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Background: A dysregulated immune response is emerging as a key feature of critical illness in COVID-19. Neutrophils are key components of early innate immunity that, if not tightly regulated, contribute to uncontrolled systemic inflammation. We sought to decipher the role of neutrophil phenotypes, functions, and homeostasis in COVID-19 disease severity and outcome., Methods: By using flow cytometry, this longitudinal study compares peripheral whole-blood neutrophils from 90 COVID-19 ICU patients with those of 22 SARS-CoV-2-negative patients hospitalized for severe community-acquired pneumonia (CAP) and 38 healthy controls. We also assessed correlations between these phenotypic and functional indicators and markers of endothelial damage as well as disease severity., Results: At ICU admission, the circulating neutrophils of the COVID-19 patients showed continuous basal hyperactivation not seen in CAP patients, associated with higher circulating levels of soluble E- and P-selectin, which reflect platelet and endothelial activation. Furthermore, COVID-19 patients had expanded aged-angiogenic and reverse transmigrated neutrophil subsets-both involved in endothelial dysfunction and vascular inflammation. Simultaneously, COVID-19 patients had significantly lower levels of neutrophil oxidative burst in response to bacterial formyl peptide. Moreover patients dying of COVID-19 had significantly higher expansion of aged-angiogenic neutrophil subset and greater impairment of oxidative burst response than survivors., Conclusions: These data suggest that neutrophil exhaustion may be involved in the pathogenesis of severe COVID-19 and identify angiogenic neutrophils as a potentially harmful subset involved in fatal outcome., (© 2022. The Author(s).)

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Non-steroidal anti-inflammatory drugs (NSAIDs) are commonly used to alleviate symptoms during community-acquired pneumonia (CAP), while neither clinical data nor guidelines encourage this use. Experimental data suggest that NSAIDs impair neutrophil intrinsic functions, their recruitment to the inflammatory site, and the resolution of inflammatory processes after acute pulmonary bacterial challenge. During CAP, numerous observational data collected in hospitalized children, hospitalized adults, and adults admitted to intensive care units (ICUs) support a strong association between pre-hospital NSAID exposure and a delayed hospital referral, a delayed administration of antibiotic therapy, and the occurrence of pleuropulmonary complications, even in the only study that has accounted for a protopathic bias. Other endpoints have been described including a longer duration of antibiotic therapy and a greater hospital length of stay. In all adult series, patients exposed to NSAIDs were younger and had fewer comorbidities. The mechanisms by which NSAID use would entail a complicated course in pneumonia still remain uncertain. The temporal hypothesis and the immunological hypothesis are the two main emerging hypotheses. Current data strongly support an association between NSAID intake during the outpatient treatment of CAP and a complicated course. This should encourage experts and scientific societies to strongly advise against the use of NSAIDs in the management of lower respiratory tract infections.

Objective: To better understand the functional state of circulating neutrophils in patients with ischemic stroke (IS) for planning future clinical trials., Methods: We analyzed by flow cytometry activation state of circulating neutrophils and the distribution of neutrophil peripheral subsets in 41 patients with acute IS less than 6 hours before admission and compared them with 22 age-matched healthy controls., Results: Our results demonstrated continuous basal hyperactivation of circulating neutrophils during acute IS, characterized by lower l-selectin expression and higher CD11b expression at the cell surface, increased ROS production by neutrophils, and greater circulating levels of neutrophil elastase. Neutrophil hyperactivation was associated with deregulation of the equilibrium between apoptotic and necrotic. Patients also had higher percentages than controls of the overactive senescent (CXCR4 bright /CD62L dim ) neutrophil subset and increased percentage of neutrophils with a reverse transendothelial migration (CD54 high CXCR1 low ) phenotype. Importantly, neutrophil alterations were associated with the clinical severity of the stroke, evaluated by its NIH Stroke Scale score., Conclusion: Altogether, our results indicate that during acute IS, the inflammatory properties of circulating neutrophils rise, associated with the expansion of harmful neutrophil subsets. These changes in neutrophil homeostasis, associated with disease severity, may play an instrumental role by contributing to systemic inflammation and to the blood-brain barrier breakdown. Our findings highlight new potential therapeutic approaches of stroke by rebalancing the ratio of senescent to immunosuppressive neutrophils or decreasing reverse neutrophil transmigration or both.

