Neutrophil Oxidative Activity Research Articles

Adrenergic effect on women's blood neutrophil oxidative activity the first day after delivery.

Recent studies have described a significant role for neutrophils in reproductive processes and their participation in the preparation of the cervix for childbirth and the activation of labor, in the postpartum involution of the uterus, and in the occurrence of preeclampsia. This study aimed to assess the formation of free radicals by neutrophils in the blood of women on the first day after childbirth and to characterize the adrenergic effect on this process. Venous blood samples from 100 female volunteers aged 26-32 years who had 2 or 3 full-term deliveries were collected and analyzed. Various adrenergic compounds were considered (agonists alphaand betaadrenoreceptors, adrenoblockers). The intensity of the respiratory burst of neutrophils and the effect of adrenergic substances on them were assessed with latex-induced luminol-dependent chemiluminescence. Neutrophil activity depends on the stage of the woman's reproductive process: it decreases during pregnancy, reaches the lowest values during childbirth, and increases significantly in the first hours after childbirth. On the first day after childbirth, alpha-1-adrenergic receptors are highly active in neutrophils, through which NADP-H-oxidase is activated and activated oxygen species are formed. At the same time, alphaor beta-agonists inhibit the radical activity of cells. Latex-induced oxidative burst of female blood neutrophils correlates with the stage of the reproductive process. Stressful conditions in the postpartum period can suppress the ability of neutrophils to release reactive oxygen species, which increases the risk of postpartum infections.

Phagocytosis and oxidative activity of neutrophils after interaction with uropathogenic <i>Escherichia coli</i> biofilms

Recurrent urinary tract infections (UTIs) are associated primarily with the ability of Escherichia coli to form biofilms. The interaction of neutrophils, factors of innate immunity, with microorganisms in biofilms is difficult compared to planktonic forms due to the lack of direct contact, as well as due to the antiphagocytic action of the extracellular matrix of biofilms. The purpose of this study was evaluation of neutrophils phagocytic and oxidative activity during interaction with biofilms of uropathogenic E. coli (UPEC) DL82 and R44. Peripheral blood neutrophils from healthy men were isolated using ficoll-urographin double gradient, incubated for 1 h with bacterial cells from biofilms or their supernatants, then leukocytes functional activity was evaluated. Phagocytic activity of neutrophils was determined by the degree of bioluminescence inhibition of bioluminescent strain E. coli K12 TG1 lux+ (pXen) upon their absorption by neutrophils. Production of extracellular reactive oxygen species (ROS) was analyzed by the intensity of luminol-dependent chemiluminescence in spontaneous and stimulated by E. coli K12 variants. Significance of differences was determined using Student’s t-test at p < 0.05. It was found that neutrophils interaction with UPEC biofilm cells or supernatants did not affect the phagocytic activity. E. coli DL82 supernatants reduce neutrophils spontaneous ROS production compared to control and biofilm cells. E. coli R44 supernatants with a low virulence potential did not affect ROS production, while biofilm cells stimulated it. When assessing stimulated ROS production, exposure to R44 strain supernatants did not cause a decrease in neutrophils activation in response to an external stimulus (E. coli K12 cells). Preliminary contact of neutrophils with E. coli R44 bacteria resulted in a high and prolonged level of ROS production compared to the control. Neutrophils interaction with DL82 cells resulted in a higher level of ROS compared to supernatants, however a subsequent rapid depletion of neutrophils oxidative potential was observed. Thus, cells and supernatants of UPEC biofilms can determine the activation of neutrophils.

