Decrease In L-selectin Expression Research Articles

Artificial shear stress effects on leukocytes at a biomaterial interface.

Medical devices, such as ventricular assist devices (VADs), introduce both foreign materials and artificial shear stress to the circulatory system. The effects these have on leukocytes and the immune response are not well understood. Understanding how these two elements combine to affect leukocytes may reveal why some patients are susceptible to recurrent device-related infections and provide insight into the development of pump thrombosis. Biomaterials-DLC: diamond-like carbon-coated stainless steel; Sap: single-crystal sapphire; and Ti: titanium alloy (Ti6 Al4 V) were attached to the parallel plates of a rheometer. Whole human blood was left between the two discs for 5minutes at +37°C with or without the application of shear stress (0s-1 or 1000s-1 ). Blood was removed and used for complete blood cell counts, flow cytometry (leukocyte activation, cell death, microparticle generation, phagocytic ability, and reactive oxygen species [ROS] production), and the production of pro-inflammatory cytokines. L-selectin expression on monocytes was decreased when blood was exposed to the biomaterials both with and without shear. Applying shear stress to blood on a Sap and Ti surface led to activation of neutrophils shown as decreased L-selectin expression. Sap and Ti blunted the LPS-stimulated macrophage migration inhibitory factor (MIF) production, most notably when sheared on Ti. The biomaterials used here have been shown to activate leukocytes in a static environment. The introduction of shear appears to exacerbate this activation. Interestingly, a widely accepted biocompatible material (Ti) utilized in many different types of devices has the capacity for immune cell activation and inhibition of MIF secretion when combined with shear stress. These findings contribute to our understanding of the contribution of biomaterials and shear stress to recurrent infections and vulnerability to sepsis in some VAD patients as well as pump thrombosis.

Sphingosine-1-phosphate increases human alveolar epithelial IL-8 secretion, proliferation and neutrophil chemotaxis

Sphingosine-1-phosphate (S1P) has been presented recently as a pro-inflammatory agent in the airway epithelium since S1P levels are increased in bronchoalveolar lavage fluid of human asthmatics. However, the effects of S1P over the alveolar epithelium and neutrophil interactions are poorly understood. Here, we show that S1P increased interleukin 8 (IL-8) gene expression and protein secretion and proliferation in alveolar epithelial cells A549 at physiological concentrations (1 μM). At the same time, S1P increased intracellular Ca 2+ concentration (potency 17.91 μM, measured by epifluorescence microscopy), phospholipase D (PLD) activity (measured by chemiluminiscence method) and extracellular matrix-regulated kinase1/2 (ERK1/2) phosphorylation (measured by western blot) via G i-coupled receptor (inhibited by pertussis toxin 100 ng/ml) in A549 cells. Both, IL-8 secretion and A549 proliferation were dependent of PLD activity (inhibited by 1-butanol 0.5%), intracellular Ca 2+ (inhibited by acetoxymethyl 1,2-bis(2-aminophenoxy)-ethane- N, N, N′, N′-tetraacetic acid (BAPTA-AM) 100 μM), ERK1/2 phosphorylation (inhibited by 2-[2-amino-3-methoxyphenyl]-4H-1-benzopyran-4-one (PD98059) 10 μM) and G i-coupled receptors (blocked by pertussis toxin 100 ng/ml). Moreover, S1P increased intercellular adhesion molecule I (ICAM-1) expression and failed in vascular cell adhesion molecule I (VCAM-1) modification (measured by flow cytometer) in A549. Indirectly, A549 supernatant fluids arising from A549-S1P 1 μM stimulation decreased L-selectin expression without CD11b/CD18 integrin modification in human neutrophils. In the same way, A549-S1P supernatant fluids increased neutrophil chemotaxis (Boyden chamber), which was inhibited by antibody against IL-8. This study demonstrates for the first time that S1P participates in the alveolar epithelial interactions in vitro.