Objective: Recent studies have underlined the effect of systemic inflammation on the pathophysiology of Alzheimer's disease (AD). Neutrophils are key components of early innate immunity and contribute to uncontrolled systemic inflammation if not tightly regulated. The aim of our study was to fully characterize human circulating neutrophils at different disease stages in AD., Methods: We analyzed neutrophil phenotypes and functions in 42 patients with AD (16 with mild cognitive impairment and 26 with dementia), and compared them to 22 age-matched healthy subjects. This study was performed directly in whole blood to avoid issues with data interpretation related to cell isolation procedures., Results: Blood samples from AD patients with dementia revealed neutrophil hyperactivation associated with increased reactive oxygen species production and increased levels of intravascular neutrophil extravascular traps. The homeostasis of circulating neutrophils in these patients also changed: The ratio between the harmful hyperreactive CXCR4 high /CD62L low senescent and the CD16 bright /CD62L dim immunosuppressive neutrophil subsets rose in the later stage of the disease. These abnormalities were greater in fast-decliner than in slow-decliner patients., Interpretation: Our results indicate that the inflammatory properties of circulating neutrophils shift as the percentage of aged neutrophils expands in patients with AD-changes that may play an instrumental role in establishing systemic chronic inflammation. Most important, our data strongly suggest that the neutrophil phenotype may be associated with the rate of cognitive decline and may thus constitute an innovative and prognostic blood biomarker in patients with AD. Ann Neurol 2018;83:387-405., (© 2018 American Neurological Association.)

We report the case of a patient with acute necrotizing colitis due to invasive amebiasis associated with CD4 lymphopenia and impaired neutrophil responses. The course of the disease was characterized by CMV reactivation and severe and recurrent bacterial and fungal infections, which might be related to the decreased CD4 T cell count and the impaired functional capacities of neutrophils, respectively. The clinical outcome was positive with normalization of both CD4 cell count and neutrophil functions.

The aim of our study was to analyze polymorphonuclear neutrophil (PMN) functions in elderly individuals compared with those in healthy young participants, directly in whole blood to avoid issues with data interpretation related to cell isolation procedures. Despite the presence of increased circulating levels of proinflammatory cytokines, resting PMNs from the elderly individuals were not activated as shown by normal CD62L and CD11b expression at the PMN surface and normal constitutive reactive oxygen species (ROS) production. However, suboptimal stimulation induced modulations of CD62L and CD11b expression, which positively correlated with the interleukin-6 circulating level, suggesting a possible in vivo preactivation of old PMNs by this cytokine. In addition, PMN phagocytosis of opsonized Escherichia Coli was decreased in elderly individuals. Furthermore, upon preincubation of elderly whole-blood samples with tumor necrosis factor-α or Toll Receptor agonists, we observed a reduced PMN oxidative burst in response to formyl peptides. Elderly participants also exhibited an increased percentage of the immunosuppressive CD16 bright /CD62L dim PMN subpopulation, which was characterized by a lower phagocytic index and a reduced ROS production compared with the CD16 bright /CD62L bright subset. Thus, the reduced phagocytosis and ROS production associated with an expansion of immunosuppressive CD16 bright /CD62L dim PMN subpopulation might be involved in the increased susceptibility to bacterial and fungal infections with old age., (© The Author 2016. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objective: Synovitis associated with osteoarthritis (OA) is directly responsible for several clinical symptoms and reflects OA's structural progression. This study sought to analyze the expression of proinflammatory mediations, including Interleukin (IL)-17 and IL-22, which play key roles in regulating inflammatory processes, in inflamed and non-inflamed areas of osteoarthritic synovium., Methods: Synovium from knees of 32 OA patients were collected at surgery. Macroscopic evaluation of inflammation enabled inflamed and non-inflamed areas to be separated. Samples were incubated to obtain tissue-conditioned media. Quantitative mRNA expression of proinflammatory mediators was analyzed by RT-PCR and protein levels by ELISA and gelatin zymography. Immunohistochemistry and histology were performed., Results: Inflamed synovium were characterized by increased leukocyte infiltration and a higher vessel-to-tissue area ratio than non-inflamed tissues. Macrophages, T and B lymphocytes, and some neutrophils were found only in the inflamed tissue, and only in the subintimal layer. Levels of proinflammatory cytokines and MMP-9 were significantly higher in tissue-conditioned media from inflamed than non-inflamed tissues. Inflamed areas were associated with higher expression of IL-17 and IL-22, both correlated with the combined release of IL-6, IL-23, and TGFβ1., Conclusion: Our results showed that inflammatory cytokines, including IL-17 and IL-22, are expressed at higher levels by inflamed OA synovium and suggest IL-22 involvement in OA pathophysiology. This study will help identify new therapeutic strategies for OA, especially the targeting of IL-22 to decrease inflammation., (Copyright © 2014. Published by Elsevier Ltd.)

