Diminished Soluble and Total Cellular L-Selectin in Cord Blood Is Associated with Its Impaired Shedding from Activated Neutrophils

Koenig, Joyce M.; Simon, J.; Anderson, Donald C.; Smith, E. O’Brian; Smith, C. W.

doi:10.1203/00006450-199604000-00009

Cited by 45 publications

(33 citation statements)

References 42 publications

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“…However, chemotactic stimulation with fMLP increased surface expression of CD11b to a greater degree in adult PMN than in cord blood PMN (Fig. 3), results that are similar to those we and others have observed (19,26,27).…”

Section: Resultssupporting

confidence: 91%

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Neonatal Neutrophils with Prolonged Survival Exhibit Enhanced Inflammatory and Cytotoxic Responsiveness

et al. 2005

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Apoptosis is critical to the resolution of inflammation, as it promotes the removal of neutrophils (PMN) by the reticuloendothelial system. In contrast, PMN persistence characterizes the early stages of chronic inflammation. Adult PMN with delayed senescence retain some functionality, although this has not been described for neonatal PMN. We hypothesized that neonatal PMN with prolonged survival retain cytotoxic and inflammatory function. To test one aspect of inflammatory function, we determined surface CD11b expression on 0-h and 24-h PMN after chemotactic formyl-methionine-leucine-phenylalanine (fMLP) stimulation. Although fMLP induced a greater percentage upregulation of CD11b on 0-h adult PMN, this was similar between nonapoptotic cord blood and adult PMN at 24 h. Furthermore, percentage up-regulation of CD11b was more robust for 24-h than for 0-h cord blood PMN. In contrast, there was no difference in responsiveness between 0-h and 24-h adult PMN. In studies of cytotoxic potential, we determined the expression of reactive oxygen intermediates (ROI) in phorbol 12-myristate 13-acetatestimulated cord blood and adult PMN at 0 h and in 24-h nonapoptotic PMN, using the dihydrorhodamine 123 assay.Stimulated cord blood PMN generated more ROI than did adult PMN at both 0 h and 24 h; in addition, ROI levels in 24-h cord blood PMN were similar to those of 0-h adult PMN. We conclude that PMN with prolonged survival retain specific cytotoxic and inflammatory functions, and these are enhanced in cord blood PMN. We speculate that neonatal PMN with prolonged survival have the functional capacity to contribute to the pathogenesis of inflammatory disorders. Abbreviations CLD, chronic lung disease DHR, dihydrorhodamine 123 fMLP, formyl-methionine-leucine-phenylalanine PE, phycoerythrin PMA, phorbol 12-myristate 13-acetate PMN, neutrophil(s) ROI, reactive oxygen intermediates 7-AAD, 7-aminoactinomycin PMN serve as the primary line of host defense during the early stages of sepsis and inflammation (1). After activation by systemic or local factors, primed endothelial cells interact with PMN through adhesion molecules that include selectins and integrins (2). These adhesive interactions initiate a process that allows the extravasation of PMN into inflamed tissue with the goal of neutralizing the inciting stimulus. In the case of inflammation mediated by bacterial or fungal microorganisms, phagocytosis and subsequent killing of these invaders occurs through mechanisms that partly involve ROI and proteases (3,4).The concept that apoptosis of inflammatory PMN and their timely removal by resident macrophages are critical to the resolution of inflammation has been well established (5,6). During the resolution phase, PMN typically undergo the phenotypic changes associated with apoptosis, which promotes their removal by resident macrophages (7). PMN can also undergo necrosis and lysis, a process that induces cytotoxicity in surrounding tissues (8), although apoptosis is the more common and physiologically preferable route. All cells a...

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Section: Resultssupporting

confidence: 91%

“…Dextran-sedimented leukocytes were subjected to density centrifugation, and the resultant was subjected to hypotonic lysis of contaminating erythrocytes, as previously described (19). Isolated PMN (10 6 cells/mL) suspended in RPMI 1640/2% FCS (Mediatech Inc., Herndon, VA) were cultured in polypropylene tubes at 37°C, 5% CO 2 , for up to 24 h.…”

Section: Methodsmentioning

confidence: 99%

Neonatal Neutrophils with Prolonged Survival Exhibit Enhanced Inflammatory and Cytotoxic Responsiveness

et al. 2005

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“…These factors make in vitro experiments with cord blood less accurate in pathophysiologic studies of MAS compared with adult blood experiments, which can be performed under strictly controlled conditions. It was shown previously that cord-blood neutrophils have lower CD62L expression compared with adult blood (48,49). Our data are in accordance with these observations.…”