Course of Platelet Activation and Platelet-Leukocyte Interaction in Cerebrovascular Ischemia

Platelet activation plays a crucial role in the pathophysiology of cerebral ischemia. The aim of this study was to investigate the contribution of platelet activation and leukocyte-platelet interactions to the disease. One hundred thirty-five patients with transient ischemic attack (TIA) or stroke were enrolled in this single-center study. They underwent cranial computer tomography within 24 hours of clinical onset and after 3 months, and systemic venous blood samples were drawn. Platelet activation (CD62P expression), leukocyte activation (L-selectin expression), and the appearance of platelet-specific antigens on leukocytes as an index of platelet-leukocyte aggregation were measured by flow cytometric techniques in the acute state and at 3-month follow-up. Patients with a completed stroke or TIA had significantly increased circulating platelet-leukocyte aggregates, increased P-selectin expression on platelets, and decreased L-selectin expression in the acute state compared with the control group (healthy volunteers). No differences in regard to the tested activation markers could be detected between patients with stroke or TIA in the acute phase of the disease. However, platelet and leukocyte activations were normalized after 3 months in patients with TIA, whereas leukocyte activation (reduced L-selectin expression) remained in stroke patients. In patients with TIA and completed stroke, platelet and leukocyte activation is substantially enhanced in the acute phase of the disease. The sustained leukocyte activation observed in stroke but not in TIA patients at 3-month follow up might play a pathophysiological role in the course of the disease.

Platelet binding to monocytes increases the adhesive properties of monocytes by up-regulating the expression and functionality of β1 and β2 integrins.

Human monocytes adhere to activated platelets, resulting in the formation of platelet-monocyte complexes (PMC). Complex formation depends on the interaction between platelet-displayed P-selectin and the specific ligand for P-selectin on leukocytes, P-selectin glycoprotein ligand-1 (PSGL-1). We have recently shown that monocytes within PMC have increased adhesive capacity to the activated endothelium. To better understand the effect of platelet binding on the capacity of monocytes to adhere to activated endothelium, the P-selectin-PSGL-1 interaction-induced changes in integrin functionality were studied. The binding of platelets to monocytes via P-selectin-PSGL-1 interactions was shown to increase expression and activity of α4 β1 and αM β2 integrin, with a concomitant decrease in L-selectin expression. Furthermore, the binding of platelets to monocytes resulted in increased monocyte adhesion to intercellular adhesion molecule-1, vascular cell adhesion molecule-1, and fibronectin. Platelet binding was also responsible for an increase in monocyte transendothelial migration. Similar effects were observed after engagement of PSGL-1 with specific antibodies or with P-selectin immunoglobulin protein. Our data suggest that platelets, by binding via P-selectin to PSGL-1 on monocytes, induce up-regulation and activation of β1 and β2 integrins and increased adhesion of monocytes to activated endothelium. Hence, monocytes within PMC are in a higher state of activation and may have, therefore, an increased atherogenic capacity.

Phosphodiesterase type 4 inhibitor rolipram inhibits activation of monocytes during extracorporeal circulation

Cardiopulmonary bypass is associated with systemic inflammatory response syndrome and risk of multiorgan injury mediated by activated leukocytes. Phosphodiesterase type 4 is the predominant phosphodiesterase isozyme in leukocytes and plays a key role in the regulation of leukocyte activation. The aim of this study was to examine the effect of rolipram, a selective phosphodiesterase type 4 inhibitor, on functional changes of monocytes during simulated extracorporeal circulation. Simulated extracorporeal circulation was established by recirculating heparinized human blood for 120 minutes on a membrane oxygenator with or without 10 micromol/L of rolipram. L-selectin and CD11b expression of monocytes were measured with flow cytometry. C4d fragment, Bb fragment, C5b-9, and interleukin-6 were measured with enzyme immunoassay. Rolipram reduced the increase in CD11b expression and the decrease in L-selectin expression of monocytes in response to simulated extracorporeal circulation. Rolipram inhibited the increase in C4d fragment and interleukin-6, but it did not affect the increase in Bb fragment or C5b-9. Rolipram inhibited changes in adhesion molecule expression and interleukin-6 release by activated monocytes in simulated extracorporeal circulation. This study suggests that phosphodiesterase type 4 inhibition could be feasible therapeutic strategy to prevent exaggerated inflammatory response and organ injury in patients undergoing cardiopulmonary bypass.

