Neutrophils are involved in the development of acute pancreatitis (AP), but it is not clear how neutrophil-induced tissue damage is regulated. In addition to secreting antimicrobial compounds, activated neutrophils eliminate invading microorganisms by expelling nuclear DNA and histones to form extracellular web-like structures called neutrophil extracellular traps (NETs). However, NETs have been reported to contribute to organ dysfunction in patients with infectious diseases. We investigated whether NETs contribute to the development of AP in mice. AP was induced in C57BL/6 mice by infusion of taurocholate into the pancreatic duct or by intraperitoneal administration of L-arginine. Pancreata were collected and extracellular DNA was detected by Sytox green staining, levels of CXC chemokines, histones, and cytokines also were measured. Cell-free DNA was quantified in plasma samples. Signal transducer and activator of transcription 3 phosphorylation and trypsin activation were analyzed in isolated acinar cells. NETs were depleted by administration of DNase I to mice. Plasma was obtained from healthy individuals (controls) and patients with severe AP. Infusion of taurocholate induced formation of NETs in pancreatic tissues of mice and increased levels of cell-free DNA in plasma. Neutrophil depletion prevented taurocholate-induced deposition of NETs in the pancreas. Administration of DNase I to mice reduced neutrophil infiltration and tissue damage in the inflamed pancreas and lung, and decreased levels of blood amylase, macrophage inflammatory protein-2, interleukin 6, and high-mobility groups protein 1. In mice given taurocholate, DNase I administration also reduced expression of integrin α M (macrophage-1 antigen) on circulating neutrophils. Similar results occurred in mice with L-arginine–induced AP. Addition of NETs and histones to acinar cells induced formation of trypsin and activation of signal transducer and activator of transcription 3; these processes were blocked by polysialic acid. Patients with severe AP had increased plasma levels of NET components compared with controls. NETs form in the pancreata of mice during the development of AP, and NET levels are increased in plasma from patients with AP, compared with controls. NETs regulate organ inflammation and injury in mice with AP, and might be targeted to reduce pancreatic tissue damage and inflammation in patients.

Blood vascular endothelial cells (BECs) control the immune response by regulating blood flow and immune cell recruitment in lymphoid tissues. However, the diversity of BEC and their origins during immune angiogenesis remain unclear. Here we profile transcriptomes of BEC from peripheral lymph nodes and map phenotypes to the vasculature. We identify multiple subsets, including a medullary venous population whose gene signature predicts a selective role in myeloid cell (vs lymphocyte) recruitment to the medulla, confirmed by videomicroscopy. We define five capillary subsets, including a capillary resident precursor (CRP) that displays stem cell and migratory gene signatures, and contributes to homeostatic BEC turnover and to neogenesis of high endothelium after immunization. Cell alignments show retention of developmental programs along trajectories from CRP to mature venous and arterial populations. Our single cell atlas provides a molecular roadmap of the lymph node blood vasculature and defines subset specialization for leukocyte recruitment and vascular homeostasis.

Recent findings indicate that platelets not only regulate thrombosis and hemostasis but may also be involved in proinflammatory activities. Herein, we hypothesized that platelets may play a role in sepsis by activating and priming circulating neutrophils for subsequent recruitment into the lung. Prospective experimental study. University Hospital Research Unit. Male C57BL/6 mice. Lung edema, bronchoalveolar infiltration of neutrophils, levels of myeloperoxidase, expression and function of membrane-activated complex-1 (Mac-1) on neutrophils and the CXC chemokines, macrophage inflammatory protein-2, and cytokine-induced neutrophil chemoattractant were determined after cecal ligation and puncture (CLP). Mice received a platelet-depleting antibody as well as antibodies directed against P-selectin glycoprotein-ligand-1 and Mac-1 before CLP induction. CLP caused significant pulmonary damage characterized by neutrophil infiltration, increased levels of CXC chemokines, and edema formation in the lung. Furthermore, CLP up-regulated Mac-1 expression on neutrophils and increased the number of neutrophils binding platelets in the circulation. Interestingly, depletion of platelets reduced CLP-induced edema and neutrophil recruitment in the bronchoalveolar space by >60%. Furthermore, depletion of platelets reduced Mac-1 expression on neutrophils. On the other hand, inhibition of P-selectin glycoprotein-ligand-1 abolished CLP-induced neutrophil–platelet aggregation but had no effect on neutrophil expression of Mac-1. These data demonstrate that platelets play a key role in regulating infiltration of neutrophils and edema formation in the lung via upregulation of Mac-1 in abdominal sepsis.

