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At least 883 records · Page 49Linked to original sources
PubMed2005

Proapoptotic effect of curcumin on human neutrophils: activation of the p38 mitogen-activated protein kinase pathway.

OBJECTIVE: Despite advances in the management of sepsis and acute respiratory distress syndrome, the mortality rate remains high. Delayed apoptosis of neutrophils is associated with multiple organ failure under those conditions. Thus, development of nontoxic neutrophil apoptosis regulating molecules may provide a novel therapeutic strategy. Curcumin is a promising dietary supplement for cancer prevention. However, the effect of curcumin on human neutrophil apoptosis remains unknown. We therefore hypothesized that curcumin would produce a proapoptotic effect on neutrophils. DESIGN: Prospective, controlled, and randomized in vitro study. SETTING: Research institute laboratory. SUBJECTS: Human peripheral neutrophils obtained from normal subjects. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: In the presence or absence of curcumin, both spontaneous neutrophil apoptosis and apoptosis of neutrophils following transmigration across a human lung endothelium-epithelium bilayer were studied by morphology and terminal dUTP nucleotide end labeling analyses, respectively. Myeloperoxidase activity and migration assays were performed to determine the impact of curcumin on neutrophil function. To elucidate the potential mechanism, the p38 mitogen-activated protein kinase pathway and caspase-3 activity were examined by Western blotting and enzymatic analyses. The data demonstrate that curcumin increased constitutive neutrophil apoptosis and abrogated the transbilayer migration-induced delay in neutrophil apoptosis. Neutrophil activation was reduced by curcumin treatment as evidenced by a decrease in migration and myeloperoxidase release. A marked increase in p38 phosphorylation and caspase-3 activity was observed following curcumin exposure. In addition, inhibition of p38 mitogen-activated protein kinase with SB203580 suppressed apoptosis and caspase-3 activation induced by curcumin. Thus, activation of p38 mitogen-activated protein kinase or an increase in caspase-3 activity appears to contribute to the proapoptotic effect of human neutrophil apoptosis by curcumin. CONCLUSION: The characteristics of curcumin, including its proapoptotic effect and antidegranulation effect, make it a potential candidate for the therapy of neutrophil-induced lung injury and sepsis.

Antineoplastic Agents↗
PubMed2005

Delayed neutrophil apoptosis attenuated by melatonin in human acute pancreatitis.

OBJECTIVES: This study evaluated the expression of neutrophil apoptosis and the effects of melatonin at different concentrations on delayed neutrophil apoptosis in different severities of acute pancreatitis in patients. METHODS: The study population was comprised of 10 patients with severe acute pancreatitis (SAP) and 10 with mild acute pancreatitis (MAP). A total of 10 mL of blood was drawn 24 hours after the onset of the clinical disease for isolation and incubation of the human neutrophils with 4 different concentrations of melatonin. Neutrophil apoptosis activity, CD18 expression, and respiratory burst activity were assessed with flow cytometry 12 hours after incubation. Another group of neutrophils from a healthy control group was used (n = 6) for comparison. RESULTS: Neutrophil apoptosis in patients with SAP is delayed compared with that of patients with MAP. Neutrophils from patients with SAP or MAP are functionally activated. Melatonin at concentrations of 10(-8), 10(-7), or 10(-6) M reverses the delayed process and enhances apoptosis activity in neutrophils in patients with MAP. Melatonin at concentrations of 10(-7) and 10(-6) M reverses the delayed process and increases apoptosis activity in neutrophils in patients with SAP. Neutrophils from patients with SAP and MAP showed significantly increased CD18 expression and respiratory burst activity. Melatonin at concentrations of 10(-7) or 10(-6) M reverses CD18 expression and respiratory burst activity in neutrophils in patients with SAP. CONCLUSIONS: This study highlights the importance of neutrophil apoptosis in patients with SAP and raises the possibility of a therapeutic strategy. Study data show that melatonin promotes neutrophil apoptosis in human acute pancreatitis.