Background: Despite control of HIV infection under antiretroviral therapy (ART), immune T-cell activation persists in patients with controlled HIV infection, who are at higher risk of inflammatory diseases than the general population. PMNs play a key role in host defenses against invading microorganisms but also potentiate inflammatory reactions in cases of excessive or misdirected responses., Objective: The aim of our study was to analyze PMN functions in 60 ART-treated and controlled HIV-infected patients (viral load, <20 RNA copies/mL; CD4 count, ≥ 350 cells/mm(3)) with (HIV[I] group) and without (HIV[NI] group) diseases related to an inflammatory process and to compare them with 22 healthy control subjects., Methods: Flow cytometry was used to evaluate PMN functions in whole-blood conditions. We studied in parallel the activation markers of T lymphocytes and monocytes and the proinflammatory cytokine environment., Results: Blood samples from HIV-infected patients revealed basal PMN hyperactivation associated with deregulation of the apoptosis/necrosis equilibrium. Interestingly, this hyperactivation was greater in HIV(I) than HIV(NI) patients and contrasted with a lack of monocyte activation in both groups. The percentage of circulating cells producing IL-17 was also significantly higher in HIV-infected patients than in control subjects and was positively correlated with markers of basal PMN activation. In addition, the detection of IL-22 overproduction in HIV(NI) patients suggests that it might contribute to counteracting chronic inflammatory processes during HIV infection., Conclusions: This study thus demonstrates the presence of highly activated PMNs in HIV-infected patients receiving effective ART and the association of these cells with a specific IL-17/IL-22 environment., (Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Common variable immunodeficiency (CVID) is a heterogeneous antibody deficiency condition with alterations in T cell regulation and function, dendritic and B-cell compartment and represents the most frequent cause of symptomatic primary immunodeficiency. We addressed whether CVID is associated with abnormalities in the polymorphonuclear neutrophil (PMN) compartment, an important component of innate immunity and plays a key role in host defenses against invading microorganisms. We used flow cytometry to examine PMN phenotypic and functional abnormalities in CVID patients, using whole-blood conditions in order to avoid artifacts due to isolation procedures. We demonstrated that PMN from CVID patients displays, at resting state, a decreased expression of CD15, CD11b and CD16b, which might be related to an abnormality in neutrophil maturation. In addition, these neutrophils exhibit a decrease in degranulation, phagocytosis and reactive oxygen species production, as well as an increased death by apoptosis. These PMN abnormalities observed in CVID patients could result in an increased risk for recurrent bacterial infections.

Aspergillus fumigatus is an opportunistic human fungal pathogen that sheds galactosaminogalactan (GG) into the environment. Polymorphonuclear neutrophils (PMNs) and NK cells are both part of the first line of defense against pathogens. We recently reported that GG induces PMN apoptosis. In this study, we show that PMN apoptosis occurs via a new NK cell-dependent mechanism. Reactive oxygen species, induced by the presence of GG, play an indispensable role in this apoptotic effect by increasing MHC class I chain-related molecule A expression at the PMN surface. This increased expression enables interaction between MHC class I chain-related molecule A and NKG2D, leading to NK cell activation, which in turn generates a Fas-dependent apoptosis-promoting signal in PMNs. Taken together, our results demonstrate that the crosstalk between PMNs and NK cells is essential to GG-induced PMN apoptosis. NK cells might thus play a role in the induction of PMN apoptosis in situations such as unexplained neutropenia or autoimmune diseases., (Copyright © 2014 by The American Association of Immunologists, Inc.)

Protein phosphatases Z that are unique to the fungal kingdom have been associated to resistance to high salt concentration, cell wall integrity, cell cycle regulation, and oxidative stress in fungi. In Aspergillus fumigatus, it was shown that PHZA is under the control of the transcription factor Skn7 and is only involved in the control of the oxidative stress. Accordingly, the ΔphzA mutant showed a defect in virulence in an experimental model of corneal infection in immunocompetent animals and that the impact on susceptibility to cell wall drugs is only secondary., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Polymorphonuclear neutrophils (PMN) are the most abundant circulating leukocytes. They represent a first line of innate immunity against a large panel of microbial pathogens, pending development of specific immune responses. The role of PMN in human immunodeficiency virus type 1 (HIV-1) disease has mainly been investigated from the point of view of the increased susceptibility of HIV-1-infected patients to bacterial and fungal infections. However, it is now clear that the relationship between PMN and HIV-1 is far more complex. This review examines both the beneficial and the detrimental effects of PMN during HIV infection., (© 2013 S. Karger AG, Basel.)

Purpose of Review: The pathogenesis of HIV infection is highly complex and involves numerous actors of the immune system. On the one hand, our immunity has a predominant role in limiting HIV replication and the depletion of its targets, but on the other hand, the persistent infection established by the virus is associated with chronic immune activation and inflammation, potentially resulting in the progressive exhaustion of the host immune resources, and in the onset of non-AIDS-defining comorbidities. The thorough study of HIV pathogenesis is increasingly more challenging., Recent Findings: New knowledge together with technological advances offers the possibility to monitor a constellation of cellular immune markers. Here, we discuss the relevance of studying these markers in order to assess the efficacy to control HIV, the inflammatory response to HIV infection, and the alteration and exhaustion of the immune compartments., Summary: Monitoring these cellular immune markers is important to reach a deeper understanding of HIV pathogenesis and to perform a comprehensive clinical follow-up of HIV-infected patients.