To define the role of CD40L in abdominal sepsis. Platelets promote sepsis-induced pulmonary recruitment of neutrophils. However, the identity of the platelet-derived molecule regulating neutrophil infiltration is not known. The hypothesis of the present study was that platelet-derived CD40L might be responsible for platelet-mediated activation and accumulation of neutrophils in sepsis. Wild-type C57BL/6 mice and CD40L gene-deficient mice were exposed to cecal ligation and puncture (CLP). Lung edema, bronchoalveolar neutrophils, CD40L and macrophage inflammatory protein-2 (MIP-2) plasma levels, myeloperoxidase activity and Mac-1 expression were determined up to 24 hours after CLP induction. For platelet depletion was an anti-GP1bα antibody administered before CLP. Plasma levels of soluble CD40L increased and surface expression of CD40L on platelets decreased in CLP mice. Platelet depletion reduced CLP-induced CD40L levels by 90%. CLP-provoked Mac-1 expression on neutrophils was abolished in CD40L-deficient mice. Interestingly, CLP-induced edema and myeloperoxidase activity in the lung as well as neutrophil infiltration in the broncoalveolar space were markedly reduced in mice lacking CD40L. In vitro experiments showed that CD40L was not capable of directly increasing Mac-1 levels on neutrophils. Instead, CLP-induced plasma levels of MIP-2 were significantly reduced in CD40L-deficient mice and inhibition of the MIP-2 receptor (CXCR2) decreased Mac-1 expression on neutrophils in septic animals. CD40L derived from platelets is a potent activator of neutrophils and mediates sepsis-induced neutrophil recruitment and lung edema. The neutrophil activating mechanism of CD40L is indirect and mediated via MIP-2 formation and CXCR2 signaling. Targeting CD40L may be an effective approach to limit pulmonary damage in abdominal sepsis.

Excessive neutrophil activation is a major component in septic lung injury. Neutrophil-derived DNA may form extracellular traps in response to bacterial invasions. The aim of the present study was to investigate the potential role of neutrophil extracellular traps (NETs) in septic lung injury. Male C57BL/6 mice were treated with recombinant human (rh)DNAse (5 mg/kg) after cecal ligation and puncture (CLP). Extracellular DNA was stained by Sytox green, and NET formation was quantified by confocal microscopy and cell-free DNA in plasma, peritoneal cavity, and lung. Blood, peritoneal fluid, and lung tissue were harvested for analysis of neutrophil infiltration, NET levels, tissue injury, as well as CXC chemokine and cytokine formation. We observed that CLP caused increased formation of NETs in plasma, peritoneal cavity, and lung. Administration of rhDNAse not only eliminated NET formation in plasma, peritoneal cavity, and bronchoalveolar space but also reduced lung edema and tissue damage 24 h after CLP induction. Moreover, treatment with rhDNAse decreased CLP-induced formation of CXC chemokines, IL-6, and high-mobility group box 1 (HMGB1) in plasma, as well as CXC chemokines and IL-6 in the lung. In vitro, we found that neutrophil-derived NETs had the capacity to stimulate secretion of CXCL2, TNF-α, and HMGB1 from alveolar macrophages. Taken together, our findings show that NETs regulate pulmonary infiltration of neutrophils and tissue injury via formation of proinflammatory compounds in abdominal sepsis. Thus we conclude that NETs exert a proinflammatory role in septic lung injury.

Neutrophils form sticky web-like structures known as neutrophil extracellular traps (NETs) as part of innate immune response. NETs are decondensed extracellular chromatin filaments comprising nuclear and cytoplasmic proteins. NETs have been implicated in many gastrointestinal diseases including colorectal cancer (CRC). However, the regulatory mechanisms of NET formation and potential pharmacological inhibitors in the context of CRC have not been thoroughly discussed. In this review, we intend to highlight roles of NETs in CRC progression and metastasis as well as the potential of targeting NETs during colon cancer therapy.