The role of neutrophils in inflammation resolution.

The fundamental role played by neutrophils for an efficient, acute inflammatory response has long been appreciated, with the underlying molecular and cellular mechanisms largely elucidated over the past decades. However, more recent work suggests that the biological functions exerted by this fascinating leucocyte are somewhat more extensive than previously acknowledged. Here we discuss how extravasated neutrophils govern the initiation of the resolution phase of inflammation by enabling activation of pro-resolving circuits to ensure the safe conclusion of the inflammatory response. The neutrophil 'alarm bell' on resolution is effected through release of soluble mediators as well as apoptotic bodies and other vesicles, which, in turn, can inform and modify the microenvironment ultimately leading to termination of the inflammatory response coinciding with re-establishment of tissue homeostasis and functionality.

Acute coronary syndrome secondary to cardiac infiltration and coronary occlusion of chronic lymphocytic leukemia - A case report.

A 72-year-old male with a history of chronic lymphocytic leukemia (CLL) was admitted to hospital with a productive cough and an episode of diarrhea and vomiting. He was initially treated for pneumonitis and sepsis. On the 12th day of his admission, he reported chest pain. Changes on his electrocardiogram were suggestive of myocardial ischemia and an elevated troponin rise was detected from his blood tests. A diagnosis of acute coronary syndrome was made but due to his frailty, he was medically managed. His echocardiogram revealed an external echogenic mass which invaded the anterolateral left ventricular wall. Further imaging with cardiac magnetic resonance imaging (MRI) and computed tomography (CT) thorax demonstrated external encasement of left circumflex coronary artery with mediastinal mass, leading into downstream myocardial ischemia and subsequent necrosis. He was considered suitable for aggressive radiotherapy/chemotherapy but passed away 7 days later. This case highlights the unusual case where an acute myocardial infarction can be attributed to direct infiltration and external compression of coronary artery by mediastinal tumor and the value of multi-modality imaging (echocardiogram, CT, and MRI) in identifying the cause of myocardial ischemia in patients with CLL in the end stages of the disease. .

Giant Circumflex Artery Aneurysm With a Coronary Sinus Fistula.

Giant coronary artery aneurysms are extremely rare entities, and the authors are aware of only 13 case reports of such aneurysms of the circumflex artery. This case report is provided to add evidence of the natural history and pathophysiology and describe the successful surgical treatment of this condition.

Microvesicle Subsets in Sepsis Due to Community Acquired Pneumonia Compared to Faecal Peritonitis.

RATIONALE: Microvesicles (MV) act as a nonsoluble means of intercellular communication, with effector roles in disease pathogenesis and potentially as biomarkers. Previously, we reported that neutrophil MV expressing alpha-2-macroglobulin (A2MG) are protective in experimental sepsis and associate with survival in a small cohort of patients with sepsis due to community acquired pneumonia (CAP). OBJECTIVES: To characterize MV profiles in sepsis due to CAP or fecal peritonitis (FP) and determine their relation to outcome. To investigate the effects of novel sepsis treatments (granulocyte-macrophage colony stimulating factor (GM-CSF) and interferon-υ (IFN-γ)) on MV production and functions in vitro. METHODS: Flow cytometry analysis of MV identified the cell of origin and the proportion of A2MG expression in the plasma of patients with sepsis secondary to CAP (n = 60) or FP (n = 40) and compared with healthy volunteers (HV, n = 10). The association between MV subsets and outcome was examined. The ability of GM-CSF and IFN-γ on A2MG MV production from whole blood was examined together with the assessment of their effect on neutrophil and endothelial functions. RESULTS: Circulating cell-derived and A2MG MV were higher in CAP compared with FP and HV. A2MG MV were higher in survivors of CAP, but not in FP. GM-CSF and IFN-γ enhanced A2MG MV production, with these MV eliciting pathogen clearance in vitro. CONCLUSIONS: Plasma MV profiles vary according to the source of infection. A2MG MV are associated with survival in CAP but not FP. We propose specific MV subsets as novel biomarkers in sepsis and potential effector for some of the actions of experimental therapeutic interventions.

Exosomal cargo including microRNA regulates sensory neuron to macrophage communication after nerve trauma.

