Neutrophil DREAM promotes neutrophil recruitment in vascular inflammation

The interaction between neutrophils and endothelial cells is critical for the pathogenesis of vascular inflammation. However, the regulation of neutrophil adhesive function remains not fully understood. Intravital microscopy demonstrates that neutrophil DREAM promotes neutrophil recruitment to sites of inflammation induced by TNF-α but not MIP-2 or fMLP. We observe that neutrophil DREAM represses expression of A20, a negative regulator of NF-κB activity, and enhances expression of pro-inflammatory molecules and phosphorylation of IκB kinase (IKK) after TNF-α stimulation. Studies using genetic and pharmacologic approaches reveal that DREAM deficiency and IKKβ inhibition significantly diminish the ligand-binding activity of β2 integrins in TNF-α–stimulated neutrophils or neutrophil-like HL-60 cells. Neutrophil DREAM promotes degranulation through IKKβ-mediated SNAP-23 phosphorylation. Using sickle cell disease mice lacking DREAM, we show that hematopoietic DREAM promotes vaso-occlusive events in microvessels following TNF-α challenge. Our study provides evidence that targeting DREAM might be a novel therapeutic strategy to reduce excessive neutrophil recruitment in inflammatory diseases.     

Junctional adhesion molecule-C regulates vascular endothelial permeability by modulating VE-cadherin-mediated cell-cell contacts

We recently reported that junctional adhesion molecule (JAM)-C plays a role in leukocyte transendothelial migration. Here, the role of JAM-C in vascular permeability was investigated in vitro and in vivo. As opposed to macrovascular endothelial cells that constitutively expressed JAM-C in cell-cell contacts, in quiescent microvascular endothelial cells, JAM-C localized mainly intracellularly, and was recruited to junctions upon short-term stimulation with vascular endothelial growth factor (VEGF) or histamine. Strikingly, disruption of JAM-C function decreased basal permeability and prevented the VEGF- and histamine-induced increases in human dermal microvascular endothelial cell permeability in vitro and skin permeability in mice. Permeability increases are essential in angiogenesis, and JAM-C blockade reduced hyperpermeability and neovascularization in hypoxia-induced retinal angiogenesis in mice. The underlying mechanisms of the JAM-C-mediated increase in endothelial permeability were studied. JAM-C was essential for the regulation of endothelial actomyosin, as revealed by decreased F-actin, reduced myosin light chain phosphorylation, and actin stress fiber formation due to JAM-C knockdown. Moreover, the loss of JAM-C expression resulted in stabilization of VE-cadherin-mediated interendothelial adhesion in a manner dependent on the small GTPase Rap1. Together, through modulation of endothelial contractility and VE-cadherin-mediated adhesion, JAM-C helps to regulate vascular permeability and pathologic angiogenesis.     

Ligand bridging mediates integrin alpha IIb beta 3 (platelet GPIIB-IIIA) dependent homotypic and heterotypic cell-cell interactions.

The aggregation of cells bearing recombinant integrin alpha IIb beta 3 (platelet GPIIb-IIIa) has been analyzed by two-color flow cytometry. As in normal platelets, aggregation requires functional alpha IIb beta 3, "activation" of alpha IIb beta 3, and fibrinogen (fg) binding to alpha IIb beta 3. Cellular aggregation required that both interacting cells express functional alpha IIb beta 3, because a binding defective mutant, alpha IIb beta 3 (D119----Y), failed to support interaction with wild type alpha IIb beta 3-bearing cells. In addition, cells bearing resting alpha IIb beta 3 were incorporated into aggregates formed by cells bearing a constitutively active mutant, alpha IIb beta 3 (beta 1-2), indicating that only one of the cells in an interacting pair must be activated. Finally, heterotypic interactions occurred between cells bearing activated alpha IIb beta 3 and cells bearing alpha V beta 3, a fg-binding integrin present on endothelial and tumor cells. Thus, ligand bridging between fg-binding integrins represents a mechanism of cell-cell interaction, cells bearing resting alpha IIb beta 3 (e.g., resting platelets) may be incorporated into aggregates formed by cells bearing activated alpha IIb beta 3, and alpha IIb beta 3 mediates heterotypic interactions with cells bearing other fg receptors.     

