The bone marrow is the primary site for neutrophil production and release into the circulation. Because the CXC chemokine receptor-4/stromal derived factor-1 (CXCR4/ SDF-1) axis plays a central role in the interactions of hematopoietic stem cells, lymphocytes, and developing neutrophils in the marrow, we investigated whether reciprocal CXCR4-dependent mechanisms might be involved in neutrophil release and subsequent return to the marrow following circulation. Neutralizing antibody to CXCR4 reduced marrow retention of infused neutrophils (45.7% ؎ 0.5% to 6.9% ؎ 0.5%) and was found to mobilize neutrophils from marrow (34.4% ؎ 4.4%). Neutrophil CXCR4 expression and SDF-1-induced calcium flux decreased with maturation and activation of the cells, corresponding to the decreased marrow homing associated with these characteristics in vivo. Infusion of the inflammatory mediator and CXCR2 ligand KC led to mobilization of neutrophils from marrow by itself and was augmented 3-fold by low doses of CXCR4-blocking antibody that otherwise had no mobilizing effect. Examination of KC and SDF-1 calcium signaling demonstrated that the effect of KC may, in part, be due to heterologous desensitization to SDF-1. These results suggest that the CXCR4/SDF-1 axis is critical in circulating neutrophil homeostasis and that it may participate in the rapid release of neutrophils from the marrow during inflammation through a novel interaction with inflammatory CXC chemokines. (Blood. 2004;104: 565-571)

The severe and protracted inflammation that characterizes acute lung injury (ALI) is driven by the ongoing recruitment of neutrophils to the lung. Although much of the cytokine signaling responsible for the initial phase of ALI has been elaborated, relatively little is known about the mechanisms governing the recruitment of neutrophils from the bone marrow to the lung in the later period of this disease. Given its previously described chemoattractant effects on marrow neutrophils, we investigated whether stromal-derived factor-1 (SDF-1) (CXCL12) might participate in this later phase of recruitment. Using immunohistochemistry to examine both banked human lung specimens from patients with ALI and lungs from mice with LPS-induced pneumonitis, we found that pulmonary SDF-1 expression increases during ALI. We further determined that both lung SDF-1 protein expression and mRNA expression rise in a delayed but sustained pattern in this mouse model and that the major source of the increase in expression appears to be the lung epithelium. Lastly, we found that expression of the SDF-1 receptor CXCR4 rises in a similar temporal pattern on neutrophils in both the blood and airspace of LPS-injured mice and that Ab-mediated SDF-1 blockade significantly attenuates late but not early pulmonary neutrophilia in this model. These results implicate SDF-1 in neutrophil recruitment to the lung in the later period of acute lung injury and suggest a novel role for this cytokine in coordinating the transition from the inflammatory response to the initiation of tissue repair.

Although obesity is implicated in numerous health complications leading to increased mortality, the relationship between obesity and outcomes for critically ill patients appears paradoxical. Recent studies have reported better outcomes and lower levels of inflammatory cytokines in obese patients with acute lung injury (ALI)/acute respiratory distress syndrome, suggesting that obesity may ameliorate the effects of this disease. We investigated the effects of obesity in leptin-resistant db/db obese and diet-induced obese mice using an inhaled LPS model of ALI. Obesity-associated effects on neutrophil chemoattractant response were examined in bone marrow neutrophils using chemotaxis and adoptive transfer; neutrophil surface levels of chemokine receptor CXCR2 were determined by flow cytometry. Airspace neutrophilia, capillary leak, and plasma IL-6 were all decreased in obese relative to lean mice in established lung injury (24 h). No difference in airspace inflammatory cytokine levels was found between obese and lean mice in both obesity models during the early phase of neutrophil recruitment (2-6 h), but early airspace neutrophilia was reduced in db/db obese mice. Neutrophils from uninjured obese mice demonstrated diminished chemotaxis to the chemokine keratinocyte cytokine compared with lean control mice, and adoptive transfer of obese mouse neutrophils into injured lean mice revealed a defect in airspace migration of these cells. Possibly contributing to this defect, neutrophil CXCR2 expression was significantly lower in obese db/db mice, and a similar but nonsignificant decrease was seen in diet-induced obese mice. ALI is attenuated in obese mice, and this blunted response is in part attributable to an obesity-associated abnormal neutrophil chemoattractant response.Keywords: adult respiratory distress syndrome; chemotaxis; cytokines; innate immunity Acute lung injury (ALI) and the acute respiratory distress syndrome (ARDS) are characterized by persistent, uncontrolled pulmonary inflammation that occurs in response to a wide range of insults, including pneumonia, sepsis, and trauma (1, 2). Alveolar recruitment of neutrophils is thought to be a central factor in the onset and progression of this syndrome (3, 4), and increases in airspace neutrophilia and plasma neutrophilic cytokine levels, including TNF-a, IL-1b, IL-6, and IL-8, are associated with increased morbidity and mortality from this disease (4-6). It is increasingly recognized that ALI pathogenesis and outcome are strongly influenced by host factors, including genetic polymorphisms and comorbid conditions (1, 2). Preliminary clinical evidence suggests that obesity may have an ameliorative effect on ALI outcome (7). Although ambiguity exists in smaller studies (8, 9), recent large cohort studies from our group and others, as well as several metaanalyses, have shown a reduction in mortality with rising body mass index in ALI and critical illness in general (7,(10)(11)(12)(13)(14)(15)(16). Such an association, though tentative, is surprising because obesity...