Following peripheral axon injury, dysregulation of non-coding microRNAs (miRs) occurs in dorsal root ganglia (DRG) sensory neurons. Here we show that DRG neuron cell bodies release extracellular vesicles, including exosomes containing miRs, upon activity. We demonstrate that miR-21-5p is released in the exosomal fraction of cultured DRG following capsaicin activation of TRPV1 receptors. Pure sensory neuron-derived exosomes released by capsaicin are readily phagocytosed by macrophages in which an increase in miR-21-5p expression promotes a pro-inflammatory phenotype. After nerve injury in mice, miR-21-5p is upregulated in DRG neurons and both intrathecal delivery of a miR-21-5p antagomir and conditional deletion of miR-21 in sensory neurons reduce neuropathic hypersensitivity as well as the extent of inflammatory macrophage recruitment in the DRG. We suggest that upregulation and release of miR-21 contribute to sensory neuron-macrophage communication after damage to the peripheral nerve.

Proresolving and cartilage-protective actions of resolvin D1 in inflammatory arthritis.

Rheumatoid arthritis (RA) is a debilitating disease characterized by persistent accumulation of leukocytes within the articular cavity and synovial tissue. Metabololipidomic profiling of arthritic joints from omega-3 supplemented mice identified elevated levels of specialized proresolving lipid mediators (SPM) including resolvin D1 (RvD1). Profiling of human RA synovial fluid revealed physiological levels of RvD1, which - once applied to human neutrophils - attenuated chemotaxis. These results prompted analyses of the antiarthritic properties of RvD1 in a model of murine inflammatory arthritis. The stable epimer 17R-RvD1 (100 ng/day) significantly attenuated arthritis severity, cachexia, hind-paw edema, and paw leukocyte infiltration and shortened the remission interval. Metabololipidomic profiling in arthritic joints revealed 17R-RvD1 significantly reduced PGE2 biosynthesis, while increasing levels of protective SPM. Molecular analyses indicated that 17R-RvD1 enhanced expression of genes associated with cartilage matrix synthesis, and direct intraarticular treatment induced chondroprotection. Joint protective actions of 17R-RvD1 were abolished in RvD1 receptor-deficient mice termed ALX/fpr2/3-/- . These investigations open new therapeutic avenues for inflammatory joint diseases, providing mechanistic substance for the benefits of omega-3 supplementation in RA.

Neutrophil-derived microvesicles enter cartilage and protect the joint in inflammatory arthritis.

Microvesicles (MVs) are emerging as a new mechanism of intercellular communication by transferring cellular lipid and protein components to target cells, yet their function in disease is only now being explored. We found that neutrophil-derived MVs were increased in concentration in synovial fluid from rheumatoid arthritis patients compared to paired plasma. Synovial MVs overexpressed the proresolving, anti-inflammatory protein annexin A1 (AnxA1). Mice deficient in TMEM16F, a lipid scramblase required for microvesiculation, exhibited exacerbated cartilage damage when subjected to inflammatory arthritis. To determine the function of MVs in inflammatory arthritis, toward the possibility of MV-based therapeutics, we examined the role of immune cell-derived MVs in rodent models and in human primary chondrocytes. In vitro, exogenous neutrophil-derived AnxA1(+) MVs activated anabolic gene expression in chondrocytes, leading to extracellular matrix accumulation and cartilage protection through the reduction in stress-adaptive homeostatic mediators interleukin-8 and prostaglandin E2. In vivo, intra-articular injection of AnxA1(+) MV lessened cartilage degradation caused by inflammatory arthritis. Arthritic mice receiving adoptive transfer of whole neutrophils displayed abundant MVs within cartilage matrix and revealed that MVs, but not neutrophils themselves, can penetrate cartilage. Mechanistic studies support a model whereby MV-associated AnxA1 interacts with its receptor FPR2 (formyl peptide receptor 2)/ALX, increasing transforming growth factor-ß production by chondrocytes, ultimately leading to cartilage protection. We envisage that MVs, either directly or loaded with therapeutics, can be harnessed as a unique therapeutic strategy for protection in diseases associated with cartilage degeneration.

Annexin A1-containing extracellular vesicles and polymeric nanoparticles promote epithelial wound repair.

Epithelial restitution is an essential process that is required to repair barrier function at mucosal surfaces following injury. Prolonged breaches in epithelial barrier function result in inflammation and further damage; therefore, a better understanding of the epithelial restitution process has potential for improving the development of therapeutics. In this work, we demonstrate that endogenous annexin A1 (ANXA1) is released as a component of extracellular vesicles (EVs) derived from intestinal epithelial cells, and these ANXA1-containing EVs activate wound repair circuits. Compared with healthy controls, patients with active inflammatory bowel disease had elevated levels of secreted ANXA1-containing EVs in sera, indicating that ANXA1-containing EVs are systemically distributed in response to the inflammatory process and could potentially serve as a biomarker of intestinal mucosal inflammation. Local intestinal delivery of an exogenous ANXA1 mimetic peptide (Ac2-26) encapsulated within targeted polymeric nanoparticles (Ac2-26 Col IV NPs) accelerated healing of murine colonic wounds after biopsy-induced injury. Moreover, one-time systemic administration of Ac2-26 Col IV NPs accelerated recovery following experimentally induced colitis. Together, our results suggest that local delivery of proresolving peptides encapsulated within nanoparticles may represent a potential therapeutic strategy for clinical situations characterized by chronic mucosal injury, such as is seen in patients with IBD.