Acute Disease↗
PubMed1993

Inhibition of neutrophil adherence to antibody by dapsone: a possible therapeutic mechanism of dapsone in the treatment of IgA dermatoses.

Dapsone is frequently effective in cutaneous diseases characterized by antibody deposition and accumulation of neutrophils. We hypothesized that this mechanism of action of dapsone may involve the inhibition of neutrophil adherence to antibody. The neutrophil adherence assay, which measures the binding of neutrophils to basement membrane zone-bound antibody on skin sections, was used to evaluate the effect of dapsone on neutrophil adherence to immunoglobulin A and immunoglobulin G. We evaluated the effect of dapsone on adherence of normal neutrophils to immunoglobulin A and immunoglobulin G from sera of linear immunoglobulin A bullous dermatosis and bullous pemphigoid patients, respectively. Linear immunoglobulin A bullous dermatosis or bullous pemphigoid antibody were bound to the basement membrane zone of normal skin sections as a substrate for the neutrophil adherence assay. Dapsone was added directly to the neutrophils or to the antibody source in concentrations of 0-50 micrograms/ml (pharmacologic range). Addition of dapsone to neutrophils produced an incremental inhibition of neutrophil adherence up to 75% at 50 micrograms/ml. Dapsone produced similar inhibition when added directly to the antibody itself, despite washing prior to usage in the neutrophil-adherence assay. Control specimens including irrelevant fractions of patient sera failed to demonstrate binding. Serum from a patient on dapsone therapy also showed inhibition of neutrophil adherence compared to the same patient on no therapy. We conclude that dapsone inhibits the adherence of neutrophils to basement membrane zone antibody in a dose-dependent manner. This may be related to an effect directly on antibody. This inhibition may contribute to the clinical efficacy of dapsone in antibody-mediated diseases.

Adult↗
PubMed2004

Abnormal LTC4 synthase RNA degradation in neutrophils from CML patients.

Neutrophils from patients with chronic myeloid leukaemia (CML) have an aberrant expression of leukotriene (LT)C4 synthase. In order to learn more about the regulation of this abnormality, LTC4 synthase mRNA expression was determined by reverse transcription polymerase chain reaction. A digoxigenin (DIG)-labelled LTC4 synthase RNA was synthesized and incubated in cytosolic extracts from CML neutrophils, normal neutrophils and eosinophils. LTC4 synthase mRNA was detected in total but not cytoplasmic RNA from normal neutrophils. In contrast, LTC4 synthase mRNA was found in the cytoplasm of CML neutrophils and in normal eosinophils, which also express the enzyme. The DIG-labelled LTC4 synthase RNA was, as opposed to normal neutrophils, degraded in cytosolic extracts from CML neutrophils. The degradation was time dependent and cell concentration dependent. Degradation was also seen in eosinophils, indicating that degradation of LTC4 synthase RNA was correlated to the expression of the protein. This study showed that the difference in expression of LTC4 synthase in normal and CML neutrophils was not because of a total lack of LTC4 synthase mRNA in normal neutrophils. However normal neutrophils lack, in contrast to CML neutrophils, LTC4 synthase mRNA in the cytoplasm. This discrepancy is not caused by a stabilized LTC4 synthase RNA in the cytosol of CML neutrophils. Instead an abnormal degradation of LTC4 synthase RNA was found in the cytosol of CML neutrophils.

Cytosol↗
PubMed2005

Kinetics of pulmonary neutrophil recruitment and clearance in a natural and spontaneously resolving model of airway inflammation.