Corticotropin-releasing hormone regulates IL-6 expression during inflammation

Stimulation of the hypothalamic-pituitary-adrenal (HPA) axis by proinflammatory cytokines results in increased release of glucocorticoid that restrains further development of the inflammatory process. IL-6 has been suggested to stimulate the HPA axis during immune activation independent of the input of hypothalamic corticotropin-releasing hormone (CRH). We used the corticotropin-releasing hormone-deficient (Crh(-/-)) mouse to elucidate the effect of CRH deficiency on IL-6 expression and IL-6-induced HPA axis activation during turpentine-induced inflammation. We demonstrate that during inflammation CRH is required for a normal adrenocorticotropin hormone (ACTH) increase but not for adrenal corticosterone rise. The paradoxical increase of plasma IL-6 associated with CRH deficiency suggests that IL-6 release during inflammation is CRH-dependent. We also demonstrate that adrenal IL-6 expression is CRH-dependent, as its basal and inflammation-induced expression is blocked by CRH deficiency. Our findings suggest that during inflammation, IL-6 most likely compensates for the effects of CRH deficiency on food intake. Finally, we confirm that the HPA axis response is defective in Crh(-/-)/IL-6(-/-) mice. These findings, along with the regulation of IL-6 by CRH, support the importance of the interaction between the immune system and the HPA axis in the pathophysiology of inflammatory diseases.     

Neutrophil myeloperoxidase and its substrates: formation of specific markers and reactive compounds during inflammation

Myeloperoxidase is an inflammatory enzyme that generates reactive hypochlorous acid in the presence of hydrogen peroxide and chloride ion. However, this enzyme also uses bromide ion or thiocyanate as a substrate to form hypobromous or hypothiocyanous acid, respectively. These species play important roles in host defense against the invasion of microorganisms. In contrast, these enzyme products modify biomolecules in hosts during excess inflammation, indicating that the action of myeloperoxidase is both beneficial and harmful. Myeloperoxidase uses other endogenous compounds, such as serotonin, urate, and l-tyrosine, as substrates. This broad-range specificity may have some biological implications. Target molecules of this enzyme and its products vary, including low-molecular weight thiols, proteins, nucleic acids, and lipids. The modified products represent biomarkers of myeloperoxidase action. Moderate inhibition of this enzyme might be critical for the prevention/modulation of excess, uncontrolled inflammatory events. Some phytochemicals inhibit myeloperoxidase, which might explain the reductive effect caused by the intake of vegetables and fruits on cardiovascular diseases.     

Methyltransferase G9A regulates T cell differentiation during murine intestinal inflammation

Inflammatory bowel disease (IBD) pathogenesis is associated with dysregulated CD4⁺ Th cell responses, with intestinal homeostasis depending on the balance between IL-17–producing Th17 and Foxp3⁺ Tregs. Differentiation of naive T cells into Th17 and Treg subsets is associated with specific gene expression profiles; however, the contribution of epigenetic mechanisms to controlling Th17 and Treg differentiation remains unclear. Using a murine T cell transfer model of colitis, we found that T cell–intrinsic expression of the histone lysine methyltransferase G9A was required for development of pathogenic T cells and intestinal inflammation. G9A-mediated dimethylation of histone H3 lysine 9 (H3K9me2) restricted Th17 and Treg differentiation in vitro and in vivo. H3K9me2 was found at high levels in naive Th cells and was lost following Th cell activation. Loss of G9A in naive T cells was associated with increased chromatin accessibility and heightened sensitivity to TGF-β1. Pharmacological inhibition of G9A methyltransferase activity in WT T cells promoted Th17 and Treg differentiation. Our data indicate that G9A-dependent H3K9me2 is a homeostatic epigenetic checkpoint that regulates Th17 and Treg responses by limiting chromatin accessibility and TGF-β1 responsiveness, suggesting G9A as a therapeutic target for treating intestinal inflammation.     