Acute pyelonephritis (APN), which is mainly caused by uropathogenic Escherichia coli (UPEC), is the most common bacterial complication in renal transplant recipients receiving immunosuppressive treatment. However, it remains unclear how immunosuppressive drugs, such as the calcineurin inhibitor cyclosporine A (CsA), decrease renal resistance to UPEC. Here, we investigated the effects of CsA in host defense against UPEC in an experimental model of APN. We show that CsA-treated mice exhibit impaired production of the chemoattractant chemokines CXCL2 and CXCL1, decreased intrarenal recruitment of neutrophils, and greater susceptibility to UPEC than vehicle-treated mice. Strikingly, renal expression of Toll-like receptor 4 (Tlr4) and nucleotide-binding oligomerization domain 1 (Nod1), neutrophil migration capacity, and phagocytic killing of E. coli were significantly reduced in CsA-treated mice. CsA inhibited lipopolysaccharide (LPS)-induced, Tlr4-mediated production of CXCL2 by epithelial collecting duct cells. In addition, CsA markedly inhibited Nod1 expression in neutrophils, macrophages, and renal dendritic cells. CsA, acting through inhibition of the nuclear factor of activated T-cells (NFATs), also markedly downregulated Nod1 in neutrophils and macrophages. Silencing the NFATc1 isoform mRNA, similar to CsA, downregulated Nod1 expression in macrophages, and administration of the 11R-VIVIT peptide inhibitor of NFATs to mice also reduced neutrophil bacterial phagocytosis and renal resistance to UPEC. Conversely, synthetic Nod1 stimulating agonists given to CsA-treated mice significantly increased renal resistance to UPEC. Renal transplant recipients receiving CsA exhibited similar decrease in NOD1 expression and neutrophil phagocytosis of E. coli. The findings suggest that such mechanism of NFATc1-dependent inhibition of Nod1-mediated innate immune response together with the decrease in Tlr4-mediated production of chemoattractant chemokines caused by CsA may contribute to sensitizing kidney grafts to APN.

Polymorphonuclear neutrophils (PMN) play a key role in host defenses against invading microorganisms but also potentiate inflammatory reactions in case of excessive or misdirected responses. Release of the alarmin high-mobility group box 1 (HMGB1) by cells that die at an inflammatory site may act as an alert signal for the immune system. We studied the effect of HMGB1 on human PMN migration, using whole-blood samples to avoid cell activation associated with isolation procedures. HMGB1 50-100 ng/ml reduced baseline PMN migration as well as formyl-methionyl-leucyl-phenylalanine- and IL-8-induced PMN chemotaxis. This inhibitory effect was mediated by the RAGE receptor. In contrast, a higher HMGB1 concentration (5,000 ng/ml) had a chemoattractant effect on PMN through IL-8 production. This effect required the engagement of Toll-like receptors 2 and 4 in addition to the RAGE receptor. The A box component of HMGB1, which antagonizes the endogenous protein, reduced chemotaxis and also strongly inhibited the enhancement of PMN migration observed with the highest HMGB1 concentration. In contrast, the B box, reported to be the active form of HMGB1, exerted a chemoattractant effect. These results strongly point to a key regulatory role of HMGB1 in PMN recruitment to inflammatory tissues. The A box component could potentially serve to inhibit inappropriate PMN recruitment during chronic inflammatory disorders associated with excessive HMGB1 release., (Copyright © 2011 S. Karger AG, Basel.)

Objective: Fibrocytes are mesenchymal progenitors involved in normal and pathologic repair. The aims of this study were: 1) to quantify fibrocytes in bronchoalveolar lavage fluid from patients with or without acute lung injury and acute respiratory distress syndrome; and 2) to evaluate the prognostic value of bronchoalveolar lavage fibrocyte percentage in patients with acute lung injury and acute respiratory distress syndrome., Design: Prospective cohort study., Setting: Three intensive care units of a large tertiary referral center., Patients: One hundred twenty-two ventilated patients requiring bronchoalveolar lavage were enrolled (62 acute respiratory distress syndrome, 30 acute lung injury, 30-ventilated patients without acute lung injury and acute respiratory distress syndrome)., Interventions: After bronchoalveolar lavage collection during standard care, the patients were followed up for 28 days and clinical outcome was recorded. Fibrocytes (CD45+/collagen 1+) were quantified in bronchoalveolar lavage by flow cytometry. Comparison of bronchoalveolar lavage fibrocyte percentage from patients with or without acute lung injury and acute respiratory distress syndrome was performed using a Wilcoxon test. A multivariate analysis using a Cox model was performed to study the independent predictors of survival., Measurements and Main Results: Fibrocytes were detected in 90 of 92 (98%) bronchoalveolar lavages from patients with acute lung injury and acute respiratory distress syndrome. The median percentage of bronchoalveolar lavage fibrocytes was significantly higher in patients with acute lung injury and acute respiratory distress syndrome (5.0%) in comparison with ventilated control subjects (0.9%, p < .0001). After adjustment for age, comorbidity of malignancy, and severity of illness, a bronchoalveolar lavage fibrocyte percentage >6% was independently associated with a higher 28-day mortality in patients with acute lung injury and acute respiratory distress syndrome (hazard ratio [95% confidence interval] 6.15 [2.78-13.64], p ≤ .0001). Addition of bronchoalveolar lavage fibrocyte percentage in a clinical model predicting mortality in patients with acute lung injury and acute respiratory distress syndrome improved global fit and discriminatory capacity (c-statistic, 0.78-0.85; p = .007)., Conclusions: Fibrocytes are detectable in bronchoalveolar lavage during acute lung injury and acute respiratory distress syndrome. A bronchoalveolar lavage fibrocyte percentage >6% provides an additive prognostic value to clinical predictors and may be useful to identify patients with acute lung injury and acute respiratory distress syndrome at highest risk of an adverse outcome.