BACKGROUND: Neutrophil apoptosis and phagocytic clearance have been proposed as key determinants affecting the resolution of airway inflammation. Objective To determine the kinetics of neutrophil priming, recruitment, activation and subsequent clearance in a naturally occurring equine disease model of neutrophilic pulmonary inflammation. METHODS AND RESULTS: A 5 h mouldy hay/straw challenge in hypersensitive horses induced transient pulmonary dysfunction lasting 4 days. At 24 h circulating neutrophils were primed and displayed delayed rates of spontaneous apoptosis in vitro. Neutrophil numbers in the airspaces peaked at 5 h and then fell abruptly, returning to pre-challenge levels by 4 days. Airspace neutrophils demonstrated increased respiratory burst activity compared with circulating cells and equine neutrophil elastase 2A concentrations increased in parallel with neutrophil numbers indicating in vivo priming and degranulation. The number of apoptotic neutrophils and proportion of alveolar macrophages containing phagocytosed apoptotic neutrophils increased significantly at 24 h and 4 days post-challenge corresponding to the period of most rapid neutrophil clearance. CONCLUSION: This is the first demonstration of spontaneous neutrophil apoptosis and phagocytic removal in a natural disease model of airway inflammation and provides critical kinetic data to support the hypothesis that this clearance pathway plays a central role in the resolution of neutrophilic inflammation.

Allergens↗
PubMed1991

Stimulation of neutrophil adherence to vascular endothelial cells by histamine and thrombin and its inhibition by PAF antagonists and dexamethasone.

1. In order to clarify the roles of platelet-activating factor (PAF) in histamine- and thrombin-induced neutrophil adhesion to vascular endothelial cells, the effects of several PAF antagonists were examined. The effects of the glucocorticoid dexamethasone were also examined in order to gain further insight into the anti-inflammatory actions of glucocorticoids. 2. In culture, histamine and thrombin stimulated the adherence of rat peritoneal neutrophils to human endothelial cells from the umbilical vein. They did not stimulate neutrophil adherence in the absence of endothelial cells, suggesting that the target cells for the histamine- and thrombin-induced adherence of neutrophils were endothelial cells, not neutrophils. 3. Several PAF antagonists, such as CV-3988, L-652,731 and Y-24,180 inhibited the histamine- and thrombin-induced neutrophil adherence in a concentration-dependent manner. Indomethacin failed to inhibit it. 4. Dexamethasone, a steroidal anti-inflammatory drug, did not inhibit the histamine- and thrombin-induced adherence of neutrophils to endothelial cells when the drug was present only during the 20 min incubation period for the adherence assay. When the endothelial cells were preincubated for 3 h with dexamethasone, the adherence of neutrophils to endothelial cells induced by histamine or thrombin was not inhibited. 5. When the neutrophils were preincubated for 3 h with dexamethasone, the histamine- and thrombin-induced adherence of neutrophils to endothelial cells was inhibited. 6. Our studies indicate that: (a) adherence of neutrophils to endothelial cells induced by histamine and thrombin is mediated by PAF production since PAF antagonists inhibited the adherence of neutrophils; and (b) neutrophils, not endothelial cells, are the target cells through which dexamethasone acts to inhibit adherence.

Animals↗
PubMed2003

Suppression of mitochondria-dependent neutrophil apoptosis with thermal injury.

Neutrophil apoptosis is delayed under trauma and/or sepsis conditions. The mechanism for the delay has remained unclear. We hypothesize that modulation of the mitochondrial pathway of apoptosis contributes to the delay in neutrophil apoptosis with burn injury. Rats were subjected to burn injury (30% of total body surface area, 98 degrees C for 10 s) and euthanatized 24 h postinjury. Blood neutrophils from sham and burn-injured rats were isolated by Ficoll gradient centrifugation and cultured for 2 or 8 h. Neutrophil apoptosis was determined by annexin V and propidium iodide (PI) labeling and flow cytometry. Neutrophil mitochondrial morphology was assessed via histochemical staining (MitoTracker GreenFM) and confocal microscopy. Neutrophils from rats with burn injury showed a decreased level of apoptosis compared with sham rat neutrophils at both 2 and 8 h of incubation. In incubated sham rat neutrophils, mitochondria showed a change from normal "tubular" to an "aggregated" morphology. In contrast, cultured neutrophils from burn rats did not exhibit this mitochondrial morphological transition until 8 h of incubation. Compared with sham rat neutrophils, neutrophils from burn rats showed decreased levels of active caspase-9 and -3. Whereas an upregulation of Bcl-xL and a downregulation of Bax seemed to contribute to decreased apoptosis in burn rat neutrophils at 2 h of incubation, the decreased apoptosis at 8 h appeared to be associated with a decrease in Bax and increased phosphorylated Bad. These data suggest that suppression of the mitochondrial pathway plays an essential role in the delay of polymorphonuclear neutrophil apoptosis with burn injury.

