The 5-Phosphatase SHIP2 Promotes Neutrophil Chemotaxis and Recruitment

Neutrophils, the most abundant circulating leukocytes in humans have key roles in host defense and in the inflammatory response. Agonist-activated phosphoinositide 3-kinases (PI3Ks) are important regulators of many facets of neutrophil biology. PIP3 is subject to dephosphorylation by several 5' phosphatases, including SHIP family phosphatases, which convert the PI3K product and lipid second messenger phosphatidylinositol 3,4,5-trisphosphate (PIP3) into PI(3,4)P2, a lipid second messenger in its own right. In addition to the leukocyte restricted SHIP1, neutrophils express the ubiquitous SHIP2. This study analyzed mice and isolated neutrophils carrying a catalytically inactive SHIP2, identifying an important regulatory function in neutrophil chemotaxis and directionality in vitro and in neutrophil recruitment to sites of sterile inflammation in vivo, in the absence of major defects of any other neutrophil functions analyzed, including, phagocytosis and the formation of reactive oxygen species. Mechanistically, this is explained by a subtle effect on global 3-phosphorylated phosphoinositide species. This work identifies a non-redundant role for the hitherto overlooked SHIP2 in the regulation of neutrophils, and specifically, neutrophil chemotaxis/trafficking. It completes an emerging wider understanding of the complexity of PI3K signaling in the neutrophil, and the roles played by individual kinases and phosphatases within

Boundary cap cells constrain spinal motor neuron somal migration at motor exit points by a semaphorin-plexin mechanism.

RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are.BACKGROUND: In developing neurons, somal migration and initiation of axon outgrowth often occur simultaneously and are regulated in part by similar classes of molecules. When neurons reach their final destinations, however, somal translocation and axon extension are uncoupled. Insights into the mechanisms underlying this process of disengagement came from our study of the behaviour of embryonic spinal motor neurons following ablation of boundary cap cells. These are neural crest derivatives that transiently reside at motor exit points, central nervous system (CNS):peripheral nervous system (PNS) interfaces where motor axons leave the CNS. In the absence of boundary cap cells, motor neuron cell bodies migrate along their axons into the periphery, suggesting that repellent signals from boundary cap cells regulate the selective gating of somal migration and axon outgrowth at the motor exit point. Here we used RNA interference in the chick embryo together with analysis of null mutant mice to identify possible boundary cap cell ligands, their receptors on motor neurons and cytoplasmic signalling molecules that control this process. RESULTS: We demonstrate that targeted knock down in motor neurons of Neuropilin-2 (Npn-2), a high affinity receptor for class 3 semaphorins, causes their somata to migrate to ectopic positions in ventral nerve roots. This finding was corroborated in Npn-2 null mice, in which we identified motor neuron cell bodies in ectopic positions in the PNS. Our RNA interference studies further revealed a role for Plexin-A2, but not Plexin-A1 or Plexin-A4. We show that chick and mouse boundary cap cells express Sema3B and 3G, secreted semaphorins, and Sema6A, a transmembrane semaphorin. However, no increased numbers of ectopic motor neurons are found in Sema3B null mouse embryos. In contrast, Sema6A null mice display an ectopic motor neuron phenotype. Finally, knockdown of MICAL3, a downstream semaphorin/Plexin-A signalling molecule, in chick motor neurons led to their ectopic positioning in the PNS. CONCLUSION: We conclude that semaphorin-mediated repellent interactions between boundary cap cells and immature spinal motor neurons regulates somal positioning by countering the drag exerted on motor neuron cell bodies by their axons as they emerge from the CNS at motor exit points. Our data support a model in which BC cell semaphorins signal through Npn-2 and/or Plexin-A2 receptors on motor neurons via a cytoplasmic effector, MICAL3, to trigger cytoskeletal reorganisation. This leads to the disengagement of somal migration from axon extension and the confinement of motor neuron cell bodies to the spinal cord

Stromal Cells Covering Omental Fat-Associated Lymphoid Clusters Trigger Formation of Neutrophil Aggregates to Capture Peritoneal Contaminants

