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

Department of Pharmacology, Faculty of Medicine of Ribeirão Preto, University of São Paulo, Av. Bandeirantes 3900, Monte Alegre, 14049-900 Ribeirão Preto, SP, Brazil.
American Journal of Respiratory and Critical Care Medicine (Impact Factor: 11.99). 04/2007; 175(5):490-7. DOI: 10.1164/rccm.200601-103OC
Source: PubMed

ABSTRACT 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
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).
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.
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.


Available from: Fabrício Rios-Santos, Jun 16, 2015
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Sepsis-induced acute lung injury is characterized by activation and injury of pulmonary microvascular endothelial cells (PMVEC), increased neutrophil-PMVEC adhesion and migration, and trans-PMVEC high-protein edema. Inducible NO synthase (iNOS) inhibits septic murine neutrophil migration in vivo and in vitro. The effects of NO in human neutrophil-PMVEC interactions are not known. We isolated human PMVEC using magnetic bead-bound anti-PECAM antibody. Confluent PMVEC at passage 3-4 were co-cultured with human neutrophils for assessment of neutrophil-PMVEC adhesion, and trans-PMVEC neutrophil migration and Evans-Blue dye-labeled albumin leak. Two NO donors (spermine-NONOate, S-nitroso-N-acetylpenicillamine) attenuated both cytomix-enhanced neutrophil-PMVEC adhesion by 64+/-14% (p<0.01) and 32+/-3% (p<0.05), respectively, and cytomix-induced trans-PMVEC neutrophil migration by 85+/-16% (p<0.01) and 43+/-5% (p<0.01), respectively. Correspondingly, iNOS inhibition with 1400W enhanced cytomix-stimulated neutrophil migration by 52+/-3% (p<0.01), but had no effect on neutrophil-PMVEC adhesion. Conversely, a peroxynitrite donor (SIN-1) increased both neutrophil-PMVEC adhesion (38+/-2% vs. 14+/-1% control, p<0.01) and trans-PMVEC neutrophil migration; with both effects were completely inhibited by scavenging of NO, superoxide, or peroxynitrite (p<0.05 for each). Scavenging of peroxynitrite also eliminated cytomix-induced neutrophil adhesion and migration. Blocking CD18-dependent neutrophil adhesion prevented cytomix-stimulated trans-PMVEC EB-albumin leak (p<0.05), while inhibiting neutrophil migration paradoxically enhanced cytomix-stimulated EB-albumin leak (11+/-1% vs. 7+/-0.5%, p<0.01). FMLP-induced neutrophil migration had no effect on trans-PMVEC EB-albumin leak. In summary, we report differential effects, including the inhibitory action of NO and stimulatory effect of ONOO(-) on human neutrophil-PMVEC adhesion and trans-PMVEC migration under cytomix stimulation. Moreover, neutrophil-PMVEC adhesion, but not trans-PMVEC migration, contributes to human PMVEC barrier dysfunction.
    Microvascular Research 06/2008; 76(2):80-8. DOI:10.1016/j.mvr.2008.06.001 · 2.43 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Sepsis continues to have a high mortality rate worldwide. The multi-step effects of this syndrome make it difficult to develop a comprehensive understanding of its pathophysiology and to identify a direct treatment. Neutrophils play a major role in controlling infection. Interestingly, the recruitment of these cells to an infection site is markedly reduced in severe sepsis. The systemic activation of Toll-like receptors and high levels of TNF-α and nitric oxide are involved in the reduction of neutrophil recruitment due to down-regulation of CXCR2 in neutrophils. By contrast, CCR2 is expressed in neutrophils after sepsis induction and contributes to their recruitment to organs far from the infection site, which contributes to organ damage. This review provides an overview of the recent advances in the understanding of the role of neutrophils in sepsis, highlighting their potential as a therapeutic target.
    Expert Review of Clinical Immunology 05/2014; DOI:10.1586/1744666X.2014.922876 · 3.34 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Sepsis is still a leading cause of death worldwide and the mechanism of shock remains to be completely understood. Several studies have aimed to evaluate the effects of several drugs and procedures in sepsis, and the most common models of this study are to challenge mice with LPS or to simulate a polymicrobial infection using a surgical procedure. Such procedure consists in exposure of the cecum by a midline laparotomy, ligature of ileocecal junction and perforation with a needle, squeezing cecum contents to the peritoneum cavity. Beyond the variations allowed by this model, the thickness of the needle used and the number of perforations seem to be an important factor, displaying different levels of sepsis severity. In this study, we used two mice strains (C57BL/6 and BALB/c) to describe the procedures of cecal ligation and puncture (CLP), comparing the survival rates of mice subjected to three different thicknesses of perforation.