Deficiency of Leukocyte Surface Glycoproteins Mol, LFA-1, and Leu M5 in a Dog With Recurrent Bacterial Infections: An Animal Model

A dog with severe recurrent bacterial infections, impaired pus formation, delayed wound healing, and severe persistent leukocytosis was the result of a mother-son mating. Assessment of leukocyte function revealed profound abnormalities in adherence-dependent activities including impaired granulocyte adhesion to glass/plastic surfaces or nylon wool, decreased granulocyte aggregation and chemotaxis, and diminished lymphocyte blastogenesis, but normal neutrophil oxidative activity, serum immunoglobulin, and complement levels. By immunofluorescence analysis, CD11b and CD18 monoclonal antibodies specific for the 155-kd alpha polypeptide of Mo1 (gp 155, 94) and the 94 kd beta peptide common to Mo1, LFA-1 (gp 170, 94), and Leu M5 (p 150, 94) (surface molecules that promote leukocyte adhesion) failed to bind to unstimulated and A23187 calcium ionophore-stimulated granulocytes or mononuclear cells of the affected dog as compared with strong specific binding to canine control cells. The Mo1 glycoproteins were only barely detectable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of immunoprecipitates from lysates of 125I surface-labeled neutrophils from the affected dog as compared with intense bands seen with canine control cell precipitates. We conclude that this dog has a severe leukocyte surface glycoprotein deficiency syndrome that is similar, if not identical, to that recently recognized in humans. Dogs with deficiency of leukocyte Mo1, LFA-1, and Leu M5 expression may represent a useful animal model to characterize further the molecular basis for an inherited disorder in leukocyte effector function.

Discrimination of Methionine Sulfoxide and Sulfone by Human Neutrophil Elastase

Human neutrophil elastase (HNE) is a uniquely destructive serine protease with the ability to unleash a wave of proteolytic activity by destroying the inhibitors of other proteases. Although this phenomenon forms an important part of the innate immune response to invading pathogens, it is responsible for the collateral host tissue damage observed in chronic conditions such as chronic obstructive pulmonary disease (COPD), and in more acute disorders such as the lung injuries associated with COVID-19 infection. Previously, a combinatorially selected activity-based probe revealed an unexpected substrate preference for oxidised methionine, which suggests a link to oxidative pathogen clearance by neutrophils. Here we use oxidised model substrates and inhibitors to confirm this observation and to show that neutrophil elastase is specifically selective for the di-oxygenated methionine sulfone rather than the mono-oxygenated methionine sulfoxide. We also posit a critical role for ordered solvent in the mechanism of HNE discrimination between the two oxidised forms methionine residue. Preference for the sulfone form of oxidised methionine is especially significant. While both host and pathogens have the ability to reduce methionine sulfoxide back to methionine, a biological pathway to reduce methionine sulfone is not known. Taken together, these data suggest that the oxidative activity of neutrophils may create rapidly cleaved elastase “super substrates” that directly damage tissue, while initiating a cycle of neutrophil oxidation that increases elastase tissue damage and further neutrophil recruitment.

Malnutrition and inflammation in hemodialysis patients: Comparative evaluation of neutrophil reactive oxygen formation

Impaired phagocytic function has been established in uremic patients. Chemotaxis, particle ingestion, and free radical and metabolic activity were all found to be disturbed in dialysis patients. Malnutrition is common among hemodialysis (HD) patients, with an estimated prevalence of 40% to 70%. Malnutrition-Inflammation Score (MIS) appears to be a useful tool for risk stratification of chronic HD patients. We assessed the correlation between MIS and phagocyte function in HD patients. Forty-four chronic HD patients were enrolled from the dialysis unit. The patients were divided into two groups according to the MIS: 1 to 12 (normal-mild) and 13 to 30 (severely malnourished). Hydrogen peroxide release by polymorphonuclear leukocytes was evaluated using the dihydrorhodamine 123 method. Phagocytic activity of neutrophils was evaluated after stimulation with Escherichia coli bacteria and phorbol 12-myristate 13-acetate (PMA) (positive control). Neutrophil oxidative activity in all HD patients versus healthy controls was significantly lower in median fluorescence intensity (MdFI)-E. coli and MdFI-PMA. We found significant correlations among MdFI-PMA and calculated MIS and other nutritional parameters in chronic HD patients. Impaired phagocytic function was identified in chronic HD patients. The severity of the impairment was associated with nutrition and inflammation parameters, as well as Malnutrition-Inflammation Score.