Animals↗
PubMed1988

Endotoxemia and neutrophil activation in vivo.

There is a growing body of data to suggest that marginated granulocytes mediate much of the pulmonary damage observed during endotoxemia. The mechanism(s) by which endotoxemia initiates neutrophil margination and cytotoxicity remain either controversial or unknown. The objectives of this study were 1) to determine the temporal relationship between endotoxin-induced decreases in mean arterial pressure and circulating neutrophils, 2) to monitor neutrophil activation in vivo by measuring myeloperoxidase (MPO) activity in the plasma and lymph, and 3) to assess the interaction between endotoxin and complement in activation of neutrophilic oxidative metabolism in vitro. We found that a bolus injection of endotoxin causes a concurrent decrease in both mean arterial pressure and circulating neutrophils at 2 min postinfusion. Blood pressure recovered to approximately 70% of control values by 180 min, whereas circulating neutrophils remain depressed at 20% of control values for the entire experimental period. Using MPO as a marker for neutrophil activation, we found that infusion of endotoxin produces a dramatic increase in plasma and lymph MPO activity, suggesting activation of neutrophilic metabolism in vivo. In vitro data showed that both endotoxin and plasma were required for optimal neutrophilic degranulation and superoxide formation. We conclude that 1) the appearance of MPO in the plasma (or lymph) may be a useful neutrophil marker for neutrophil activation in vivo and may prove useful in following the course of neutrophil-mediated tissue injury during endotoxemia, and 2) endotoxin-activated complement (C5a) activates neutrophils to produce cytotoxic oxidants.

Animals↗
PubMed2006

Functional and genomic changes induced by alveolar transmigration in human neutrophils.

Although the accumulation of neutrophils in the lungs and airways is common to many inflammatory lung diseases, including acute lung injury, the alterations that neutrophils undergo as they leave the peripheral circulation and migrate into the lungs have not been well characterized. Human volunteers were exposed to endotoxin by bronchoscopic instillation. The resulting air space neutrophil accumulation and peripheral blood neutrophils were isolated 16 h later, compared with circulating neutrophils isolated before or after to the pulmonary endotoxin exposure, and compared with circulating neutrophils exposed to endotoxin in vitro. Microarray analysis was performed on air space, circulatory, and in vitro endotoxin-stimulated neutrophils. Functional analysis included the determination of neutrophil apoptosis, chemotaxis, release of cytokines and growth factors, and superoxide anion release. Dramatic gene expression differences were apparent between air space and circulating neutrophils: approximately 15% of expressed genes have altered expression levels, including broad increases in inflammatory- and chemotaxis-related genes, as well as antiapoptotic and IKK-activating pathways. Functional analysis of air space compared with circulating neutrophils showed increased superoxide release, diminished apoptosis, decreased IL-8-induced chemotaxis, and a pattern of IL-8, macrophage inflammatory protein-1beta, monocyte chemoattractant protein-1, and tumor necrosis factor-alpha release different from either unstimulated or LPS-stimulated circulating neutrophils. Many of these changes are not elicited by in vitro treatment with endotoxin. Limited differences were detected between circulating neutrophils isolated before and 16 h after pulmonary endotoxin instillation. These results suggest that neutrophils sequestered in the lung become fundamentally different from those resident in the circulation, and this difference is distinct from in vitro activation with endotoxin.