Angiopoietin-2 causes inflammation in vivo by promoting vascular leakage

Angiopoietins (Angs) are endothelium-selective ligands that exert most of their actions through the Tie-2 receptor. It is widely accepted that Ang-1 promotes the structural integrity of blood vessels and exhibits anti-inflammatory properties. In contrast, the role of Ang-2 remains less clear because it has been shown to behave as a Tie-2 agonist or antagonist under different experimental conditions. To define the role of Ang-2 in acute inflammation, we studied the effects of recombinant Ang-2 administration in vivo. We show herein that Ang-2, but not Ang-1, induces edema formation in the mouse paw in a dose-dependent manner; the edema seems to be fast-peaking (maximum at 30 min) and resolves within 4 h. The effect of Ang-2 is blocked by the coadministration with a soluble form of the Tie-2 receptor or Ang-1. NO and prostaglandin E(2) levels in mouse paw following the injection of Ang-2 remained unaltered, suggesting that the action of Ang-2 does not involve these mediators. In addition, Ang-2 exerted a weak stimulatory effect on leukocyte migration in the mouse paw. Similarly, Ang-2 injected into the mouse air pouch produced only a modest effect on cell extravasation that peaked at 30 min. However, when cell migration was elicited using zymosan, Ang-2 significantly inhibited leukocyte migration. We conclude that Ang-2 by itself stimulates the extravasation of cell-poor fluid, but in the presence of ongoing inflammation it reduces cellular infiltration in tissues.     

Purinergic receptor P2Y1 regulates polymodal C-fiber thermal thresholds and sensory neuron phenotypic switching during peripheral inflammation

We have recently found that, following complete Freund’s adjuvant (CFA)-induced inflammation, cutaneous polymodal nociceptors (CPM) lacking the transient receptor potential vanilloid 1 (TRPV1) are sensitized to heat stimuli. In order to determine possible mechanisms playing a role in this change, we examined gene expression in the L2/L3 sensory ganglia following CFA injection into the hairy hind paw skin and found that G-protein-coupled purinoreceptor P2Y1 expression was increased. This receptor is of particular interest, as most CPMs innervating mouse hairy skin bind isolectin B4, which co-localizes with P2Y1. Additionally, our recent findings have shown that cutaneous CPMs in P2Y1−/− mice displayed significantly reduced thermal sensitivity. Together, these findings suggested a possible role for P2Y1 in inflammation-induced heat sensitization in these fibers. To test this hypothesis, we utilized our in vivo small interfering RNA technique to knock down the inflammation-induced increase in P2Y1 expression and then examined the functional effects using ex vivo recording. We found that the normal reduction of heat thresholds in CPM fibers induced by CFA was completely blocked by inhibition of P2Y1. Surprisingly, inhibition of P2Y1 during inflammation also significantly increased the number of CPM neurons expressing TRPV1 without a change in the total number of TRPV1-positive cells in the L2 and L3 dorsal root ganglia. These results show that the inflammation-induced enhanced expression of P2Y1 is required for normal heat sensitization of cutaneous CPM fibers. They also suggest that P2Y1 plays a role in the maintenance of phenotype in cutaneous afferent fibers containing TRPV1.     

NG2 expression regulates vascular morphology and function in human brain tumours

Tumour angiogenesis is a tightly regulated process involving cross-talk between tumour cells and the host tissue. The underlying mechanisms that regulate such interactions remain largely unknown. NG2 is a transmembrane proteoglycan whose presence on transformed cells has been demonstrated to increase proliferation in vitro and angiogenesis in vivo. To study the effects of NG2 during tumour growth and progression, we engineered an NG2 positive human glioma cell line (U251-NG2) from parental NG2 negative cells (U251-WT) and implanted both cell types stereotactically into immunodeficient nude rat brains. The tumours were longitudinally monitored in vivo using multispectral MRI employing two differently sized contrast agents (Gd-DTPA-BMA and Gadomer) to assess vascular leakiness, vasogenic oedema, tumour volumes and necrosis. Comparisons of Gd-DTPA-BMA and Gadomer revealed differences in their spatial distribution in the U251-NG2 and U251-WT tumours. The U251-NG2 tumours exhibited a higher leakiness of the larger molecular weight Gadomer and displayed a stronger vasogenic oedema (69.9 +/- 15.2, P = 0.018, compared to the controls (10.7 +/- 7.7). Moreover, immunohistochemistry and electron microscopy revealed that the U251-NG2 tumours had a higher microvascular density (11.81 +/- 0.54; P = 0.0010) compared to controls (5.76 +/- 0.87), with vessels that displayed larger gaps between the endothelial cells. Thus, tumour cells can regulate both the function and structure of the host-derived tumour vasculature through NG2 expression, suggesting a role for NG2 in the cross-talk between tumour-host compartments.     

