier integrity. This pattern is mimicked by experimental activation of healthy neutrophils. The effect of septic neutrophils on the endothelium depends upon the initial inflammatory event, correlates with organ dysfunction and resolution of sepsis, and is ameliorated by β-glucan.

Simvastatin antagonizes CD40L secretion, CXC chemokine formation, and pulmonary infiltration of neutrophils in abdominal sepsis

Statins have been reported to exert anti-inflammatory actions and protect against septic organ dysfunction. Herein, we hypothesized that simvastatin may attenuate neutrophil activation and lung damage in abdominal sepsis. Male C57BL/6 mice were pretreated with simvastatin (0.5 or 10 mg/kg) before CLP. In separate groups, mice received an anti-CD40L antibody or a CXCR2 antagonist (SB225002) prior to CLP. BALF and lung tissue were harvested for analysis of neutrophil infiltration, as well as edema and CXC chemokine formation. Blood was collected for analysis of Mac-1 and CD40L expression on neutrophils and platelets, as well as soluble CD40L in plasma. Simvastatin decreased CLP-induced neutrophil infiltration and edema formation in the lung. Moreover, Mac-1 expression increased on septic neutrophils, which was significantly attenuated by simvastatin. Inhibition of CD40L reduced CLP-induced up-regulation of Mac-1 on neutrophils. Simvastatin prevented CD40L shedding from the surface of platelets and reduced circulating levels of CD40L in septic mice. CXC chemokine-induced migration of neutrophils in vitro was decreased greatly by simvastatin. Moreover, simvastatin abolished CLP-evoked formation of CXC chemokines in the lung, and a CXCR2 antagonist attenuated pulmonary accumulation of neutrophils. Our data suggest that the inhibitory effect of simvastatin on pulmonary accumulation of neutrophils may be related to a reduction of CD40L secretion into the circulation, as well as a decrease in CXC chemokine formation in the lung. Thus, these protective mechanisms help to explain the beneficial actions exerted by statins, such as simvastatin, in sepsis.

Neutrophil Apoptosis by Fas/FasL: Harmful or Advantageous in Cardiac Surgery?

Polymorphonuclear leukocytes or neutrophils are the main executors of cellular death, both in septic inflammation during bacterial infection and in sterile inflammation during trauma or surgery. Whereas in septic inflammation neutrophils perform a useful function to fortify the host's defense against infection, in sterile inflammation, by contrast, they contribute to unwelcome tissue damage. Regardless of the situation, activated neutrophils exhibit a prolonged lifespan and delayed apoptotic death which, under normal conditions, is a prerequisite for their natural renewal. Traditionally, delayed neutrophil apoptosis was considered to promote trauma or surgical injury. According to the results of recent studies, however surprising they may appear, the reverse might be in keeping with what happens IN VIVO. Apoptotic signaling in neutrophils could, by contrast, contribute to intrinsic protection of the host's tissues. This review article, aimed preferentially but not exclusively at the cardiac surgeon, presents some new information in support of this viewpoint, which fits in with our own observations.

Circulating neutrophils of septic patients constitutively express IL-10R1 and are promptly responsive to IL-10

Previous studies have demonstrated that neutrophils isolated from the blood of healthy donors do not respond to IL-10 in terms of either activation of signal transducer and activator of transcription-3 (STAT3) tyrosine phosphorylation or induction of suppressor of cytokine signalling (SOCS)-3 protein, unlike autologous mononuclear cells. This was explained by the fact that circulating neutrophils of healthy donors express only IL-10R2, but not IL-10R1, the latter IL-10R chain being essential for mediating IL-10 responsiveness. In this study, we report that peripheral blood neutrophils of septic patients constitutively display, besides IL-10R2, also abundant levels of surface IL-10R1. Consequently, septic neutrophils are promptly responsive to IL-10 in vitro, as revealed by a direct IL-10-mediated induction of STAT3 tyrosine phosphorylation and SOCS-3 gene transcription, mRNA and protein expression. Consistent with the presence of a fully functional IL-10R, modulation of LPS-induced CXCL8, CCL4, tumour necrosis factor-alpha and IL-1ra gene expression was also rapidly induced by IL-10 in septic, but not normal, neutrophils. Collectively, these data uncover that neutrophils of septic patients are predisposed to be promptly responsive to IL-10, presumably to help limiting their pro-inflammatory state. They also fully validate our previous observations, herein in the context of a human disease, that responsiveness of human neutrophils to IL-10 is strictly dependent upon the modulation of IL-10R1 expression.