Differential diagnostics and correction of metabolic and immunological disorders in patients with hepatic cirrhosis, complicated hepatosplenomegalia and portal hypertension

The study of the role of the spleen in the pathogenesis of liver cirrhosis is important for treatment. Currently available literature data indicate the role of altered spleen functions in the pathogenesis of liver cirrhosis. The aim of this study was to clarify the features of the etiology and pathogenetic disorders in patients with hepatosplenomegaly. Materials and methods. We investigated the uniform elements of blood and blood serum from 58 patients with hepatosplenomegaly amid cirrhosis, portal hypertension complicated. Patients were divided into two groups depending on the antigenic load: I - patients with liver cirrhosis against the background of hepatitis viruses HBV/HCV and II - patients with autoimmune hepatitis against the background of herpes viruses CMV/VEB. Treatment of patients included the use of antiherpetic immunoglobulins and hematopoietic cord blood progenitor cells. Results. In patients of group I was detected an increase in the spontaneous oxidative activity of neutrophils and depletion of the oxidative reserve in oxygen-dependent phagocytosis. It activated the signaling pathways of pro-inflammatory cytokines IL-6 (an increase of 60 times). We found an increased content of CD22+ -plasma B-lymphocyte that elevated synthesized concentrations of IgA. In patients of group II were found an increase in the activity of C3 and a decrease in the C4-components of complement; thrombocytopenia due to the presence of platelet autoantibodies; increased synthesis of immunoglobulins IgM, IgG, increased synthesis of IFN-. All patients showed disturbances in the metabolism of amino acids in various metabolic cycles - ornithine, which disrupted the formation of urea and indicates the accumulation of ammonia, which can enhance the cytotoxic effect in group I; methionine with a violation of the formation of cystine from cysteine and the accumulation of homocysteine, which enhances the synthesis of IFN-γ, which was increased in group II by 4 times. Conclusions. Some etiological and pathogenetic features of the formation of liver cirrhosis complicated by portal hypertension and recurrent bleeding from varicose veins of the esophagus were revealed. The immunological and metabolic changes verified by us correlated with the disappearance of the clinical manifestations of the disease, improved well-being, and the absence of recurrence of bleeding.

Mesenchymal stem cells increase expression of heme oxygenase-1 leading to anti-inflammatory activity in treatment of acute liver failure

BackgroundMesenchymal stem cells (MSCs) have been studied for the treatment of acute liver failure (ALF) for several years. MSCs may exert their effect via complex paracrine mechanisms. Heme oxygenase (HO) 1, a rate-limiting enzyme in heme metabolism, exerts a wide range of anti-inflammatory, anti-apoptotic and immunoregulatory effects in a variety of diseases. However, the relationship between MSCs and HO-1 in the treatment of ALF is still unclear. We investigated the preventive and therapeutic potential of intravenously administered BMSCs.MethodsBone marrow-derived mesenchymal stem cells (BMSCs) obtained from Sprague–Dawley rats were isolated and cultured. We employed BMSCs, hemin (a HO-1 inducer) and zinc protoporphyrin (ZnPP, the HO-1 activity inhibitor) in D-galactosamine (D-Gal)/lipopolysaccharides (LPS)-induced ALF rats. Rats were sacrificed at days 1, 3, 5, and 7 post-transfusion, respectively. Blood samples and liver tissues were collected. Hepatic injury, HO-1 activity, chemokines, inflammatory cytokines, the number and oxidative activity of neutrophils, ki67, and TUNEL-positive cells were evaluated.ResultsHO-1 induction or BMSCs transplantation attenuated D-galactosamine/lipopolysaccharide-induced increases in alanine aminotransferase, aspartate aminotransferase, total bilirubin (TBIL), ammonia, and inflammatory cytokines. Treatment with hemin or BMSCs also inhibited neutrophil infiltration, oxidative activity, and hepatocyte apoptosis. The protective effect of BMSCs was partially neutralized by ZnPP, suggesting the key role of HO-1 in the process.ConclusionsThese findings may correlate with inhibition of nuclear factor-κ B activation. BMSCs ameliorated ALF by increasing the HO-1 expression, which reduced PMN infiltration and function, and played an important anti-inflammatory and anti-apoptotic role.Graphical Proposed mechanism by which BMSCs reduce inflammation, neutrophil activation, and hepatocyte apoptosis and promote hepatocyte proliferation via HO-1. BMSCs increase HO-1 expression in liver via Nrf2. HO-1 protects against LPS/D-Gal-induced ALF by inhibiting neutrophil infiltration and inflammatory burst, and hepatocyte apoptosis and necrosis. HO-1 also promotes hepatocyte proliferation.