Cell Movement↗
PubMed1999

Effects of endogenous and exogenous catecholamines on LPS-induced neutrophil trafficking and activation.

Endotoxemia produces elevations in catecholamine levels in the pulmonary and systemic circulation as well as rapid increases in neutrophil number and proinflammatory cytokine expression in the lungs. In the present experiments, we examined the effects of endogenous and exogenous adrenergic stimulation on endotoxin-induced lung neutrophil accumulation and activation. Levels of interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, and macrophage inflammatory protein (MIP)-2 mRNAs were increased in lung neutrophils from endotoxemic mice compared with those present in lung neutrophils from control mice or in peripheral blood neutrophils from endotoxemic or control mice. Treatment with the beta-adrenergic antagonist propranolol before endotoxin administration did not affect trafficking of neutrophils to the lungs or the expression of IL-1beta, TNF-alpha, or MIP-2 by lung neutrophils. Administration of the alpha-adrenergic antagonist phentolamine before endotoxemia did not alter lung neutrophil accumulation as measured by myeloperoxidase (MPO) levels but did result in significant increases in IL-1beta, TNF-alpha, and MIP-2 mRNA expression by lung neutrophils compared with endotoxemia alone. Administration of the alpha1-adrenergic agonist phenylephrine before endotoxin did not affect trafficking of neutrophils to the lungs but was associated with significantly increased expression of TNF-alpha and MIP-2 mRNAs by lung neutrophils compared with that found after endotoxin alone. In contrast, treatment with the alpha2-adrenergic agonist UK-14304 prevented endotoxin-induced increases in lung MPO and lung neutrophil cytokine mRNA levels. The suppressive effects of UK-14304 on endotoxin-induced increases in lung MPO were not affected by administration of the nitric oxide synthase inhibitor N-nitro-L-arginine methyl ester. These data demonstrate that the initial accumulation and activation of neutrophils in the lungs after endotoxemia can be significantly diminished by alpha2-adrenergic stimulation. Therapy with alpha2-adrenergic agents may have a role in modulating inflammatory pulmonary processes associated with sepsis-induced acute lung injury.

Adrenergic alpha-Agonists↗
PubMed2001

Neutrophil maturation and activation determine anatomic site of clearance from circulation.

The long-term disposition of circulating neutrophils and the site of disappearance from circulation remain unclear. We investigated neutrophil localization in mice using (111)In-labeled murine peripheral blood neutrophils, mature bone marrow neutrophils, and peritoneal exudate neutrophils to track in vivo localization of these different cell populations. Infused peripheral neutrophils were found to localize equally between liver and marrow sites by 4 h (31.2 +/- 1.9 vs. 31.9 +/- 1.8%), whereas exudate neutrophils predominantly localized to liver (42.0 +/- 1.1%) and marrow-derived neutrophils to the marrow (65.9 +/- 6.6%) where they were found to localize predominantly in the hematopoietic cords. Stimulation of marrow neutrophils before infusion caused a shift in localization from marrow to liver, and subsequent induction of an inflammatory site after infusion and marrow sequestration led to remobilization of infused marrow neutrophils but not of peripheral neutrophils. These results indicate that the marrow participates in removing neutrophils from circulation, with evidence supporting both storage and perhaps disposal functions. Furthermore, models for circulating neutrophil homeostasis should consider that the site of retention is governed by the maturation and activation states of the cell.

Actins↗
PubMed1984

Pulmonary neutrophil kinetics after thrombin-induced intravascular coagulation.

