Oxygen Radicals and Arachidonate Metabolites in Lung Injury a

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/72476/1/j.1749-6632.1986.tb18477.x.pd

Prostacyclin protects ischemic reperfused myocardium in the dog by inhibition of neutrophil activation

Prostacyclin (PGI2) and the stable PGI2 analogue SC39902 (6,9[alpha]-epoxy, 5S-fluoro-11[alpha], 15S-dehydroxyprosta-6, 13E-dien-1-oic acid, sodium salt) were studied in anesthetized open-chest dogs subjected to 90 minutes of left circumflex coronary artery (LCCA) occlusion and 6 hours of reperfusion. PGI2 (50 ng/kg/min, infused into the left atrium) reduced infarct mass by 59% compared to control, but SC39902 (1.5 [mu]g/kg/min) failed to produce a significant reduction in infarct size. Both PGI2 and SC39902 reduced mean arterial blood pressure, heart rate, and rate-pressure product to the same extent. Regional myocardial blood flow measured with radiolabelled tracer microspheres did not demonstrate an increase in regional blood flow to the ischemic myocardium during the 90 minutes of LCCA occlusion in the PGI2 and control treatment groups. Canine neutrophils were isolated from whole blood and activated with opsonized zymosan. PGI2 produced a concentration-dependent inhibition of neutrophil activation as measured by superoxide production in vitro, whereas SC39902 failed to effectively inhibit neutrophil activation. Neutrophil migration into inflammatory skin lesions was effectively attenuated when dogs were pretreated with PGI2 (50 ng/kg/min, intravenously). Therefore, it is suggested that the cytoprotective effect of PGI2 during myocardial ischemia and reperfusion is related to an inhibition of neutrophil migration and the production of cytotoxic activated oxygen species.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/26892/1/0000458.pd

Nucleobindin Co-Localizes and Associates with Cyclooxygenase (COX)-2 in Human Neutrophils

The inducible cyclooxygenase isoform (COX-2) is associated with inflammation, tumorigenesis, as well as with physiological events. Despite efforts deployed in order to understand the biology of this multi-faceted enzyme, much remains to be understood. Nucleobindin (Nuc), a ubiquitous Ca2+-binding protein, possesses a putative COX-binding domain. In this study, we investigated its expression and subcellular localization in human neutrophils, its affinity for COX-2 as well as its possible impact on PGE2 biosynthesis. Complementary subcellular localization approaches including nitrogen cavitation coupled to Percoll fractionation, immunofluorescence, confocal and electron microscopy collectively placed Nuc, COX-2, and all of the main enzymes involved in prostanoid synthesis, in the Golgi apparatus and endoplasmic reticulum of human neutrophils. Immunoprecipitation experiments indicated a high affinity between Nuc and COX-2. Addition of human recombinant (hr) Nuc to purified hrCOX-2 dose-dependently caused an increase in PGE2 biosynthesis in response to arachidonic acid. Co-incubation of Nuc with COX-2-expressing neutrophil lysates also increased their capacity to produce PGE2. Moreover, neutrophil transfection with hrNuc specifically enhanced PGE2 biosynthesis. Together, these results identify a COX-2-associated protein which may have an impact in prostanoid biosynthesis

Impact of Anti-Inflammatory Agents on the Gene Expression Profile of Stimulated Human Neutrophils: Unraveling Endogenous Resolution Pathways

Adenosine, prostaglandin E2, or increased intracellular cyclic AMP concentration each elicit potent anti-inflammatory events in human neutrophils by inhibiting functions such as phagocytosis, superoxide production, adhesion and cytokine release. However, the endogenous molecular pathways mediating these actions are poorly understood. In the present study, we examined their impact on the gene expression profile of stimulated neutrophils. Purified blood neutrophils from healthy donors were stimulated with a cocktail of inflammatory agonists in the presence of at least one of the following anti-inflammatory agents: adenosine A2A receptor agonist CGS 21680, prostaglandin E2, cyclic-AMP-elevating compounds forskolin and RO 20-1724. Total RNA was analyzed using gene chips and real-time PCR. Genes encoding transcription factors, enzymes and regulatory proteins, as well as secreted cytokines/chemokines showed differential expression. We identified 15 genes for which the anti-inflammatory agents altered mRNA levels. The agents affected the expression profile in remarkably similar fashion, suggesting a central mechanism limiting cell activation. We have identified a set of genes that may be part of important resolution pathways that interfere with cell activation. Identification of these pathways will improve understanding of the capacity of tissues to terminate inflammatory responses and contribute to the development of therapeutic strategies based on endogenous resolution