A new polysaccharide secreted by the human opportunistic fungal pathogen Aspergillus fumigatus has been characterized. Carbohydrate analysis using specific chemical degradations, mass spectrometry, ¹H and ¹³C nuclear magnetic resonance showed that this polysaccharide is a linear heterogeneous galactosaminogalactan composed of α1-4 linked galactose and α1-4 linked N-acetylgalactosamine residues where both monosacharides are randomly distributed and where the percentage of galactose per chain varied from 15 to 60%. This polysaccharide is antigenic and is recognized by a majority of the human population irrespectively of the occurrence of an Aspergillus infection. GalNAc oligosaccharides are an essential epitope of the galactosaminogalactan that explains the universal antibody reaction due to cross reactivity with other antigenic molecules containing GalNAc stretches such as the N-glycans of Campylobacter jejuni. The galactosaminogalactan has no protective effect during Aspergillus infections. Most importantly, the polysaccharide promotes fungal development in immunocompetent mice due to its immunosuppressive activity associated with disminished neutrophil infiltrates.

Subversion or exacerbation of antigen-presenting cells (APC) death modulates host/pathogen equilibrium. We demonstrated during in vitro differentiation of monocyte-derived macrophages and monocyte-derived dendritic cells (DCs) that HIV sensitizes the cells to undergo apoptosis in response to TRAIL and FasL, respectively. In addition, we found that HIV-1 increased the levels of pro-apoptotic Bax and Bak molecules and decreased the levels of anti-apoptotic Mcl-1 and FLIP proteins. To assess the relevance of these observations in the context of an experimental model of HIV infection, we investigated the death of APC during pathogenic SIV-infection in rhesus macaques (RMs). We demonstrated increased apoptosis, during the acute phase, of both peripheral blood DCs and monocytes (CD14(+)) from SIV(+)RMs, associated with a dysregulation in the balance of pro- and anti-apoptotic molecules. Caspase-inhibitor and death receptors antagonists prevented apoptosis of APCs from SIV(+)RMs. Furthermore, increased levels of FasL in the sera of pathogenic SIV(+)RMs were detected, compared to non-pathogenic SIV infection of African green monkey. We suggest that inappropriate apoptosis of antigen-presenting cells may contribute to dysregulation of cellular immunity early in the process of HIV/SIV infection.

Background: Polymorphonuclear neutrophils (PMN) play a key role in host defences against invading microorganisms but can also potentiate detrimental inflammatory reactions in case of excessive or misdirected responses. Intravenous immunoglobulins (IVIg) are used to treat patients with immune deficiencies and, at higher doses, in autoimmune, allergic and systemic inflammatory disorders., Methodology/principal Findings: We used flow cytometry to examine the effects of IVIg on PMN functions and survival, using whole-blood conditions in order to avoid artifacts due to isolation procedures. IVIg at low concentrations induced PMN activation, as reflected by decreased L-selectin and increased CD11b expression at the PMN surface, oxidative burst enhancement, and prolonged cell survival. In contrast, IVIg at higher concentrations inhibited LPS-induced CD11b degranulation and oxidative burst priming, and counteracted LPS-induced PMN lifespan prolongation., Conclusions/significance: IVIg appears to have differential, concentration-dependent effects on PMN, possibly supporting the use of IVIg as either an anti-microbial or an anti-inflammatory agent.

Background: Inflammatory bowel diseases (IBDs), such as ulcerative colitis and Crohn's disease, are chronic inflammatory disorders that increase the risk for colorectal cancer. The mitochondrial translocator protein (TSPO) is a high-affinity drug- and cholesterol-binding protein expressed in the colon and its expression is increased in colon cancers. The aim of this study was to investigate TSPO expression in IBD biopsies and to establish an animal model of IBD to examine the role of TSPO. In addition, we evaluated the potential use of TSPO drug ligands in diagnosing and treating IBD., Methods: TSPO expression in IBD biopsies was evaluated using immunohistochemistry. IBD was induced in a rat experimental model via treatment with dextran sodium sulfate (DSS). Colon morphology, TSPO expression, and proinflammatory cytokine production were evaluated in addition to the effect of TSPO drug ligands on disease pathology., Results: TSPO protein levels were elevated in the enterocytes of IBD biopsies. TSPO expression was localized to the enterocyte mitochondria. DSS treatment induced a time-dependent phenotype mimicking IBD with tissue injury and subsequent tissue regeneration. Coadministration of DSS and the TSPO drug ligands PK 11195 or Ro5-4864 increased both the rate of colon ulceration and regeneration, whereas administration of the TSPO drug ligand flunitrazepam partially prevented this pathology. These data correlated with changes in proinflammatory cytokine plasma levels, as well as increased cytokine production and secretion from the colon., Conclusions: TSPO may serve as a marker of the IBD repair process, and TSPO drug ligands should be further evaluated for IBD treatment.