Development of a wearable bioartificial kidney using the Bioartificial Renal Epithelial Cell System (BRECS).

Cell therapy for the treatment of renal failure in the acute setting has proved successful, with therapeutic impact, yet development of a sustainable, portable bioartificial kidney for treatment of chronic renal failure has yet to be realized. Challenges in maintaining an anticoagulated blood circuit, the typical platform for solute clearance and support of the biological components, have posed a major hurdle in advancement of this technology. This group has developed a Bioartificial Renal Epithelial Cell System (BRECS) capable of differentiated renal cell function while sustained by body fluids other than blood. To evaluate this device for potential use in end-stage renal disease, a large animal model was established that exploits peritoneal dialysis fluid for support of the biological device and delivery of cell therapy while providing uraemic control. Anephric sheep received a continuous flow peritoneal dialysis (CFPD) circuit that included a BRECS. Sheep were treated with BRECS containing 1×108 renal epithelial cells or acellular sham devices for up to 7days. The BRECS cell viability and activity were maintained with extracorporeal peritoneal fluid circulation. A systemic immunological effect of BRECS therapy was observed as cell-treated sheep retained neutrophil oxidative activity better than sham-treated animals. This model demonstrates that use of the BRECS within a CFPD circuit embodies a feasible approach to a sustainable and effective wearable bioartificial kidney. Copyright © 2016 John Wiley & Sons, Ltd.

Influence of progesterone on functional activity of neutrophils in threatening preterm labor

Aim of the study. To assess the influence of exogenous progesterone on the functional activity of neutrophils in patients, whose pregnancies proceeded with clinical signs of threatened abortion. Materials and methods. The material used for the study were peripheral blood neutrophils of 32 women, who were hospitalized with a diagnosis of "threatening premature labor" in the term of 22-34 weeks and whose pregnancies later ended in spontaneous preterm (group 1, 10 patients) and term (group 2, 22 patients) birth. Women of 1st and 2nd groups were comparable in age, gestational age and serum concentration of endogenous progesterone. The study was conducted in vitro. Neutrophils were isolated by settling for NBT-test and phagocytosis reaction, and by centrifugation on a double density gradient of ficoll-verografin in order to assess the formation of neutrophil extracellular traps. Neutrophils were added to a suspension of exogenous progesterone at concentrations of 10 ng/ml, 20 ng/ml, 100 ng/ml. The oxidation activity of neutrophils was measured in the spontaneous and stimulated (with Staphylococcus aureus at a concentration of 1×108 bacterial cells in 1 ml) types of NBT-test with microscopic evaluation of the results. The ability of neutrophils to form extracellular traps was evaluated in spontaneous and stimulated (with Staphylococcus aureus at a concentration of 1×108 bacterial cells in 1 ml) types of the test using color dye Hoechst 33342, followed by fluorescent microscopy. Results. Progesterone (10 ng/ml) significantly decreased spontaneous neutrophil oxidative activity [17.0% (12.0-26.0%) and 7.0% (4.0-10.0%), p<0.05] in patients of group 1, but not in patients of group 2. There was no effect of progesterone at studied concentrations on stimulated neutrophil oxidative activity in women of groups 1 and 2. When 20 ng/ml progesterone was added there was a decrease in neutrophil phagocytic index in patients of group 2 [66% (55-77%) and 51% (40-69%), p<0.05]. Progesterone (100 ng/ml) caused an increase in spontaneous NET-forming function in both groups and had no effect on stimulated function. Conclusions. Progesterone is able to provide various immunomodulatory effects on neutrophils of patients, whose pregnancies ended in preterm and term birth. Exogenous progesterone is able to provide various immunomodulatory effects on neutrophils of patients, whose pregnancies ended in preterm and term birth. It reduced the increased spontaneous oxidative activity of neutrophils in women with adverse pregnancy outcomes and lowered phagocytic number in patients with favorable pregnancy outcome. This drug also stimulated spontaneous NET-forming function of the cells, regardless of the pregnancy outcome.