Previous studies have indicated that neutrophils are required for the development of increased lung vascular permeability after thrombin-induced pulmonary microembolization. In this study, we examined neutrophil kinetics and uptake in the sheep lung before and after lung vascular injury. Sheep neutrophils were isolated by a Percoll-gradient method and labeled with indium-111 oxine. A maximum lung activity of 40% of the injected indium-111 neutrophil activity was attained 8-12 min after the injection. The calculated half-lives of both circulating and pulmonary neutrophils were 700 min. The rate of washout of labeled neutrophils from the lungs was the same as the loss of the peripheral blood activity, indicating removal of neutrophils from the lung and blood by a common pathway (e.g., liver and spleen). Intravenous infusion of alpha-thrombin resulted in an immediate uptake of neutrophils of 14% above the base-line activity. The increased uptake was associated with an immediate decrease in the blood activity, indicating sequestration of the neutrophils in the pulmonary circulation. The neutrophil uptake after alpha-thrombin was transient, reaching a maximum 15 min after infusion. Neutrophil uptake did not occur with alpha-thrombin (which lacks the fibrinogen recognition site), suggesting that the uptake was secondary to intravascular coagulation. An increase in the pulmonary blood volume cannot explain the increased neutrophil sequestration because pulmonary blood volume determined by [99mTc]pertechnetate-labeled erythrocytes did not increase after the alpha-thrombin infusion. Therefore, alpha-thrombin results in a transient neutrophil sequestration in the lung, and the response is secondary to the intravascular coagulation induced by the alpha-thrombin.

Animals↗
PubMed1987

Neutrophil kinetics and lung injury.

The fact that the lung stores a large proportion of the marginated pool of neutrophils is a result of the anatomy of the pulmonary microcirculation. This capillary bed is made up of a network of a very large number of segments, many of which have smaller diameters than the neutrophils. Both neutrophils and erythrocytes must deform to pass through the capillary bed, but as the neutrophils are larger and less deformable, they pass through the segments more slowly. The slower movement of neutrophils through the pulmonary capillaries produces little obstruction to erythrocyte flow because the very large number of segments allows the erythrocytes to stream around those that are filled with neutrophils. The size of the marginated pool of neutrophils in the lung depends on the balance between forces that propel the neutrophils through the capillary segments and those that tend to retard them. Activation of the PMN both decreases their deformability and increases their adherence to endothelium, which results in increased numbers of neutrophils in the lung and a reduction in the number circulating in the blood. There is growing evidence that the adherence of the neutrophil to the endothelial cell is mediated by the CD omega 18 on the surface of the neutrophil. The absence of these proteins leads to lack of neutrophil adherence and migration, whereas factors that enhance adherence lead to increased expression of these glycoproteins. The fact that activated neutrophils can also damage normal tissue has led to several attractive hypotheses concerning the pathogenesis of lung diseases such as emphysema and the ARDS. Further elucidation of factors that cause cells to marginate in the pulmonary circulation and a more complete understanding of the factors that control their adherence to endothelium, migration into the interstitial and airspace, and phagocytic function will undoubtedly lead to a better understanding of both the physiology and pathology of the lung.

Animals↗
PubMed1989

Neutrophil function in ischemic heart disease.

Neutrophils contribute to the healing of and scar formation in myocardium after ischemic injury. Many recent studies indicate that neutrophils may be involved in the genesis and propagation of myocardial ischemia. To characterize neutrophil function in ischemic heart disease, neutrophil chemotaxis, leukotriene B4 (LTB4) generation, and elastase release in plasma were measured in 20 patients with stable angina, 17 patients with unstable angina or acute myocardial infarction (AMI), and 20 age-matched control subjects. Neutrophils from patients with stable angina exhibited markedly increased chemotactic activity and LTB4 generation as compared with the age-matched control subjects (p less than 0.01). Neutrophils of nine of 17 patients with unstable angina or AMI clumped spontaneously ex vivo and exhibited marked pseudopod formation and granule extrusion on electron microscopy. Subsequent chemotactic activity and LTB4 generation by neutrophils from these patients was less than in patients with stable angina, suggesting previous in vivo activation. Plasma levels of peptide B beta, a product of fibrin degradation by human neutrophil elastase, were approximately 15-fold higher (p less than 0.001) in patients with unstable angina or AMI (588 +/- 171 pmol/l, mean +/- SEM) compared with those in patients with stable angina (37 +/- 25 pmol/l) or control subjects (40 +/- 22 pmol/l), confirming intense in vivo neutrophil activation. Our study shows enhanced neutrophil function in patients with ischemic heart disease. The increased neutrophil chemotactic activity and LTB4 generation may be markers of stable angina pectoris. Intense neutrophil activation in unstable angina or AMI, as manifested by morphologic changes in neutrophils and elastase release, may relate to ongoing in vivo cellular activation.