Neutrophil retention in and release from the bone marrow is a critical process that remains incompletely understood. Previous work has implicated the CXCR4/stromal derived factor-1 (SDF-1) chemokine axis in the marrow retention of neutrophils, yet the adhesion pathways responsible for this retention are unknown. Because α4β1 integrin (VLA-4) and its ligand VCAM-1 play a central role in the interactions of hematopoietic stem cells, lymphocytes, and developing neutrophils in the marrow, we investigated whether this integrin might be involved in marrow neutrophil retention and release. In this study, we show that VLA-4 is expressed on murine marrow neutrophils and decreases with maturation, whereas blockade of this integrin leads to the release of marrow neutrophils. Marrow neutrophils adhere via VLA-4 to VCAM-1, which is expressed on marrow endothelium and stroma, and inhibition of VCAM-1 causes release of marrow neutrophils. Furthermore, SDF-1 (CXCL12) signaling through neutrophil CXCR4 augments VLA-4 adhesion to VCAM-1 in vitro, an effect that is blocked by preincubation with pertussis toxin. In vivo blockade of both CXCR4 and α4 causes synergistic release of marrow neutrophils, showing that cross-talk between CXCR4 and VLA-4 modulates marrow retention of these cells. Taken together, these results indicate that the VLA-4/VCAM adhesion pathway is critical in the retention and maturation-controlled release of neutrophils from the marrow, while providing an important link between the CXCR4/SDF-1 signaling axis and the adhesion events that govern this process.

Low tidal volume (Vt) ventilation is protective against ventilator-induced lung injury but can promote development of atelectasis. Periodic deep inflation (DI) can open the lung, but if delivered too frequently may cause damage via repeated overdistention. We therefore examined the effects of varying DI frequency on lung mechanics, gas exchange, and biomarkers of injury in mice. C57BL/6 males were mechanically ventilated with positive end-expiratory pressure (PEEP) of 2 cmH2O for 2 h. One high Vt group received a DI with each breath (HV). Low Vt groups received 2 DIs after each hour of ventilation (LV) or 2 DIs every minute (LVDI). Control groups included a nonventilated surgical sham and a group receiving high Vt with zero PEEP (HVZP). Respiratory impedance was measured every 4 min, from which tissue elastance (H) and damping (G) were derived. G and H rose progressively during LV and HVZP, but returned to baseline after hourly DI during LV. During LVDI and HV, G and H remained low and gas exchange was superior to that of LV. Bronchoalveolar lavage fluid protein was elevated in HV and HVZP but was not different between LV and LVDI. Lung tissue IL-6 and IL-1beta levels were elevated in HVZP and lower in LVDI compared with LV. We conclude that frequent DI can safely improve gas exchange and lung mechanics and may confer protection from biotrauma. Differences between LVDI and HV suggest that an optimal frequency range of DI exists, within which the benefits of maintaining an open lung outweigh injury incurred from overdistention.