Time-dependent inhibition of oxygen radical induced lung injury

Experimental acute lung injury mediated by reactive metabolites of oxygen can be inhibited by the antioxidant enzymes catalase and Superoxide dismutase (SOD). However, the specific time interval during which these enzymes must be present in order to cause protection is not well defined. Using two experimental models of oxidant-dependent acute lung injury, one involving the intratracheal injection of glucose, glucose oxidase, and lactoperoxidase and the other involving the intravenous injection of cobra venom factor (CVF), we investigated the effects of delaying antioxidant administration on the outcome of the inflammatory response. In both cases, the protective effects of catalase and SOD were rapidly attenuated when their administration was delayed for a short period of time. For example, intratracheal catalase resulted in 98% protection when given simultaneously with the glucose oxidase and lactoperoxidase, but only 13% protection when the catalase was delayed 4 min. Likewise, in the CVF-induced lung injury model, intravenous catalase resulted in 40% protection when given simultaneously with the CVF, but only 2% protection when the catalase was delayed 20 min, even though the peak of the injury occurred hours after the initiation of the injury. A similar time dependence was seen with SOD. These results indicate that antioxidant therapy is required early in the course of oxygen radical-mediated acute lung injury for effective protection.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44504/1/10753_2004_Article_BF00914272.pd

IL-17A Expression Is Localised to Both Mononuclear and Polymorphonuclear Synovial Cell Infiltrates

This study examines the expression of IL-17A-secreting cells within the inflamed synovium and the relationship to in vivo joint hypoxia measurements.IL-17A expression was quantified in synovial tissue (ST), serum and synovial fluid (SF) by immunohistochemistry and MSD-plex assays. IL-6 SF and serum levels were measured by MSD-plex assays. Dual immunofluorescence for IL-17A was quantified in ST CD15+ cells (neutrophils), Tryptase+ (mast cells) and CD4+ (T cells). Synovial tissue oxygen (tpO(2)) levels were measured under direct visualisation at arthroscopy. Synovial infiltration was assessed using immunohistochemistry for cell specific markers. Peripheral blood mononuclear and polymorphonuclear cells were isolated and exposed to normoxic or 3% hypoxic conditions. IL-17A and IL-6 were quantified as above in culture supernatants.IL-17A expression was localised to mononuclear and polymorphonuclear (PMN) cells in inflamed ST. Dual immunoflourescent staining co-localised IL-17A expression with CD15+ neutrophils Tryptase+ mast cells and CD4+T cells. % IL-17A positivity was highest on CD15+ neutrophils, followed by mast cells and then CD4+T-cells. The number of IL-17A-secreting PMN cells significantly correlated with sublining CD68 expression (r = 0.618, p<0.01). IL-17A SF levels correlated with IL-6 SF levels (r = 0.675, p<0.01). Patients categorized according to tp0(2)< or >20 mmHg, showed those with low tp0(2)<20 mmHg had significantly higher IL-17A+ mononuclear cells with no difference observed for PMNs. Exposure of mononuclear and polymorphonuclear cells to 3% hypoxia, significantly induced IL-6 in mononuclear cells, but had no effect on IL-17A expression in mononuclear and polymorphonuclear cells.This study demonstrates IL-17A expression is localised to several immune cell subtypes within the inflamed synovial tissue, further supporting the concept that IL-17A is a key mediator in inflammatory arthritis. The association of hypoxia with Il-17A expression appears to be indirect, probably through hypoxia-induced pro-inflammatory pathways and leukocyte influx within the joint microenvironment

Cationic polyelectrolytes: A new look at their possible roles as opsonins, as stimulators of respiratory burst in leukocytes, in bacteriolysis, and as modulators of immune-complex diseases (A review hypothesis)

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44497/1/10753_2004_Article_BF00915991.pd

Lipoteichoic acid-antilipoteichoic acid complexes induce superoxide generation by human neutrophils

Human neutrophils (PMNs) which have been incubated with lipoteichoic acid (LTA) from group A streptococci generated large amounts of Superoxide (O 2 − chemiluminescence and hydrogen peroxide when challenged with anti-LTA antibodies. Cytochalasin B further enhanced O 2 * generation. The onset of Of generation by the LTA-anti-LTA complexes was much faster than that induced by BSA-anti-BSA complexes. LTA-treated PMNs generated much less O 2 * when challenged with BSA complexes, suggesting that LTA might have blocked, nonspecifically, some of the Fc receptors on PMNs. PMNs treated with LTA-anti-LTA complexes further interacted with bystander nonsensitized PMNs resulting in enhanced Of generation, suggesting that small numbers of LTA-sensitized PMNs might recruit additional PMNs to participate in the generation of toxic oxygen species. Protelolytic enzyme treatment of PMNs further enhanced the generation of O 2 − by PMNs treated with LTA-anti-LTA. Superoxide generation could also be induced when PMNs and anti-LTA antibodies interacted with target cells (fibroblasts, epithelial cells) pretreated with LTA. This effect was also further enhanced by pretreatment of the target cells with proteases. PMNs incubated with LTA released lysosomal enzymes following treatment with anti-LTA antibodies. The amounts of phosphatase, Β -glucoronidase, N -acetylglucosaminidase, mannosidase, and lysozyme release by LTA-anti-LTA complexes were much smaller than those released by antibody or histone-opsonized streptococci, suggesting that opsonized particles are more efficient lysosomal enzyme releasers. However, since the amounts of O 2 − generated by the LTA complexes equaled those generated by the opsonized particles, it is assumed that the signals for triggering a respiratory burst and lysosomal enzyme secretion might be different.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/44498/1/10753_2004_Article_BF00914316.pd