Chemotaxis, Leukocyte↗
PubMed2005

Generation of reactive oxygen species by neutrophils and endothelial cell injury in normal and preeclamptic pregnancies.

The aim of this study was to investigate the role of neutrophil-derived reactive oxygen species on endothelial cell dysfunction in preeclampsia. We first assessed the correlation between nitrite and superoxide anion production in normal nonpregnant (n=10), normal pregnant (n=15), and preeclamptic women (n=12). We then examined neutrophil-mediated oxygen radical damage to human umbilical vein endothelial cells in vitro. Neutrophil superoxide release was measured by cytochrome C reduction; nitrite release was measured by the modified Griess reaction, and endothelial cell injury was measured by 51Cr release. N-formyl-methionyl-leucyl-phenylalanine-stimulated superoxide release by neutrophils was significantly increased in women with preeclampsia compared with the other 2 groups. Nitrite release by neutrophils was significantly decreased in preeclampsia compared with normal pregnancy. When neutrophils were pretreated with superoxide dismutase, nitrite release by neutrophils did not differ between normal pregnancy and preeclampsia, suggesting that excess superoxide anion in preeclampsia could reduce bioavailability of nitric oxide through neutrophil autocrine function. Neutrophil-mediated endothelial cell injury was significantly greater in women with preeclampsia than in the other 2 groups. Hydrogen peroxide was important in neutrophil-mediated endothelial cell injury in preeclampsia as catalase inhibited endothelial cell injury. When neutrophils were pretreated with NG-nitro-L-arginine methyl ester, neutrophil-mediated endothelial cell injury in preeclampsia was decreased, indicating a role for peroxynitrite formation as a mechanism of endothelial cell injury. In conclusion, the modulation of neutrophils causing superoxide production to dominate over nitrite release provides a reasonable explanation for endothelial cell dysfunction in preeclampsia.

Adult↗
PubMed1988

Fibrin is a determinant of neutrophil sequestration in the lung.

We investigated the role of fibrin in the sequestration of neutrophils in lungs challenged with microembolism. Lungs of chronically prepared awake sheep were imaged after infusion of 111In-neutrophils and 131I-fibrinogen. Thrombin (80 units/kg) was administered to induce fibrin sequestration in the lung. One group received tranexamic acid (4 mg/kg i.v.) to inhibit fibrinolysis, and the control group received a saline infusion. Thrombin infusion increased both 131I-fibrinogen and 111In-neutrophils in the lung, but the increases were greater and more prolonged in the fibrinolysis-inhibited group. We examined the adherence of isolated neutrophils to fibrin matrix to investigate how neutrophil-fibrin interactions could mediate neutrophil sequestration. Unstimulated neutrophils showed a 37.5 +/- 3.1% adherence to fibrin in contrast to a 3.9 +/- 0.4% neutrophil adherence to endothelium and a 6.5 +/- 0.5% adherence to agarose. Neutrophil adherence to fibrin increased to 81.7 +/- 6.2% with activation by phorbol myristate acetate (10(-8) M) and was inhibited by a monoclonal antibody directed against the beta-chain of the CD18 leukocyte adhesion glycoprotein. The findings indicate that retention of pulmonary fibrin microthrombi is associated with increased lung uptake of neutrophils. Fibrin serves as a substrate for neutrophil adherence, and this adherence is mediated by expression of the CD18 glycoprotein complex on neutrophils. Fibrin sequestration in the lung may contribute to lung vascular injury by inducing pulmonary neutrophil uptake.