Endothelial C3a receptor mediates vascular inflammation and blood-brain barrier permeability during aging

Dysfunction of immune and vascular systems has been implicated in aging and Alzheimer disease; however, their interrelatedness remains poorly understood. The complement pathway is a well-established regulator of innate immunity in the brain. Here, we report robust age-dependent increases in vascular inflammation, peripheral lymphocyte infiltration, and blood-brain barrier (BBB) permeability. These phenotypes were subdued by global inactivation and by endothelial cell–specific ablation of C3ar1. Using an in vitro model of the BBB, we identified intracellular Ca²⁺ as a downstream effector of C3a/C3aR signaling and a functional mediator of vascular endothelial cadherin junction and barrier integrity. Endothelial C3ar1 inactivation also dampened microglia reactivity and improved hippocampal and cortical volumes in the aging brain, demonstrating a crosstalk between brain vasculature dysfunction and immune cell activation and neurodegeneration. Further, prominent C3aR-dependent vascular inflammation was also observed in a tau-transgenic mouse model. Our studies suggest that heightened C3a/C3aR signaling through endothelial cells promotes vascular inflammation and BBB dysfunction and contributes to overall neuroinflammation in aging and neurodegenerative disease.     

Circular RNA circEsyt2 regulates vascular smooth muscle cell remodeling via splicing regulation

Circular RNAs (circRNAs) have been recently recognized as playing a role in the pathogenesis of vascular remodeling–related diseases by modulating the functions of miRNAs. However, the interplay between circRNAs and proteins during vascular remodeling remains poorly understood. Here, we investigated a previously identified circRNA, circEsyt2, whose expression is known to be upregulated during vascular remodeling. Loss- and gain-of‑function mutation analyses in vascular smooth muscle cells (VSMCs) revealed that circEsyt2 enhanced cell proliferation and migration and inhibited apoptosis and differentiation. Furthermore, the silencing of circEsyt2 in vivo reduced neointima formation, while circEsyt2 overexpression enhanced neointimal hyperplasia in the injured carotid artery, confirming its role in vascular remodeling. Using unbiased protein–RNA screening and molecular validation, circEsyt2 was found to directly interact with polyC-binding protein 1 (PCBP1), an RNA splicing factor, and regulate PCBP1 intracellular localization. Additionally, circEsyt2 silencing substantially enhanced p53β splicing via the PCBP1–U2AF65 interaction, leading to the altered expression of p53 target genes (cyclin D1, p21, PUMA, and NOXA) and the decreased proliferation of VSMCs. Thus, we identified a potentially novel circRNA that regulated vascular remodeling, via altered RNA splicing, in atherosclerotic mouse models.     

The A2B adenosine receptor protects against inflammation and excessive vascular adhesion

Adenosine has been described as playing a role in the control of inflammation, but it has not been certain which of its receptors mediate this effect. Here, we generated an A2B adenosine receptor-knockout/reporter gene-knock-in (A2BAR-knockout/reporter gene-knock-in) mouse model and showed receptor gene expression in the vasculature and macrophages, the ablation of which causes low-grade inflammation compared with age-, sex-, and strain-matched control mice. Augmentation of proinflammatory cytokines, such as TNF-alpha, and a consequent downregulation of IkappaB-alpha are the underlying mechanisms for an observed upregulation of adhesion molecules in the vasculature of these A2BAR-null mice. Intriguingly, leukocyte adhesion to the vasculature is significantly increased in the A2BAR-knockout mice. Exposure to an endotoxin results in augmented proinflammatory cytokine levels in A2BAR-null mice compared with control mice. Bone marrow transplantations indicated that bone marrow (and to a lesser extent vascular) A2BARs regulate these processes. Hence, we identify the A2BAR as a new critical regulator of inflammation and vascular adhesion primarily via signals from hematopoietic cells to the vasculature, focusing attention on the receptor as a therapeutic target.     