Polymorphonuclear neutrophils (PMN) are terminally differentiated cells with a short life span, in the blood circulation. The prolongation of the life span of PMN is critical in their effectiveness against pathogens, in particular in the tissues. This review summarizes the effect of cytokines on PMN apoptosis and points to some examples of pathological situations characterized by inappropriate cytokine production associated with dysregulation of PMN apoptosis.

Divergent Toll-like receptor 7 (TLR7) and TLR9 signaling has been proposed to distinguish pathogenic from nonpathogenic simian immunodeficiency virus infection in primate models. We demonstrate here that increased expression of type I interferon in pathogenic rhesus macaques compared to nonpathogenic African green monkeys was associated with the recruitment of plasmacytoid dendritic cells in the lymph nodes and the presence of an inflammatory environment early after infection, instead of a difference in the TLR7/9 response.

IL-17 is a potent effector cytokine involved in inflammatory response and antimicrobial defense. We report that SIV infection of rhesus macaques (RMs) results in the emergence of IL-17-expressing cells during the acute phase. This subpopulation appears at day 14 postinfection concomitantly with an increase in TGF-beta and IL-18 expression. This subset, which exhibits phenotypic markers of NK T cells (NKT), rather than Th17 CD4 cells, persists during the chronic phase and is higher in noncontrollers SIV-infected RMs compared with controllers SIV-infected RMs. In contrast, in the nonpathogenic model of SIVagm infection of African green monkeys, no change in the level of IL-17-expressing cells is observed in lymphoid organs. Consistent with the emergence of TGF-beta and IL-18 during the acute phase in SIV-infected RMs, but not in SIV-infected African green monkeys, we demonstrate that in vitro TGF-beta and IL-18 induce the differentiation and expansion of IL-17+NKT+. Altogether, these results demonstrate that IL-17-producing NKT are associated with the pathogenesis of SIV in RMs and suggest that TGF-beta and IL-18 play a role in their development.

We report the first case of granulomatous mastitis due to Corynebacterium kroppenstedtii linked to strongly impaired neutrophil responses to Nod2 agonist and a single nucleotide polymorphism within the NOD2 gene (SNP13 [Leu1007fsinsC]) in a heterozygous state. These findings provided the first demonstration of impaired Nod2 function associated with corynebacterial infection.

Apoptosis, or programmed cell death, is a highly conserved cellular suicide mechanism. Apoptosis is critical to the effective resolution of inflammation, particularly in regulating the lifespan of neutrophils. Neutrophils are key components of the first line of defense against microorganisms. Thus, subversion of this critical host defense mechanism by pathogens can contribute to susceptibility to severe and recurrent infections. In this review, we describe the molecular mechanisms involved in PMN death in relationship with viral infections.

Flow cytometric analysis provides a rapid screen for abnormalities of polymorphonuclear neutrophils (PMN) function and reflect their behavior in vivo more accurately. This review summarizes the major fluorescent probes used to study PMN oxidative burst and apoptosis using flow cytometry (FCM). We also provide examples of FCM studies in physiological and pathological situations, illustrating the advantages of FCM for assessment of PMN oxidative burst and PMN apoptosis. These data point to the role of FCM in detecting primary immunodeficiencies such as IRAK4 deficiency and support the use of the assessment of the PMN oxidative burst for routine testing in patients with bacterial infections. We also demonstrate the utility of whole-blood analysis using FCM for a better understanding of PMN functionality, i.e., tuning PMN responses to inflammatory stimuli. Finally, FCM permits a simultaneous analysis of phenotypic, functional and morphometric parameters assessing whole-blood PMN apoptosis, in particular in response to Toll-like receptor agonists and during simian immunodeficiency virus infection.

Polymorphonuclear neutrophils (PMN) from chronically HIV-infected individuals have been reported to be more prone to die. However, although non-human primates models have been extensively used for improving our knowledge on T cell immunity, the impact of SIV-infection on PMN, in relationships with disease severity, has never been assessed. In our study, we demonstrate that PMN from Rhesus macaques (RMs) of Chinese origin chronically infected with the virulent strain SIVmac251 display increased susceptibility to undergo apoptosis as compared to PMN from RMs infected with the non-pathogenic SIVDeltanef strain. PMN apoptosis was significantly increased in RMs progressing faster to AIDS as compared to non-progressors RMs. Furthermore, the percentage of apoptotic cells correlated with PMN activation state reflected by increased CD11b expression and reactive oxygen species production. Interestingly, whereas inflammatory cytokines IL-8 and IL-1beta prevent in vitro PMN death, the levels of those cytokines were low in RMs progressing towards AIDS. Altogether, increased PMN death during SIV infection is a new pathogenic effect associated with AIDS progression, adding to the long list of markers associated with disruption of defense against infection.