The omentum is a visceral adipose tissue rich in fat-associated lymphoid clusters (FALCs) that collects peritoneal contaminants and provides a first layer of immunological defense within the abdomen. Here, we investigated the mechanisms that mediate the capture of peritoneal contaminants during peritonitis. Single-cell RNA sequencing and spatial analysis of omental stromal cells revealed that the surface of FALCs were covered by CXCL1+ mesothelial cells, which we termed FALC cover cells. Blockade of CXCL1 inhibited the recruitment and aggregation of neutrophils at FALCs during zymosan-induced peritonitis. Inhibition of protein arginine deiminase 4, an enzyme important for the release of neutrophil extracellular traps, abolished neutrophil aggregation and the capture of peritoneal contaminants by omental FALCs. Analysis of omental samples from patients with acute appendicitis confirmed neutrophil recruitment and bacterial capture at FALCs. Thus, specialized omental mesothelial cells coordinate the recruitment and aggregation of neutrophils to capture peritoneal contaminants

TCR-stimulated changes in cell surface CD46 expression generate type 1 regulatory T cells

International audienc

Chemical modulation of <i>in vivo</i> macrophage function with subpopulation-specific fluorescent prodrug conjugates

Immunomodulatory agents represent one of the most promising strategies for enhancing tissue regeneration without the side effects of traditional drug-based therapies. Tissue repair depends largely on macrophages, making them ideal targets for proregenerative therapies. However, given the multiple roles of macrophages in tissue homeostasis, small molecule drugs must be only active in very specific subpopulations. In this work, we have developed the first prodrug–fluorophore conjugates able to discriminate closely related subpopulations of macrophages (i.e., proinflammatory M1 vs anti-inflammatory M2 macrophages), and employed them to deplete M1 macrophages <i>in vivo</i> without affecting other cell populations. Selective intracellular activation and drug release enabled simultaneous fluorescence cell tracking and ablation of M1 macrophages <i>in vivo</i>, with the concomitant rescue of a proregenerative phenotype. <i>Ex vivo</i> assays in human monocyte-derived macrophages validate the translational potential of this novel platform to develop chemical immunomodulatory agents as targeted therapies for immune-related diseases

Alternatively activated macrophages promotes necrosis resolution following acute liver injury

Background & Aim Following acetaminophen (APAP) overdose, acute liver injury (ALI) can occur in patients that present too late for N-acetylcysteine treatment, potentially leading to acute liver failure, systemic inflammation, and death. Macrophages influence the progression and resolution of ALI due to their innate immunological function and paracrine activity. Syngeneic primary bone marrow-derived macrophages (BMDMs) were tested as a cell-based therapy in a mouse model of APAP-induced ALI (APAP-ALI). Methods Several phenotypically distinct BMDM populations were delivered intravenously to APAP-ALI mice when hepatic necrosis was established, and then evaluated based on their effects on injury, inflammation, immunity, and regeneration. In vivo phagocytosis assays were used to interrogate the phenotype and function of alternatively activated BMDMs (AAMs) post-injection. Finally, primary human AAMs sourced from healthy volunteers were evaluated in immunocompetent APAP-ALI mice. Results BMDMs rapidly localised to the liver and spleen within 4 h of administration. Injection of AAMs specifically reduced hepatocellular necrosis, HMGB1 translocation, and infiltrating neutrophils following APAP-ALI. AAM delivery also stimulated proliferation in hepatocytes and endothelium, and reduced levels of several circulating proinflammatory cytokines within 24 h. AAMs displayed a high phagocytic activity both in vitro and in injured liver tissue post-injection. Crosstalk with the host innate immune system was demonstrated by reduced infiltrating host Ly6Chi macrophages in AAM-treated mice. Importantly, therapeutic efficacy was partially recapitulated using clinical-grade primary human AAMs in immunocompetent APAP-ALI mice, underscoring the translational potential of these findings. Conclusion We identify that AAMs have value as a cell-based therapy in an experimental model of APAP-ALI. Human AAMs warrant further evaluation as a potential cell-based therapy for APAP overdose patients with established liver injury. Lay summary After an overdose of acetaminophen (paracetamol), some patients present to hospital too late for the current antidote (N-acetylcysteine) to be effective. We tested whether macrophages, an injury-responsive leukocyte that can scavenge dead/dying cells, could serve as a cell-based therapy in an experimental model of acetaminophen overdose. Injection of alternatively activated macrophages rapidly reduced liver injury and reduced several mediators of inflammation. Macrophages show promise to serve as a potential cell-based therapy for acute liver injury