Interleukin 25 regulates type 2 cytokine-dependent immunity and limits chronic inflammation in the gastrointestinal tract

The cytokine interleukin (IL) 25 has been implicated in the initiation of type 2 immunity by driving the expression of type 2 cytokines such as IL-5 and IL-13, although its role in the regulation of immunity and infection-induced inflammation is unknown. Here, we identify a dual function for IL-25: first, in promoting type 2 cytokine-dependent immunity to gastrointestinal helminth infection and, second, in limiting proinflammatory cytokine production and chronic intestinal inflammation. Treatment of genetically susceptible mice with exogenous IL-25 promoted type 2 cytokine responses and immunity to Trichuris. IL-25 was constitutively expressed by CD4+ and CD8+ T cells in the gut of mouse strains that are resistant to Trichuris, and IL-25-deficient mice on a genetically resistant background failed to develop a type 2 immune response or eradicate infection. Furthermore, chronically infected IL-25(-/-) mice developed severe infection-induced intestinal inflammation associated with heightened expression of interferon-gamma and IL-17, identifying a role for IL-25 in limiting pathologic inflammation at mucosal sites. Therefore, IL-25 is not only a critical mediator of type 2 immunity, but is also required for the regulation of inflammation in the gastrointestinal tract.     

牛磺酸铜干预感染愈合创面血管内皮生长因子A的表达*☆

背景:铜具有广谱杀菌及耐药性强的特点,而且铜作为人体必需微量元素在创面愈合过程中发挥着重要作用.作者前期的实验得到了利于生物体吸收的牛磺酸铜有机化合物.目的:检测牛磺酸铜的抗菌活性及其对感染创面愈合过程中血管内皮生长因子A表达的影响.方法:①通过MTT比色法检测牛磺酸铜对金黄色葡萄球菌的最低抑菌浓度.②72只SD大鼠随机分为牛磺酸铜组与生理盐水组两组,于每只大鼠背部制作1个直径1.5 cm的圆形金黄色葡萄球菌感染创面.牛磺酸铜组创面应用最低抑菌浓度的牛磺酸铜溶液1 mL,生理盐水组创面应用生理盐水1 mL,隔日用药1次,直到创面完全愈合.结果与结论:①经测定牛磺酸铜对金黄色葡萄球菌的最低抗菌浓度为4 g/L.②用药后第7,10天,牛磺酸铜组创面愈合率明显高于生理盐水组(P <0.05);至第21天,两组创面都完全愈合.③组织学观察显示:用药后第7天,牛磺酸铜组肉芽组织更成熟;用药后第14天,牛磺酸铜组胶原纤维排列整齐,瘢痕窄,胶原内可见再生的毛囊和皮脂腺.④免疫组织化学检测创面血管内皮生长因子A的表达变化显示:牛磺酸铜组血管内皮生长因子A在用药后第3天表达明显高于生理盐水组(P <0.01),第7,10天,牛磺酸铜组血管内皮生长因子A表达量下降,但仍高于生理盐水组(P <0.05).其余时间点表达未见明显差异(P >0.05).证实感染创面外用牛磺酸铜药液可以发挥有效的杀菌作用,提高创面愈合率,促进大鼠感染创面血管内皮生长因子A的表达,从而提高创面的愈合质量.     

IL-31-IL-31R interactions negatively regulate type 2 inflammation in the lung

Interleukin (IL) 31Ralpha (glycoprotein 130-like monocyte receptor and glycoprotein 130-like receptor) heterodimerizes with oncostatin M receptor beta to bind IL-31, a cytokine expressed preferentially by CD4(+) T helper type 2 (Th2) cells. However, the functions of IL-31-IL-31R signaling in immune regulation remain unknown. Here, we identify a novel role for IL-31R in limiting type 2 inflammation in the lung. After intravenous injection of Schistosoma mansoni eggs, IL-31Ralpha(-/-) mice developed severe pulmonary inflammation, characterized by an increase in the area of granulomatous inflammation, increased numbers of resistin-like molecule alpha(+) cells, and enhanced collagen deposition compared to WT counterparts. In vitro, macrophages generated from IL-31Ralpha(-/-) mice promoted enhanced ovalbumin-specific CD4(+) T cell proliferation and purified naive CD4(+) T cells from IL-31Ralpha(-/-) mice exhibited enhanced proliferation and expression of Th2 cytokines, identifying a T cell- and macrophage-intrinsic regulatory function for IL-31R signaling. In contrast, the generation of CD4(+) T cell-mediated Th1 responses were normal in IL-31Ralpha(-/-) mice, suggesting that the regulatory role of IL-31R signaling is limited to type 2 responses. Together, these data implicate IL-31R signaling as a novel negative regulatory pathway that specifically limits type 2 inflammation.     