The influence of Rubus idaeus and Rubus caesius leaf extracts on platelet aggregation in whole blood. Cross-talk of platelets and neutrophils

Recently, polyphenols have gained attention as potential natural cardioprotective therapeutics, due to their antiplatelet, anti-inflammatory and anticoagulant activity. Species belonging to the genus Rubus sp. have been reported to be a source of polyphenolic compounds with antioxidative proprieties and beneficial biological activities. This study investigates the effects of leaf extracts obtained from red raspberry (Rubus idaeus L.) and European dewberry (Rubus caesius L.) on the reactivity of blood platelets.In ADP-stimulated blood, raspberry and dewberry extracts (15 µg/ml) markedly decreased platelet surface membrane expression of activated GPIIbIIIa receptor by 16% and 21%, respectively (P < 0.01) and significantly inhibited platelet aggregation (by 31–41% for raspberry and by 38–55% for dewberry, P < 0.01). In platelet-rich plasma (PRP), the extracts had no effect on ADP-induced platelet aggregation.The effectiveness of the extracts in whole blood and the lack of their activity in PRP indicate that leukocytes are likely to participate in the platelet response to the extracts. Our experiments show that the extracts significantly reduced the amount of free radicals released by activated neutrophils in whole blood (P < 0.001), as well as in suspensions of isolated neutrophils (P < 0.05). Moreover, the reduced number of neutrophils leads to the decreased efficiency of the extracts in the inhibition of platelet aggregation. In summary, our findings show that the raspberry and dewberry leaf extracts considerably modulated blood platelet reactivity in whole blood: they influenced blood platelet aggregation, possibly via the modulation of the redox status dependent on the oxidative activity of neutrophils.

Effect of mixed culture supernatants of Pseudomonas aeruginosa and Escherichia coli on apoptosis, necrosis, and oxidative activity of neutrophils.

Effect of mixed culture supernatants of Pseudomonas aeruginosa and Escherichia coli on apoptosis, necrosis, and oxidative activity of neutrophils.

Adaptation to Resistance Training Is Associated with Higher Phagocytic (but Not Oxidative) Activity in Neutrophils of Older Women.

Failure in antimicrobial activity contributes to high morbidity and mortality in the geriatric population. Little is known about the potential effect of resistance training (RT) on the functional properties of the innate immunity. This study aimed to investigate the influence of long-term RT on the endocytic and oxidative activities of neutrophils and monocytes in healthy older women. Our results indicate that the phagocytosis index (PhI) of neutrophils (but not of monocytes) in the RT-adapted group was significantly higher (P < 0.001; effect size, (d) = 0.90, 95% CI: [0.75–1.04]) compared to that in sedentary subjects. In contrast, the oxidative activity of either neutrophils or monocytes was not significantly influenced by RT. Also, total energy and carbohydrate intake as well as serum IL6 levels had a significant influence on the phagocytic activity of neutrophils (P = 0.04), being considered in the model. Multivariate regression identified the physical condition of the subject (β = 0.425; P = 0.01) as a significant predictor of PhI. In conclusion, circulating neutrophils of older women adapted to a long-term RT program expressed higher phagocytic activity.

Innate immune response by oxidative activity of neutrophils in newborn calves supplemented with vitamin E