Osteoclast stimulation factor 1 (Ostf1) KNOCKOUT increases trabecular bone mass in mice

Osteoclast stimulation factor 1 (OSTF1) is an SH3-domain containing protein that was initially identified as a factor involved in the indirect activation of osteoclasts. It has been linked to spinal muscular atrophy in humans through its interaction with SMN1, and is one of six genes deleted in a human developmental microdeletion syndrome. To investigate the function of OSTF1, we generated an Ostf1 knockout mouse model, with exons 3 and 4 of Ostf1 replaced by a LacZ orf. Extensive X-Gal staining was performed to examine the developmental and adult expression pattern, followed by phenotyping. We show widespread expression of the gene in the vasculature of most organs and in a number of cell types in adult and embryonic mouse tissues. Furthermore, whilst SHIRPA testing revealed no behavioural defects, we demonstrate increased trabecular mass in the long bones, confirming a role for OSTF1 in bone development

Boundary cap cells constrain spinal motor neuron somal migration at motor exit points by a semaphorin-plexin mechanism.

RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are.BACKGROUND: In developing neurons, somal migration and initiation of axon outgrowth often occur simultaneously and are regulated in part by similar classes of molecules. When neurons reach their final destinations, however, somal translocation and axon extension are uncoupled. Insights into the mechanisms underlying this process of disengagement came from our study of the behaviour of embryonic spinal motor neurons following ablation of boundary cap cells. These are neural crest derivatives that transiently reside at motor exit points, central nervous system (CNS):peripheral nervous system (PNS) interfaces where motor axons leave the CNS. In the absence of boundary cap cells, motor neuron cell bodies migrate along their axons into the periphery, suggesting that repellent signals from boundary cap cells regulate the selective gating of somal migration and axon outgrowth at the motor exit point. Here we used RNA interference in the chick embryo together with analysis of null mutant mice to identify possible boundary cap cell ligands, their receptors on motor neurons and cytoplasmic signalling molecules that control this process. RESULTS: We demonstrate that targeted knock down in motor neurons of Neuropilin-2 (Npn-2), a high affinity receptor for class 3 semaphorins, causes their somata to migrate to ectopic positions in ventral nerve roots. This finding was corroborated in Npn-2 null mice, in which we identified motor neuron cell bodies in ectopic positions in the PNS. Our RNA interference studies further revealed a role for Plexin-A2, but not Plexin-A1 or Plexin-A4. We show that chick and mouse boundary cap cells express Sema3B and 3G, secreted semaphorins, and Sema6A, a transmembrane semaphorin. However, no increased numbers of ectopic motor neurons are found in Sema3B null mouse embryos. In contrast, Sema6A null mice display an ectopic motor neuron phenotype. Finally, knockdown of MICAL3, a downstream semaphorin/Plexin-A signalling molecule, in chick motor neurons led to their ectopic positioning in the PNS. CONCLUSION: We conclude that semaphorin-mediated repellent interactions between boundary cap cells and immature spinal motor neurons regulates somal positioning by countering the drag exerted on motor neuron cell bodies by their axons as they emerge from the CNS at motor exit points. Our data support a model in which BC cell semaphorins signal through Npn-2 and/or Plexin-A2 receptors on motor neurons via a cytoplasmic effector, MICAL3, to trigger cytoskeletal reorganisation. This leads to the disengagement of somal migration from axon extension and the confinement of motor neuron cell bodies to the spinal cord