Section: Discussionsupporting

confidence: 94%

Complement C5a Is a Key Mediator of Meconium-Induced Neutrophil Activation

Castellheim

Pharo

Fung³

et al. 2005

Pediatr Res

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Meconium aspiration syndrome is a serious condition of the newborn characterized by pulmonary inflammation with substantial neutrophil infiltration. We recently showed that meconium is a potent activator of complement. The aim of the present study was to investigate a possible role for complement in meconiuminduced neutrophil activation. Meconium was incubated in human whole blood anticoagulated with lepirudin, a specific thrombin inhibitor that does not affect complement activation. Complement activation was detected by measuring the terminal complement complex. Neutrophil oxidative burst and changes in CD11b and L-selectin expression were measured by flow cytometry. Complement was inhibited using the MAb 166-32 and 137-26, which block factor D and neutralize C5a, respectively. Meconium markedly activated the neutrophils, as revealed by up-regulation of CD11b, accentuation of L-selectin shedding, and induction of oxidative burst. Complement inhibition using the anti-factor D antibody completely (95-100%) blocked meconium-induced changes in CD11b and L-selectin expression, whereas oxidative burst was reduced by 60 -70%. The anti-C5a antibody inhibited the neutrophil activation to the same extent as anti-factor D. The data suggest that complement activation is largely responsible for the neutrophil inflammatory responses induced by meconium in vitro and that C5a is a key mediator of this response. Abbreviations ARDS, acute respiratory distress syndrome AU, arbitrary units DHR, dihydrorhodamine LPS, lipopolysaccharide MAS, meconium aspiration syndrome MFI, median fluorescence intensity PE, phycoerythrin PMA, phorbol-12-␤-myristat-13-␣-acetate TCC, terminal SC5b-9 complement complex Meconium aspiration syndrome (MAS) may cause a profound lung inflammation similar to acute respiratory distress syndrome (ARDS). A substantial increase in neutrophils in the airways; increased levels of inflammatory cytokines, eicosanoids, superoxide anions, and endothelin-1; expression of inducible nitric oxide synthase and cyclooxygenase-2; and activation of nuclear factor-B have been described in MAS (1-8). A profound inflammatory response and the release of leukocyte cytotoxic products have an important role in initiation and propagation of lung injury in ARDS, in which neutrophils and their cytotoxic products play a crucial role (9).The complement system is a key mediator of inflammation. Complement-mediated activation of neutrophils and monocytes leads to cytokine production; release of lipid mediators and histamine; oxidative burst; and altered expression of adhesion molecules, including increased expression of CD11b and shedding, revealed as decreased expression of CD62L. We showed recently that meconium is a potent activator of the alternative complement pathway in vitro (10). To study the interaction between all participating inflammatory systems, we used an in vitro human whole-blood model that is based on the use of lepirudin, a thrombin-specific anticoagulant that does not interfere with complement activation (11). We hypot...

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“…Koenig et al (21) have reported that lysates of PMN from term neonatal cord blood have lower total L-selectin than adults, corroborating a cellular deficiency in the neonate. However, our findings of a similar loss of L-selectin compared with baseline between neonatal and adult neutrophils after FMLP stimulation differ from most previous studies, which have reported less shedding of L-selectin from neonates (4,5,21). We used high doses of FMLP, which may have masked differential losses at lower FMLP concentration.…”

Section: Discussioncontrasting

confidence: 86%

Comparison of L-Selectin and CD11b on Neutrophils of Adults and Neonates during the First Month of Life

KIM

2003

Pediatric Research

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The newborn infant is particularly susceptible to infection in the first weeks of life and this may be, in part, related to functional impairment of neonatal neutrophils in regard to adherence, chemotaxis, and migration. Differences in expression of the neutrophil adherence molecules, L-selectin and CD11b/CD18 (Mac-1), have been previously demonstrated in cord blood and in very young infants (Յ 48 h) compared with adults, but it is unknown for how long these differences persist. We measured surface expression of neutrophil L-selectin and CD11b using flow cytometry in healthy term human newborns from 24 h through 4 wk of age. We also measured levels of soluble L-selectin using an ELISA in neonates though the age of 4 wk. Compared with adults, neonates expressed significantly less L-selectin on resting neutrophils through 4 wk of age, with the lowest levels being at 24 h. Expression of L-selectin on the neutrophil after activation with N-formyl-methionyl-leucyl-phenylalanine was less in the neonate than in the adult (p Ͻ 0.05) for the first week of life. Soluble L-selectin showed a steady increase with age in the neonates. Soluble L-selectin was significantly lower in the 24-h neonate compared with the adult and was higher in the 4-wk neonate compared with the adult. CD11b expression was similar between neonates and adults on unstimulated neutrophils, but upon activation, the neonatal neutrophil demonstrated significantly lower up-regulation of CD11b on the neutrophil surface through 4 wk of age compared with adults (p Ͻ 0.05 for all ages except 1 wk). These findings that differences in expression of L-selectin and CD11b from that in adults persist throughout the neonatal period provide further evidence that these differences may play a role in the neutrophil defects observed during the neonatal period. The newborn infant is particularly susceptible to severe bacterial and fungal infections (1, 2). It is unknown for how long this susceptibility persists, but it is a clinical practice to treat these infants as immunocompromised for at least the length of the neonatal period (first 28 d of life). Neonatal neutrophils, compared with those of adults, have been found to have functional impairments in their ability to adhere to an activated endothelial surface and to migrate in response to chemotaxins (2-6). These impairments may delay or prevent neutrophils from reaching inflammatory sites, thus inhibiting the ability of the neonate to effectively fight infection.Adherence of neutrophils to the vascular endothelium is an early step in the process of neutrophil migration into inflamed tissue and is mediated by proteins expressed on the surface of the neutrophil and the endothelium. Members of two important families of adhesion molecules located on the surface of the neutrophil are L-selectin and the ␤2-integrins. L-Selectin is found on neutrophils and other leukocytes and recognizes carbohydrate ligands on the endothelial cell, including sialyl Lewis (7). L-Selectin is thought to mediate, along with other selectin prote...

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Diminished Soluble and Total Cellular L-Selectin in Cord Blood Is Associated with Its Impaired Shedding from Activated Neutrophils

Cited by 45 publications

References 42 publications

Neonatal Neutrophils with Prolonged Survival Exhibit Enhanced Inflammatory and Cytotoxic Responsiveness

Neonatal Neutrophils with Prolonged Survival Exhibit Enhanced Inflammatory and Cytotoxic Responsiveness

Complement C5a Is a Key Mediator of Meconium-Induced Neutrophil Activation

Comparison of L-Selectin and CD11b on Neutrophils of Adults and Neonates during the First Month of Life

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