Diazoxide ameliorates microcirculatory disturbances through PKC-dependent Pathway in I/R-injured rat cremaster muscles

Diazoxide is a selective mitochondria ATP-sensitive potassium (K(ATP)) channel opener, which has been reported to preserve the microvascular integrity of ischemia-reperfusion (I/R)-injured tissues. Our study aimed to assess diazoxide's effects on I/R-injured cremaster muscles and to further elucidate its underlying mechanisms. Male Sprague Dawley (SD) rats were randomized (n = 8 per group) into four groups: sham-operated control group, I/R group (4 h of pudic epigastic artery ischemia followed by 2 h of reperfusion), diazoxide + I/R group, and chelerythrine (PKC inhibitor)+diazoxide+I/R group. Microscopically, we observed that I/R markedly increased the number of rolling, adhering, and transmigrating leukocytes. I/R also markedly decreased the number of functional capillaries. Biochemically, we found that I/R significantly increased TNF-alpha, E-selectin,L-selectin and P-selectin expressions. However, I/R did not cause significant changes in ICAM-1 and PECAM-1 expressions. On the other hand, in I/R + diazoxide group, we found that diazoxide reduced the number of rolling, adhering, and transmigrating leukocytes. Furthermore, biochemical study revealed that diazoxide caused only a decrease in L-selectin expression but had no effect on TNF-alpha, E-selectin, P-selectin, ICAM-1, and PECAM-1 expressions. Finally, in chelerythrine + diazoxide + I/R group, we observed that diazoxide's protective effects were blocked by the addition of chelerythrine. Diazoxide's ability to protect against I/R injury was confirmed by the observation that it reduced the number of rolling, adhering, and transmigrating leukocytes, and increased the number of functional capillaries. Our results indicated that diazoxide operated via a PKC-dependent pathway to achieve protection against I/R injury.

A marked decrease in L-selectin expression by leucocytes in infants with Bordetella pertussis infection: leucocytosis explained?

Infants with Bordetella pertussis infection (whooping cough) have an unexplained lymphocytosis and leucocytosis characterized by an increase in small lymphocytes with convoluted and cleaved nuclei. To characterize these cells immunophenotyping using multiparameter flow cytometry was performed on leucocytes from a group of 11 infants aged 3-6 months with proven pertussis and from uninfected control subjects. The panel of monoclonal antibodies used to elucidate leucocyte subtypes included activation, adhesion, costimulatory, memory, T-helper (Th) 1 and Th2 markers. Patients with pertussis showed an increase in absolute numbers of neutrophils, monocytes, T lymphocytes (both CD4 and CD8), B lymphocytes (including CD10+/CD19+ haematogones) and natural killer (NK) cells. All leucocyte subgroups showed a marked decrease in L-selectin (CD62L) expression. The expression of other adhesion molecules CD11a, CD44 and CD54 on all leucocyte subgroups was unchanged. Expression of costimulatory molecules, CD49D and CD28 on T cells and CD80 and CD86 on monocytes, was unchanged. Lymphocyte activation markers CD69, CD25 and HLA-DR were unchanged. There was an increase in CD45RA+/CD45RO+/CD4+ cells (activated) and CD62L-/CD45RO+/CD4+ cells (Th1-like) but no increase in CD7-/CD4+ T cells (Th2-like). L-Selectin expression mediates extravasation of leucocytes into tissues and is important for homing of peripheral blood lymphocytes to lymph nodes. The significant down-regulation of L-selectin on leucocytes in pertussis infection may prevent leucocyte migration to areas of infection and homing and adhesion of T and B cells to peripheral lymphoid tissues. The increase in lymphocytes with Th1 phenotype may be required for effective immune response to the infective organism. These data provide a possible explanation for the absolute leucocytosis observed in this disease.