II SIMPÓSIO DE QUALIDADE DE LEITE O II Simpósio de Qualidade de Leite aconteceu nos dias 05, 06 e 07 de setembro de 2014, no Centro de Convenções da Faculdade de Ciências Agrárias e Veterinárias, Unesp, Câmpus de Jaboticabal, SP. Foi um evento organizado por pós-graduandos do Departamento de Medicina Veterinária Preventiva, sob a supervisão do Prof. Dr . Antonio Nader Filho (homenageado do V Congresso Brasileiro de Qualidade de Leite, 2013, pelos importantes trabalhos desenvolvidos na área). Tendo em vista o sucesso obtido na edição anterior, o II Simpósio de Qualidade de Leite apresentou uma nova programação, com palestras que abordaram assuntos atuais relacionados à cadeia do leite no Brasil aliados à presença de palestrantes renomados. Temas como fraudes e adulteração do leite; resíduos químicos; programas de autocontrole e boas práticas de fabricação; zoonoses veiculadas pelo leite; tratamento da mastite; manejo alimentar e ambiental do gado de leite; aproveitamento de dejetos; a importância da extensão rural como uma aliada; o controle estratégico de parasitas e análises sobre CBT e CCS, e uma mesa redonda para discutir a Instrução Normativa nº 62, do Ministério da Agricultura, Pecuária e Abastecimento, foram alguns dos principais assuntos abordados durante o evento. O público esperado envolveu profissionais de indústrias de laticínios, criadores de gado de leite, estudantes de graduação e de pós-graduação da área de qualidade de leite, pesquisadores e demais profissionais do segmento envolvidos com a pecuária leiteira. Outro aspecto inovador do Simpósio foi a apresentação de resumos expandidos de trabalhos científicos e de extensão rural relacionados ao tema de qualidade do leite. Os resumos estão publicados na Revista Brasileira de Higiene e Sanidade Animal , no formato de resumo expandido, o que permite maior visibilidade e divulgação das pesquisas relacionadas ao tema. Dessa forma, a Comissão Organizadora do II Simpósio de Qualidade de Leite, agradece a todos que de alguma forma colaboraram para o sucesso dessa edição. Em especial, agradecemos nossos patrocinadores, pelo apoio essencial ao evento e aos palestrantes, que foram de suma importância para o enriquecimento e sucesso do Simpósio. Muito obrigado a todos pela participação! Comissão Organizadora II Simpósio de Qualidade de Leite URL do Portal: http://www.higieneanimal.ufc.br/seer/index.php/index/index 2014 setembro; 8(5 Supl 1): p. 01- 552

Performance and immune response of suckling calves fed organic selenium

The objective of this research was to explore whether supplementation of Se above the recommended levels or application directly to the abomasum is superior to rumen supplementation in terms of boosting the immune system and promoting the growth of suckling calves. Thirty newborn calves, 18 Jersey breed and 12 crossbred (Holstein×Jersey), during 75 days, were assigned in a randomized block design (sex and breed) with the following treatments C=control (no supplementation and selenium derived only from milk and concentrate); SeA=0.80mg of Se/animal per day to the abomasum (milk); and SeR=0.80mg of Se/animal per day to the rumen (oral). The blood concentration of Se was higher in the SeR- and SeA-treated animals than in the control animals (P=0.001) at 30 days of age. The phagocytic activity of macrophages was higher in animals receiving Se supplementation compared to the control and in SeA animals compared to SeR (P=0.007). No difference in the oxidative activity of neutrophils or hematocrit was observed among treatments regimes when the calves were 30 days old. Dry matter intake (P=0.059) and HGG (P=0.059) tended to be higher in SeR-treated animals than in SeA-treated animals at 30 days of age. Feed conversion tended to be more efficient in Se-supplemented animals compared to the control (P=0.075). There were no significant differences in WG, HG or LG among treatment regimes at 30 days of age. At 75 days of age, Se-supplemented animals tended to present higher concentrations of Se in the blood (P=0.079) and greater heart girth gain (P=0.088). Supplementation of suckling calves with 0.80mg of organic Se increased the serum levels of this trace mineral in the animals and boosted their immune systems at 30 days of age. A superior immune parameter response was observed in calves supplemented with Se to the abomasum. In addition, Se supplementation maintained the performance of animals that had a diagnosis of diarrhea. Selenium supplementation did not act as a growth promoter but did improve immune system function during this phase of compromised health.