Animals↗
PubMed2004

Pulmonary neutrophil infiltration in murine sepsis: role of inducible nitric oxide synthase.

Nitric oxide (NO) derived from inducible NO synthase (iNOS) contributes to the pathophysiology of acute lung injury (ALI). The effect of iNOS on pulmonary neutrophil infiltration in ALI is not known. Thus, we assessed pulmonary microvascular neutrophil sequestration through intravital videomicroscopy and pulmonary neutrophil infiltration, reflected by myeloperoxidase activity and lavage neutrophil counts, after induction of sepsis by cecal ligation/perforation in wild-type (iNOS+/+) versus iNOS-/- mice. Pulmonary microvascular neutrophil sequestration was attenuated in septic iNOS-/- versus iNOS+/+ mice (15 +/- 1 vs. 20 +/- 1 leukocytes per field, p < 0.05), but lavage neutrophil counts were greater in iNOS-/- mice (5.7 +/- 1.5% vs. 0.7 +/- 0.1%, p < 0.05) between 6 and 18 hours after cecal ligation and perforation. When iNOS+/+ bone marrow was transplanted into bone marrow-depleted iNOS-/- mice (+ to - chimeras; iNOS limited to marrow-derived inflammatory cells), septic pulmonary microvascular neutrophil sequestration and lavage neutrophil counts were restored to levels seen in septic iNOS+/+ mice. In contrast, in - to + chimeras, pulmonary neutrophil trafficking was similar to iNOS-/- mice. In vitro cytokine-stimulated neutrophil transendothelial migration was significantly greater for iNOS-/- versus iNOS+/+ neutrophils (7.9 +/- 0.7% vs. 3.8 +/- 0.6%, p < 0.05) but was independent of endothelial iNOS. Thus, neutrophil iNOS-derived NO is an important autocrine modulator of pulmonary neutrophil infiltration in murine sepsis.

Acute Disease↗
PubMed2006

Down-regulation of CXCR2 on neutrophils in severe sepsis is mediated by inducible nitric oxide synthase-derived nitric oxide.

RATIONALE: The failure of neutrophils to migrate to an infection focus during severe sepsis is an important determinant of the inability of a host to deal with an infectious insult. Our laboratory has shown that inducible nitric oxide synthase (iNOS) induction and NO production contribute to the failure of neutrophils to migrate in the context of sepsis. OBJECTIVES AND METHODS: We investigated whether CXCR2 expression contributed to the failure of neutrophils to migrate during severe sepsis and the role of NO in modulating CXCR2 expression on neutrophils in mice subjected to nonsevere (NS) or severe (S) cecal ligation and puncture (CLP). RESULTS: Neutrophil migration to the infection focus was deficient in S-CLP mice, a phenomenon prevented by pharmacologic (aminoguanidine, l-canavanine) or genetic (iNOS gene deletion) inhibition of iNOS. The expression of CXCR2 on neutrophils from S-CLP mice was significantly reduced when compared with neutrophils from NS-CLP or sham-operated mice. CXCR2 expression was reestablished by pharmacologic and genetic inhibition of iNOS. Immunofluorescence and confocal analysis revealed that iNOS blockade reduced neutrophil CXCR2 internalization. Adhesion and emigration of neutrophils in macrophage inflammatory protein-2-stimulated mesentery microcirculation were reduced in S-CLP mice, compared with NS-CLP mice, and reestablished by pretreatment with aminoguanidine or l-canavanine. The NO donor S-nitroso-N-acetyl-d,l-penicillamine inhibited CXCL8-induced human neutrophil chemotaxis and CXCR2 expression on human and murine neutrophils. CONCLUSION: These results highlight evidences that the failure of neutrophils to migrate to an infection focus during severe sepsis is associated with excessive NO production and NO-dependent regulation of the expression of CXCR2 on the neutrophil surface.

Animals↗