Effect of recombinant human deoxyribonuclease on the expression of cell adhesion molecules of thawed and processed cord blood hematopoietic progenitors.

The integrity of granulocytic cells and platelets is compromised within cryopreserved stem cell transplants, and consequent DNA release during the thawing procedure can therefore lead to clotting phenomena or microaggregate formation and that in turn may cause loss of progenitor cells. To circumvent this problem a new processing protocol was introduced using recombinant human deoxyribonuclease (rhDNase) to prevent cell aggregate formation. In addition, the impact of this new processing protocol on CD34+ umbilical cord blood (UCB) cells was assessed. Fifty samples derived from 7 buffy coat (BC) volume reduced UCB units were cryopreserved, thawed, and processed with washing solutions that were supplemented with rhDNase in various concentrations. Thereafter, clotting and microaggregate formation was scored microscopically. In addition, expression of the adhesion molecules leukocyte function-associated antigen 1 (LFA-1, n = 6), intercellular adhesion molecule-1 (ICAM-1, n = 11), and L-selectin (n = 11) on CD34+ UCB cells was analyzed by flow cytometry after incubating the samples with either dimethyl sulfoxide (DMSO) 5.5%, rhDNase 10 or 50 U/mL, or a combination of DMSO 5.5% and rhDNase 50 U/mL. At a minimal concentration of 10 U rhDNase/mL, clotting or microaggregate formation could be prevented for all tested samples, whereas cell clots could be observed for concentrations up to 8 U/mL. The expression of adhesion molecules on untreated CD34+ UCB cells (L-selectin: 64.6 +/- 18.8%; LFA-1: 62.6 +/- 7.5%; ICAM-1: 14.8 +/- 4.1%) did not show any significant difference compared with cells that were incubated with up to 50 U/mL rhDNase (L-selectin: 62.2 +/- 19.3%; LFA-1: 63.1 +/- 5.9%; ICAM-1: 17.5 +/- 6.7%). However, after a combined treatment with DMSO 5.5% and rhDNase 50 U/mL, a slight but significant decrease in L-selectin expression could be observed (P < 0.03). The supplementation of rhDNase to a final concentration of 10 U/mL cell suspension proved to be effective in preventing clot formation under the conditions examined and did not lead to decreased expression levels of adhesion molecules. We therefore recommend the use of rhDNase for the prevention of clot formation and cell loss during the processing of thawed UCB transplants.

Modulación de la expresión de la L-selectina por agentes quimiotácticos y GM-CSF

L-selectin is an adhesion molecule with constitutive expression located on the membrane of granulocytes, monocytes and lymphocytes. It is involved in the early stages of migration of these cells toward either the sites of inflammation or lymphoid tissues. After the cells are activated, L-selectin is down-regulated with shedding of a soluble fragment. Flow cytometry was used to measure L-selectin expression levels on the granulocyte surface, after incubation with a phorbol esther (PMA), two chemotactic factors (fMLP and LTB4) and a cytokine (GM-CSF). Under basal conditions, the expression of L-selectin was found in a high percentage (95.0 +/- 0.7) of granulocytes; PMA stimulation led to a marked decrease in expression (3.2 +/- 0.6). Chemotactic factors also led to a significant decrease in L-selectin expression (69.9 +/- 5.0 for LTB4, and 53.70 +/- 4.3 for fMLP), whereas the incubation with GM-CSF produced no significant changes (89.1 +/- 4.8). When all the conditions were compared, the PMA effect was significantly higher than those observed with other stimuli; furthermore, the expression upon incubation with fMLP and LTB4 was statistically significant. These results suggest that the level of activation reached by granulocytes is directly related to their capacity for shedding L-selectin from the cell surface, and that these levels are lowered after the stimulation by chemotactic factors. GM-CSF activates several important functions of granulocyte cells, however it does not induce L-selectin shedding.