Genetically Determined Severity of Anti-Myeloperoxidase Glomerulonephritis

Myeloperoxidase (MPO) is a target antigen for antineutrophil cytoplasmic autoantibodies (ANCA). There is evidence that MPO-ANCA cause necrotizing and crescentic glomerulonephritis (NCGN) and vasculitis. NCGN severity varies among patients with ANCA disease, and genetic factors influence disease severity. The role of genetics in MPO-ANCA NCGN severity was investigated using 13 inbred mouse strains, F1 and F2 hybrids, bone marrow chimeras, and neutrophil function assays. Mouse strains include founders of the Collaborative Cross. Intravenous injection of anti-MPO IgG induced glomerular crescents in >60% of glomeruli in 129S6/SvEv and CAST/EiJ mice, but <1% in A/J, DBA/1J, DBA/2J, NOD/LtJ, and PWK/PhJ mice. C57BL6J, 129S1/SvImJ, LP/J, WSB/EiJ, NZO/HILtJ, and C3H mice had intermediate severity. High-density genotypes at 542,190 single nucleotide polymorphisms were used to identify candidate loci for disease severity by identifying genomic regions that are different between 129S6/SvEv and 129S1/SvImJ mice, which are genetically similar but phenotypically distinct. C57BL/6 × 129S6 F2 mice were genotyped at 76 SNPs to capture quantitative trait loci for disease severity. The absence of a dominant quantitative trait locus suggests that differences in severity are the result of multiple gene interactions. Invivo studies using bone marrow chimeric mice and invitro studies of neutrophil activation by anti-MPO IgG indicated that severity of NCGN is mediated by genetically determined differences in the function of neutrophils.

The influence of glycosaminoglycans on IL-8-mediated functions of neutrophils

Glycosaminoglycans (GAGs) of the extracellular matrix (ECM) contribute to the regulation of physiological processes by binding various immune-competent proteins. Due to their large structural diversity, the analysis of the binding properties and their functional consequences is challenging. The cytokine interleukin-8 (IL-8) is involved in the recruitment of neutrophils to inflammatory sites. Here, we investigated the interaction of heparin hexasaccharides and recombinant human IL-8, consisting of 77 amino acids using fluorescence and NMR spectroscopy. A dissociation constant of 2.0±0.4μM was determined for the heparin–IL-8 complex, which is slightly higher than what has been found for chondroitin-6-sulfate (KD=1.4±0.4μM) [Pichert, A.; Samsonov, S. A.; Theisgen, S.; Thomas, L.; Baumann, L.; Schiller, J.; Beck-Sickinger, A. G.; Huster, D.; Pisabarro, M. T. Glycobiology2012, 22, 134–145], suggesting an important role of the sulfate group at position 6 of the second ring in the disaccharide unit of the GAGs in this interaction. In addition, the influence of long-chain hyaluronan, chondroitin sulfate, and heparin on IL-8-induced chemotaxis and oxidative activity of neutrophils was examined. Only the incubation of heparin with IL-8 affected the IL-8-mediated chemotaxis of neutrophils. However, all investigated GAGs enhanced the IL-8-induced formation of reactive oxygen species in neutrophils, which is an entirely new finding. This work provides a representative example of how protein functions can be regulated by different GAGs of the ECM.

In vivo hydroquinone exposure alters circulating neutrophil activities and impairs LPS-induced lung inflammation in mice

Hydroquinone (HQ) is an environmental contaminant which causes immune toxicity. In this study, the effects of exposure to low doses of HQ on neutrophil mobilization into the LPS-inflamed lung were investigated. Male Swiss mice were exposed to aerosolized vehicle (control) or 12.5, 25 or 50ppm HQ (1h/day for 5 days). One hour later, oxidative burst, cell cycle, DNA fragmentation and adhesion molecules expressions in circulating neutrophils were determined by flow cytometry, and plasma malondialdehyde (MDA) levels were measured by HPLC. Also, 1h later the last exposures, inflammation was induced by LPS inhalation (0.1mg/ml/10min) and 3h later, the numbers of leukocytes in peripheral blood and in the bronchoalveolar lavage fluid (BALF) were determined using a Neubauer chamber and stained smears; adhesion molecules expressed on lung microvessel endothelial cells were quantified by immunohistochemistry; myeloperoxidase (MPO) activity was measured in the lung tissue by colorimetric assay; and cytokines in the BALF were determined by ELISA. In vivo HQ exposure augmented plasma MDA levels and oxidative activity of neutrophils, but did not cause alterations in cell cycle and DNA fragmentation. Under these conditions, the number of circulating leukocytes was not altered, but HQ exposure reduced LPS-induced neutrophil migration into the alveolar space, as these cells remained in the lung tissue. The impaired neutrophil migration into BALF may not be dependent on reduced cytokines secretions in the BALF and lung endothelial adhesion molecules expressions. However, HQ exposure increased the expression of β2 and β3 integrins and platelet-endothelial cell adhesion molecule-1 (PECAM-1) in neutrophils, which were not further enhanced by fMLP in vitro stimulation, indicating that HQ exposure activates circulating neutrophils, impairing further stimulatory responses. Therefore, it has been shown, for the first time, that neutrophils are target of lower levels of in vivo HQ exposure, which may be considered in host defense in infectious diseases.