Neutrophil elastase, MIP-2, and TLR-4 expression during human and experimental sepsis

Highly activated neutrophils play a critical role in mediating organ injury in sepsis by releasing neutrophil elastase (NE). Toll-like receptors (TLRs) play an important role in the host defense against invading microbes, and their signaling pathway is critical to the activation of the proinflammatory response. However, the relationship between TLR expression and the host defense mechanism during sepsis has not been fully elucidated. In this paper, we investigated the relationships among chemokine (MIP-2), TLR-4, and NE expression in human sepsis and murine peritonitis (CLP). TLR-4 expression on monocytes/macrophages was examined in patients with sepsis and in murine peritonitis and was markedly increased in both populations. LPS-induced MIP-2 production by bronchoalveolar cells and liver mononuclear cells in mice with peritonitis was also significantly increased compared with sham-operated mice. Pretreatment of the macrophage cell line, RAW 264.7 cells, with a NE inhibitor before their exposure to LPS resulted in a significant dose-dependent decrease in MIP-2 production, which was comparable to that seen following pretreatment with TLR-4 antibody. Furthermore, NE and LPS both up-regulated TLR-4 expression on human peripheral blood monocytes. Thus, chemokine-induced recruitment of neutrophils in sepsis may result in further increased chemokine production and increased expression of TLR-4. Neutrophil-derived NE may be associated with increased expression of monocyte/macrophage TLR-4, thereby serving as a positive feedback loop for the inflammatory response among the different cell populations.     

Marked increase in vascular endothelial growth factor concentrations during Escherichia coli endotoxin-induced acute inflammation in humans

BACKGROUND: Bacterial endotoxins can induce the synthesis and release of vascular endothelial growth factor (VEGF), which may alter vascular permeability and cause vascular leakage. MATERIALS AND METHODS: The effect of acute systemic inflammation on VEGF concentration was measured in healthy males after an intravenous bolus infusion of Escherichia coli endotoxin (lipopolysaccharide, LPS, 20 IU kg-1) in a double-blind, placebo-controlled parallel group study. LPS administration was followed by an infusion of lepirudin (bolus 0.1 mg kg-1, continuous infusion of 0.1 mg kg-1 h-1, n = 12) or saline (n = 12). RESULTS: Plasma VEGF increased from a mean of 15.1 pg mL-1 to 74.6 pg mL-1 5 h after LPS (P < 0.003). Body temperature, pulse rate, leukcytes, prothrombin fragment 1 + 2 (F1 + 2) and lactoferrin increased and platelets decreased after LPS (P < 0.05). The LPS-induced increase in VEGF was paralleled by the neutrophil cell degranulation marker lactoferrin but not by F1 + 2, and was not affected by lepirudin, which blunted F1 + 2 formation (P < 0.05). CONCLUSIONS: Inflammation-induced activation of leukcytes rather than platelets plays a role in the marked increase in VEGF, which cannot be abrogated by antithrombotic therapy.     

CXCR2- and E-selectin-induced neutrophil arrest during inflammation in vivo

The signaling events leading to the activation of integrins and firm arrest of rolling neutrophils in inflamed venules have yet to be elucidated. In vitro assays suggest that both E-selectin and chemokines can trigger arrest of rolling neutrophils, but E-selectin(-/-) mice have normal levels of adherent neutrophils in inflamed venules. To test whether chemokine-induced neutrophil arrest in vivo can be unmasked by blocking E-selectin, we investigated neutrophil adhesion in inflamed cremaster muscle venules in tumor necrosis factor (TNF)-alpha-treated CXCR2(-/-) or wild-type (WT) mice injected with E-selectin blocking monoclonal antibody (mAb) 9A9. To block chemokine receptor signaling, we investigated E-selectin(-/-) or WT mice treated with pertussis toxin (PTx) intravenously. Neutrophil adhesion was unchanged in CXCR2(-/-), E-selectin(-/-), PTx-treated WT, or mAb 9A9-treated WT mice. However, TNF-alpha-induced neutrophil adhesion was almost completely abrogated in E-selectin(-/-) mice treated with PTx and significantly reduced in CXCR2(-/-) mice treated with the E-selectin blocking mAb. In thioglycollate-induced peritonitis, PTx treatment blocked neutrophil recruitment into the peritoneum of E-selectin(-/-) mice, but had only a partial effect in WT animals. These data show that E-selectin- and chemokine-mediated arrest mechanisms are overlapping in this model and identify CXCR2 as an important neutrophil arrest chemokine in vivo.