Decreased Expression of L-Selectin on Peripheral Blood Polymorphonuclear Leukocytes in Neonates with Severe Asphyxia

The expression of adherent leukocyte surface molecules, L-selectin (CD62L), and Mac-1 (CD11b/CD18) in peripheral blood polymorphonuclear leukocytes (PMNs) was studied in 48 neonates and 24 healthy adults using fluorescence flow cytometry. L-selectin expression was decreased significantly in neonates, especially those with severe asphyxia (n = 10) compared with adults. Mac-1 expression was significantly increased in neonates compared with adults, but did not differ between 24 normal neonates, 14 neonates with mild asphyxia, and 10 neonates with severe asphyxia. These results suggest that the magnitude of the decrease in L-selectin expression reflects the severity of asphyxia in neonates.

Elevated serum L-selectin levels and decreased L-selectin expression on CD8(+) lymphocytes in systemic sclerosis.

L-selectin is expressed on most circulating leucocytes and mediates leucocyte rolling on endothelium at sites of inflammation. Following rolling or activation of leucocytes, cell surface L-selectin is released as soluble L-selectin (sL-selectin). In the present study, we assessed serum levels of sL-selectin by ELISA and blood leucocyte L-selectin expression by flow cytometry in patients with systemic sclerosis (SSc). Serum levels of sL-selectin in patients with SSc (n = 51) were significantly higher than in normal controls (n = 30) while sL-selectin levels were similar for systemic lupus erythematosus patients (n = 20) and normal controls. Furthermore, SSc patients with elevated sL-selectin levels had inflammatory joint involvement, pitting scar/ulcers, and diffuse pigmentation more frequently than those with normal sL-selectin levels. The frequency of L-selectin(+) population among CD8(+) T cells was significantly decreased in SSc patients (n = 30) compared with normal controls (n = 20), while that among CD4(+) T cells, B cells, monocytes, and neutrophils was similar for SSc patients and normal controls. These suggest that elevated sL-selectin levels and decreased frequency of L-selectin+ CD8(+) T cells in SSc patients may be involved in inflammation associated with SSc.

A Potential Role for Annexin 1 as a Physiologic Mediator of Glucocorticoid-Induced L-Selectin Shedding from Myeloid Cells

Glucocorticoids can dampen inflammatory responses by inhibiting neutrophil recruitment to tissue sites. The detailed mechanism by which glucocorticoids exert this affect on neutrophils is unknown. L-selectin is a leukocyte cell surface receptor that is implicated in several steps of neutrophil recruitment. Recently, several studies have shown that systemic treatment of animals and humans with glucocorticoids induces decreased L-selectin expression on neutrophils, suggesting one mechanism by which inflammation may be negatively regulated. However, when neutrophils are treated in vitro with glucocorticoids, no effect on L-selectin expression is observed. Thus, the existence of an additional mediator is plausible. In this study, we investigate whether annexin 1 (ANX1), a recognized second messenger of glucocorticoids, could be such a mediator. We show that ANX1 induces a dose- and time-dependent decrease in L-selectin expression on both peripheral blood neutrophils and monocytes but has no effect on lymphocytes. The loss of L-selectin from neutrophils is due to shedding that is mediated by a cell surface metalloprotease ("sheddase"). Using cell shape and a beta(2) integrin activation epitope, we show that the ANX1-induced shedding of L-selectin appears to occur without overt cell activation. These data may provide the basis for further understanding of mechanisms involved in the down-regulation of inflammatory responses.