Endotoxemia and the host systemic response during experimental gingivitis

To assess endotoxemia episodes and subsequent changes in serum inflammatory biomarkers using the experimental gingivitis model. Data from 50 healthy black and white adult males and females were compared for serum concentrations of endotoxin, and serum biomarkers [neutrophil oxidative activity, interleukin (IL)-1β, IL-6, IL-8, C-reactive protein (CRP), and fibrinogen] at baseline, at 3 weeks of experimental gingivitis, and after 2 weeks of recovery. Means were compared using repeated measures analysis of variance. Endotoxemia was reported in 56% of the serum samples at 3 weeks of induced gingivitis. At 2 weeks of recovery, endotoxin levels decreased to levels similar to those reported at baseline. Neutrophil oxidative activity increased significantly following 3 weeks of gingivitis versus baseline (p<0.05). In the endotoxin-negative group this increase was associated with the black subjects whereas in the endotoxin-positive group change in neutrophil activity was driven by the female subpopulation. Serum cytokines, CRP, and fibrinogen levels did not change during the study. Experimental gingivitis was associated with endotoxemia and hyperactivity of circulating neutrophils, but not with changes in systemic levels of cytokines and acute-phase proteins. This may be attributed to the mild nature and the short duration of the induced gingivitis.

Cystic fibrosis neutrophils have normal intrinsic reactive oxygen species generation

Previous studies have identified abnormalities in the oxidative responses of the neutrophil in cystic fibrosis (CF), but it is unclear whether such changes relate to loss of membrane cystic fibrosis transmembrane conductance regulator (CFTR) or to the inflammatory environment present in this disease. The aim of the present study was to determine whether neutrophils from CF patients demonstrate an intrinsic abnormality of the respiratory burst. The respiratory burst activity of neutrophils isolated from stable DeltaF508 homozygote CF patients and matched healthy controls was quantified by both chemiluminscence and cytochrome C reduction. Expression of NADPH oxidase components and CFTR was determined by Western blotting and RT-PCR. The oxidative output from neutrophils from CF in response to receptor-linked and particulate stimuli did not differ from that of controls. Expression of NADPH oxidase components was identical in CF and non-CF neutrophils. While low levels of CFTR mRNA could be identified in the normal human neutrophil, we were unable to detect CFTR protein in human neutrophil lysates or immunoprecipitates. CFTR has no role in controlling neutrophil oxidative activity; previously reported differences in neutrophil function between CF and non-CF subjects most likely relate to the inflammatory milieu from which the cells were isolated.

Role of Cyclooxygenase in the Chorionic Gonadotropin Regulation of Human Neutrophil Activity

The involvement of cyclooxygenase in the regulation of the main functions of human neutrophils by chorionic gonadotropin was studied. The inhibitory effect of the hormone on phagocytosis and oxidative activity of neutrophils is completely or partially canceled under conditions of cyclooxygenase blocking. In addition, gonadotropin directly induces the expression of cyclooxygenase inducible form mRNA in these cells. These data indicate that realization of the suppressive effects of chorionic gonadotropin in the neutrophils is partially linked with activation of cyclooxygenase (both the constitutive and inducible forms of the enzyme).