We used pathogenic and nonpathogenic simian models of SIV infection of Chinese and Indian rhesus macaque (RMs) and African green monkeys (AGMs), respectively, to investigate the relationship between polymorphonuclear neutrophil (PMN) death and the extent of viral replication and disease outcome. In this study, we showed that PMN death increased early during the acute phase of SIV infection in Chinese RMs and coincided with the peak of viral replication on day 14. The level of PMN death was significantly more severe in RMs that progressed more rapidly to AIDS and coincided with neutropenia. Neutropenia was also observed in Indian RMs and was higher in non-Mamu-A*01 compared with Mamu-A*01 animals. In stark contrast, no changes in the levels of PMN death were observed in the nonpathogenic model of SIVagm-sab (sabaeus) infection of AGMs despite similarly high viral replication. PMN death was a Bax and Bak-independent mitochondrial insult, which is prevented by inhibiting calpain activation but not caspases. We found that BOB/GPR15, a SIV coreceptor, is expressed on the PMN surface of RMs at a much higher levels than AGMs and its ligation induced PMN death, suggesting that SIV particle binding to the cell surface is sufficient to induce PMN death. Taken together, our results suggest that species-specific differences in BOB/GPR15 receptor expression on PMN can lead to increased acute phase PMN death. This may account for the decline in PMN numbers that occurs during primary SIV infection in pathogenic SIV infection and may have important implications for subsequent viral replication and disease progression.

Chronic granulomatous disease (CGD) results from constitutional inactivating mutations in the CYBB, NCF1, CYBA or NCF2 genes that encode subunits of phagocyte NADPH oxidase. We report the findings of molecular analysis of 80 kindred. In 75 unrelated male and 5 female probands, CGD was suspected on the basis of clinical symptoms, and biological samples were referred to our laboratory between 2000 and 2007. Seventy seven patients were found to have mutations in CYBB, NCF1, CYBA or NCF2 (52 different mutations including 31 mutations not previously reported). CYBB was the most frequently mutated gene (58 males and 3 females, 76%). In autosomal recessive forms of the disease, mutations were found in NCF1 (11 patients), NCF2 (3 patients) and CYBA (2 patients). We observed that significantly fewer females were affected by autosomal recessive CGD than expected (2 females/14 males; p=0.002), suggesting that female patients with CGD may be under diagnosed.

The events that contribute to the progression to AIDS during the acute phase of a primate lentiviral infection are still poorly understood. In this study, we used pathogenic and nonpathogenic simian models of simian immunodeficiency virus (SIV) infection of rhesus macaques (RMs) and African green monkeys (AGMs), respectively, to investigate the relationship between apoptosis in lymph nodes and the extent of viral replication, immune activation, and disease outcome. Here, we show that, in SIVmac251-infected RMs, a marked increased in lymphocyte apoptosis is evident during primary infection at the level of lymph nodes. Interestingly, the levels of apoptosis correlated with the extent of viral replication and the rate of disease progression to AIDS, with higher apoptosis in RMs of Indian genetic background than in those of Chinese origin. In stark contrast, no changes in the levels of lymphocyte apoptosis were observed during primary infection in the nonpathogenic model of SIVagm-sab infection of AGMs, despite similarly high rates of viral replication. A further and early divergence between SIV-infected RMs and AGMs was observed in terms of the dynamics of T- and B-cell proliferation in lymph nodes, with RMs showing significantly higher levels of cycling cells (Ki67(+)) in the T-cell zones in association with relatively low levels of Ki67(+) in the B-cell zones, whereas AGMs displayed a low frequency of Ki67(+) in the T-cell area but a high proportion of Ki67(+) cells in the B-cell area. As such, this study suggests that species-specific host factors determine an early immune response to SIV that predominantly involves either cellular or humoral immunity in RMs and AGMs, respectively. Taken together, these data are consistent with the hypotheses that (i) high levels of T-cell activation and lymphocyte apoptosis are key pathogenic factors during pathogenic SIV infection of RMs and (ii) low T-cell activation and apoptosis are determinants of the AIDS resistance of SIVagm-infected AGMs, despite high levels of SIVagm replication.