Adhesion receptors of polymorphonuclear granulocytes on titanium in contact with whole blood

Titanium sheets were exposed to whole blood, and the TiO2 surface was investigated regarding the presence of cells, receptor expression on adherent polymorphonuclear (PMN) granulocytes, and the ability of these cells to mount a respiratory burst when challenged with opsonized zymosan. The techniques used were immunofluorescence with computer-aided image analysis and chemiluminescence. Surface coverage of erythrocytes (9% to 10%), granulocytes (9% to 14%), and platelets (1% to 4%) dominated during the first 2 hours of blood contact. PMN granulocyte adhesion to titanium was associated with a rapid decrease in L-selectin expression within 16 minutes. Initially FcγIII receptor (CD16) expression dominated on the adherent cells. After 30 minutes, a shift toward integrin expression (CD11b) was found on the adherent cells. All investigated receptors were down-regulated within 1 hour of blood-titanium contact. Attempts were made to inhibit the initial adhesion of PMN granulocytes to titanium by adding specific antibodies or 2,3-diphosphoglyceric acid (phospholipase D inhibitor) to blood before surface contact. Adding anti-CD16 resulted in a 67% reduction in cell adhesion, whereas a 35% reduction was found with 2,3-diphosphoglyceric acid. No spontaneous respiratory burst was detected from adherent PMN granulocytes residing on the TiO2 surface. The cells were, however, able to mount a respiratory burst in response to opsonized zymosan. (J Lab Clin Med 2001;137:56-63)

Interleukin-6 induces demargination of intravascular neutrophils and shortens their transit in marrow.

Recombinant human interleukin-6 (IL-6) causes both a thrombocytosis and leukocytosis. The thrombocytosis is caused by an accelerating thrombocytopoiesis, but the mechanism of the leukocytosis is unknown. This study was designed to determine the relative contributions of marrow stimulation and intravascular demargination to the IL-6 induced neutrophilia. IL-6 (2 microgram/kg), administered intravenously to rabbits, caused a biphasic neutrophilia with an initial peak at 3 h and a second peak at 9 h. Using the thymidine analog 5'-bromo-2'-deoxyuridine (BrdU) to label dividing polymorphonuclear leukocytes (PMNs) in the bone marrow, we showed that IL-6 treatment mobilizes PMNs from the marginated pool into the circulating pool at 2-6 h with a decrease in L-selectin expression on PMNs and also accelerates the release of PMNs from the postmitotic pool in the bone marrow at 12-24 h. We have concluded that IL-6 causes a biphasic neutrophilia wherein the first peak results from the mobilization of PMNs into the circulating pool from the marginated pool and the second peak results from an accelerated bone marrow release of PMNs.

Blood neutrophil functions and cytokine release in severe alcoholic hepatitis: effect of corticosteroids

Background/Aims: Several observations point to an important role of interactions between polymorphonuclear neutrophils and cytokines in severe alcoholic hepatitis. The polymorphonuclear neutrophil activation status and the local and systemic pro- and anti-inflammatory cytokine responses were quantified. The effect of corticosteroids, widely used in this setting, was evaluated using these parameters. Methods: We studied blood polymorphonuclear neutrophil functions in terms of L-selectin and β2-integrin expression, H 2O 2 production and IL-8 and tumor necrosis factor α synthesis capacity. We also measured IL-8, tumor necrosis factor α and IL-10 plasma and liver tissue levels. Fifteen patients with alcoholic hepatitis were compared to 15 patients with alcoholic cirrhosis without alcoholic hepatitis, and to 10 healthy volunteers. The impact of a 28-day course of corticosteroids on blood neutrophils activation status and cytokine levels was evaluated in patients with alcoholic hepatitis. Results: Blood polymorphonuclear neutrophils were activated, as shown by increased H 2O 2 production (48±6 vs 29±6 MFI in healthy controls), and decreased L-selectin expression (300±61 vs 449±59 in healthy controls). Upon stimulation, polymorphonuclear neutrophils synthesized large amounts of IL-8 (21.7±9.2 ng/ml vs 8.8±10 ng/ml in healthy controls) and tumor necrosis factor α (524±132 pg/ml vs 79±144 pg/ml in healthy controls). Tumor necrosis factor α and IL-8 plasma and tissue levels were markedly increased as IL-10 was barely detectable in alcoholic hepatitis patients, compared to cirrhotic patients and healthy controls. During steroid therapy, plasma levels of the pro-inflammatory cytokine IL-8 fell as early as day 14, while levels of the anti-inflammatory cytokine IL-10 increased on day 21. Finally, polymorphonuclear neutrophil functions returned to normal after treatment. Conclusion: Severe alcoholic hepatitis appears to be associated with polymorphonuclear neutrophil activation and an imbalance between pro- and anti-inflammatory cytokines; during steroid therapy a normalization of these parameters was observed.