Effects of the presence and community composition of earthworms on microbial community functioning.

Earthworms are a major component of many terrestrial ecosystems. By modifying decomposition processes and soil structure, they function as driving factors of the soil microbial community. Using microcosms, we investigated the effects of the presence and community composition of earthworms on the in situ respiratory response of a microbial community to an array of organic substrates including carbohydrates, amino acids, a polymer and an amide. Both the actual in situ catabolic response of non-growing microorganisms and the potential response of growing microorganisms were investigated. Three questions were studied: (1) does the presence of one of the main functional groups of earthworms (endogeic species) affect microbial community functioning; (2) does the presence of two functional groups (endogeic and epigeic species) alter microbial community functioning; (3) does the number of species within functional groups matter. The presence of endogeic earthworms significantly reduced microbial biomass and affected the physiological profile and functioning of the microbial community. In contrast, in the presence of endo- and epigeic species microbial biomass was not reduced significantly, indicating that epigeic species counteracted the effect of endogeic species. The physiological profile of the microbial community significantly differed between the treatment with endogeic species only and the treatment with both endo- and epigeic species. Also, the physiological profile of the microbial community was significantly affected by the number of species per functional group, which at least in part may have been caused by a sampling effect. Overall, the actual in situ catabolic response of non-growing microorganisms appears to be more sensitive than the potential response of growing microorganisms. In addition, the direction of the actual response (negative) was diametrically opposed to that of the potential response (positive). We conclude that the catabolic response of growing microorganisms does not reflect the actual case in situ. For earthworms, loss in both species number and functional group number has the potential to change soil microbial community functioning.

Phenylalanine 4-monooxygenase and the S-oxidation of S-carboxymethyl-L-cysteine.

The identity of the enzyme responsible for the S-oxidation of the mucolytic S-substituted L-cysteine drug, S-carboxymethyl-L-cysteine (SCMC), has been actively investigated for the last 10 years. A genetic polymorphism exists in the oxidation of the thioether moiety that has been identified as a disease susceptibility factor in a number of degenerative diseases. This polymorphism has also been implicated in the wide variation in clinical response to SCMC therapy in man. To date little is known about the molecular enzymology of this reaction but a previous investigation revealed that rat activated phenylalanine 4-monooxygenase (PAH) could S-oxidise both Met- and S-methyl-L-cysteine (SMC) to their S-oxide metabolites. We have investigated the hypothesis that SCMC was also a substrate for activated PAH in the cytosolic faction of the Wistar rat. 1. Substrate and inhibitor investigation revealed that SCMC was a substrate for activated PAH activity in vitro. 2. The large aromatic amino acid hydroxylase monoclonal antibody and the Fe3+ chelator, deferoxamine, completely inhibited both Phe and SCMC oxidation to their respective metabolites. 3. Analysis of the Dixon plots revealed that both Phe and SCMC competitively inhibited each other's oxidation. 4. Correlation studies showed that the rate of production of Tyr was positively correlated to the production of both SCMC and SMC S-oxides in 20 female Wistar rat hepatic cytosolic fractions. These results strongly support the hypothesis that PAH is the enzyme responsible for SCMC S-oxidation in the rat.

Cutting-edge analysis of extracellular microparticles using ImageStream(X) imaging flow cytometry.

Interest in extracellular vesicle biology has exploded in the past decade, since these microstructures seem endowed with multiple roles, from blood coagulation to inter-cellular communication in pathophysiology. In order for microparticle research to evolve as a preclinical and clinical tool, accurate quantification of microparticle levels is a fundamental requirement, but their size and the complexity of sample fluids present major technical challenges. Flow cytometry is commonly used, but suffers from low sensitivity and accuracy. Use of Amnis ImageStream(X) Mk II imaging flow cytometer afforded accurate analysis of calibration beads ranging from 1 µm to 20 nm; and microparticles, which could be observed and quantified in whole blood, platelet-rich and platelet-free plasma and in leukocyte supernatants. Another advantage was the minimal sample preparation and volume required. Use of this high throughput analyzer allowed simultaneous phenotypic definition of the parent cells and offspring microparticles along with real time microparticle generation kinetics. With the current paucity of reliable techniques for the analysis of microparticles, we propose that the ImageStream(X) could be used effectively to advance this scientific field.