Polymorphonuclear neutrophils (PMN) play a key role in innate immunity. Their activation and survival are tightly regulated by microbial products via pattern recognition receptors such as TLRs, which mediate recruitment of the IL-1R-associated kinase (IRAK) complex. We describe a new inherited IRAK-4 deficiency in a child with recurrent pyogenic bacterial infections. Analysis of the IRAK4 gene showed compound heterozygosity with two mutations: a missense mutation in the death domain of the protein (pArg12Cys) associated in cis-with a predicted benign variant (pArg391His); and a splice site mutation in intron 7 that led to the skipping of exon 7. A nontruncated IRAK-4 protein was detected by Western blotting. The patient's functional deficiency of IRAK-4 protein was confirmed by the absence of IRAK-1 phosphorylation after stimulation with all TLR agonists tested. The patient's PMNs showed strongly impaired responses (L-selectin and CD11b expression, oxidative burst, cytokine production, cell survival) to TLR agonists which engage TLR1/2, TLR2/6, TLR4, and TLR7/8; in contrast, the patient's PMN responses to CpG-DNA (TLR9) were normal, except for cytokine production. The surprisingly normal effect of CpG-DNA on PMN functions and apoptosis disappeared after pretreatment with PI3K inhibitors. Together, these results suggest the existence of an IRAK-4-independent TLR9-induced transduction pathway leading to PI3K activation. This alternative pathway may play a key role in PMN control of infections by microorganisms other than pyogenic bacteria in inherited IRAK-4 deficiency.

Interleukin-10 (IL-10) exerts its anti-inflammatory properties by down-regulating polymorphonuclear neutrophil (PMN) functions such as reactive oxygen species (ROS) production via NADPH oxidase. The molecular mechanisms underlying this process are unclear. Partial phosphorylation of the NADPH oxidase cytosolic component p47(PHOX) induced by proinflammatory cytokines, such as granulocyte-macrophage colony stimulating factor (GM-CSF) and tumor necrosis factor (TNF)-alpha, is essential for priming ROS production by PMN. The aim of this study was to determine whether IL-10 inhibits GM-CSF- and TNFalpha-induced p47(PHOX) phosphorylation and to investigate the molecular mechanisms involved in this effect. We found that IL-10 selectively inhibited GM-CSF- but not TNFalpha-induced p47PHOX phosphorylation in a concentration-dependent manner. As GM-CSF-induced p47PHOX phosphorylation is mediated by extracellular signal-regulated kinase 1/2 (ERK1/2), we tested the effect of IL-10 on this pathway. We found that IL-10 inhibited GM-CSF-induced ERK1/2 activity in an immunocomplex kinase assay. This inhibitory effect was confirmed by analyzing the phosphorylation status of the endogenous substrate of ERK1/2, p90RSK, in intact PMN. Furthermore, IL-10 decreased ROS production by adherent GM-CSF-treated PMN in keeping with the higher ROS production observed in whole blood from IL-10 knockout mice compared to their wild-type counterparts. Together, these results suggest that IL-10 inhibits GM-CSF-induced priming of ROS production by inhibiting p47PHOX phosphorylation through a decrease in ERK1/2 activity. This IL-10 effect could contribute to the tight regulation of NADPH oxidase activity at the inflammatory site.

Neutrophils are polynuclear cells essential to innate immunity. They are the first cells to migrate from the blood to the inflammatory site where they kill pathogens and secrete various mediators that regulate innate and adaptive immunity. Functional steps required for their microbicidal activity include: transendothelial migration, migration towards the invading pathogens, and then recognition, adhesion, engulfment, and killing of the target. Primary deficiencies of these stages are expressed by repeated and/or severe bacterial and fungal infections. These deficiencies include granule abnormalities and leukocyte adhesion deficiencies Type I and II, defective pathogen recognition and the defective oxidative burst that characterizes chronic granulomatous disease.

The superoxide-producing phagocyte NADPH oxidase consists of a membrane-bound flavocytochrome b(558), the cytosol factors p47(phox), p67(phox), p40(phox), and the small GTPase Rac2, which translocate to the membrane to assemble the active complex following neutrophil activation. Interleukin-8 (IL-8) does not activate NADPH oxidase, but potentiates the oxidative burst induced by stimuli such as formyl-methionyl-leucyl-phenylalanine (fMLP) via a priming mechanism. The effect of IL-8 on the components of NADPH oxidase during the priming process has never been investigated in human neutrophils. Here we showed that within 3 min, IL-8 treatment enhanced the Btk- and ERK1/2-dependent phosphorylation of p47(phox), as well as the recruitment of flavocytochrome b(558), p47(phox), and Rac2 into cholesterol-enriched detergent-resistant microdomains (or lipid rafts). Conversely, IL-8 treatment lasting 15 min failed to recruit flavocytochrome b(558), p47(phox), or Rac2, but did enhance the Btk- and p38 MAPK-dependent phosphorylation and the translocation of p67(phox) into detergent-resistant microdomains. Moreover, methyl-beta-cyclodextrin, which disrupts lipid rafts, inhibited IL-8-induced priming in response to fMLP. Our findings indicate that IL-8-induced priming of the oxidative burst in response to fMLP involves a sequential assembly of the NADPH oxidase components in the lipid rafts of neutrophils.