Decreased expression of L-selectin (CD62L) on lymphocytes in enzootic bovine leukaemia.

Expression of L-selectin was determined by single- and two-colour immunofluorescence on granulocytes, peripheral blood mononuclear cells (PBMC) and blasts of bovine origin by means of a monoclonal antibody IVA94 which recognizes bovine L-selectin (CD62L). Cells were separated from peripheral blood of healthy cattle and colleagues infected with bovine leukaemia virus (BLV). BLV-infected animals comprised lymphocytotic and non-lymphocytotic cows. L-selectin was expressed on 90-98% of granulocytes in all tested animals. The percentage of PBMC expressing L-selectin was lower in cattle with persistent lymphocytosis than in non-lymphocytotic or BLV-free cattle, and inversely correlated with lymphocyte counts. The ratio of B lymphocytes stained for L-selectin was significantly decreased from 60.2 +/- 1.9% in BLV-free cattle to 43.8 +/- 3.6 and 22.5 +/- 5.7% in non-lymphocytotic and lymphocytotic cattle, respectively. B-lymphocytes stained for L-selectin exhibited about 50% reduction in L-selectin expression in BLV-infected cattle compared with BLV-free cattle, as judged by the mean fluorescence intensity (MFI). The percentage of L-selectin-positive PBMC not bearing surface immunoglobulin M (predominantly T lymphocytes) was comparable in BLV-free and BLV-infected cattle. However, L-selectin expression on T lymphocytes was reduced (about 50%) in BLV-infected cattle, as judged by the MFI. We suppose that BLV infection results in a decreased L-selectin expression on lymphocytes, and accordingly, it may contribute to deregulation of the host immune system.

Neutrophil activation in sickle cell disease.

Vascular occlusion is the main cause of the morbidity and mortality observed in patients with sickle cell disease (SCD). Increasing evidence indicates that (activated) neutrophils could play an important role in the initiation and propagation of vaso-occlusive processes in SCD. In this study, the activation state of neutrophils in sickle cell patients was analyzed by determining the level of expression of neutrophil antigens such as CD62L, CD11b, CD66b, CD63, and Fcgamma receptors. We also analyzed plasma levels of lactoferrin, elastase, soluble (s)CD16 (sFcgammaRIII), and serum levels of soluble (s)CD62L (sL-selectin) as neutrophil activation markers in these patients. Significant differences were observed in the activation state of neutrophils in non-symptomatic sickle cell patients compared to healthy HbAA controls as exemplified by significant decrease in L-selectin expression, enhanced expression of CD64, and increased levels of soluble markers like sL-selectin, elastase, and sCD16. During vaso-occlusive crisis the differences were even more pronounced. These results show neutrophils to be activated in sickle cell patients, suggesting a role of importance in the pathophysiology of sickle cell disease.

Increase in granulocyte-macrophage-colony-stimulating factor secretion and the respiratory burst with decreased L-selectin expression in hyper-IgE syndrome patients.

The hyper-IgE syndrome is a primary immunodeficiency characterized by severe recurrent abscesses, pneumonia with pneumatocele formation, and elevated serum IgE. Eosinophilia, neutrophil chemotactic defects, and marked tissue damage are frequently present in this syndrome. To study whether functional changes in cytokines, adhesion molecules, and neutrophils might help explain these clinical observations. The following functions were analyzed in patients with the hyper-IgE syndrome and in controls: (1) production of granulocyte-macrophage-colony-stimulating factor by peripheral blood mononuclear cells by ELISA; (2) respiratory burst and reactive oxygen intermediates production by peripheral neutrophils using the luminol-enhanced chemiluminescense technique; and (3) expression of L-selectin on granulocytes and lymphocytes by flow cytometry. Patients with hyper-IgE syndrome had significantly increased production of granulocyte-macrophage-colony-stimulating factor by resting or stimulated mononuclear cells, increased generation of reactive oxygen intermediates by neutrophils treated with opsonized zymosan, and reduced L-selectin expression on quiescent and activated granulocytes and lymphocytes. Our results suggest that an important feature of the hyper-IgE syndrome is the increased production of granulocyte-macrophage-colony-stimulating factor, which may explain the reduced L-selectin expression, decreased chemotaxis, and increased oxygen radical production and tissue damage in this disease.

The Effect of Glucocorticoids on the Expression of L-Selectin on Polymorphonuclear Leukocyte

When active bone marrow release is induced by inflammatory stimuli, it is associated with an increase in L-selectin expression on circulating polymorphonuclear leukocyte (PMN). This contrasts sharply with glucocorticoid-induced granulocytosis that is associated with decreased L-selectin expression on PMN. The present study was designed to determine if the reduced L-selectin expression observed after glucocorticoid treatment is the result of suppression of L-selectin synthesis in the bone marrow. New Zealand white rabbits treated with dexamethasone (2.0 mg/kg, a single dose intravenously) were shown to have decreased L-selectin expression on circulating PMN 12 to 24 hours after treatment (P &lt; .01) with a return to baseline levels by 48 hours. When dexamethasone was administered 48 hours after the bone marrow PMN were pulse labeled with the thymidine analogue, 5′-bromo-2′-deoxyuridine (BrdU), L-selectin expression on BrdU-labeled PMN released from the bone marrow was decreased (P&lt; .01). Dexamethasone decreased L-selectin expression on segmented PMN in the bone marrow (P &lt; .05) but not on PMN already in the circulation. We conclude that glucocorticoids decrease L-selectin expression on circulating PMN by downregulating L-selectin expression in the maturation pool of bone marrow and speculate that this is an important glucocorticoid effect that influences the recruitment of PMN into inflammatory sites.

The Effect of Glucocorticoids on the Expression of L-Selectin on Polymorphonuclear Leukocyte

AbstractWhen active bone marrow release is induced by inflammatory stimuli, it is associated with an increase in L-selectin expression on circulating polymorphonuclear leukocyte (PMN). This contrasts sharply with glucocorticoid-induced granulocytosis that is associated with decreased L-selectin expression on PMN. The present study was designed to determine if the reduced L-selectin expression observed after glucocorticoid treatment is the result of suppression of L-selectin synthesis in the bone marrow. New Zealand white rabbits treated with dexamethasone (2.0 mg/kg, a single dose intravenously) were shown to have decreased L-selectin expression on circulating PMN 12 to 24 hours after treatment (P < .01) with a return to baseline levels by 48 hours. When dexamethasone was administered 48 hours after the bone marrow PMN were pulse labeled with the thymidine analogue, 5′-bromo-2′-deoxyuridine (BrdU), L-selectin expression on BrdU-labeled PMN released from the bone marrow was decreased (P< .01). Dexamethasone decreased L-selectin expression on segmented PMN in the bone marrow (P < .05) but not on PMN already in the circulation. We conclude that glucocorticoids decrease L-selectin expression on circulating PMN by downregulating L-selectin expression in the maturation pool of bone marrow and speculate that this is an important glucocorticoid effect that influences the recruitment of PMN into inflammatory sites.