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The Aedes aegypti mosquito is a vector of many infectious agents, including flaviviruses such as Zika virus. Components of mosquito saliva have pleomorphic effects on the vertebrate host to enhance blood feeding, and these changes also create a favorable niche for pathogen replication and dissemination. Here, we demonstrate that human CD47, which is known to be involved in various immune processes, interacts with a 34-kilodalton mosquito salivary protein named Nest1. Nest1 is up-regulated in blood-fed female A. aegypti and facilitates Zika virus dissemination in human skin explants. Nest1 has a stronger affinity for CD47 than its natural ligand, signal regulatory protein α, competing for binding at the same interface. The interaction between Nest1 with CD47 suppresses phagocytosis by human macrophages and inhibits proinflammatory responses by white blood cells, thereby suppressing antiviral responses in the skin. This interaction elucidates how an arthropod protein alters the human response to promote arbovirus infectivity.

Background: Incidences of inflammatory bowel disease (IBD), including Crohn's disease and ulcerative colitis, are escalating worldwide and can be considered a global public health problem. Given that the gold standard approach to IBD therapeutics focuses on reducing the severity of symptoms, there is an urgent unmet need to develop alternative therapies that halt not only inflammatory processes but also promote mucosal repair. Previous studies have identified increased stem cell factor (SCF) expression in inflamed intestinal mucosal tissues. However, the role that SCF plays in mediating intestinal inflammation and repair has not been explored., Methods: Changes in the expression of SCF were evaluated in the colonic tissue of healthy mice and during dextran sodium sulfate (DSS)-induced colitis. Furthermore, mucosal wound healing and colitis severity were analyzed in mice subjected to either mechanical biopsy or DSS treatment, respectively, following intestinal epithelial cell-specific deletion of SCF or anti-SCF antibody administration., Results: We report robust expression of SCF by intestinal epithelial cells during intestinal homeostasis with a switch to immune cell-produced SCF during colitis. Data from mice with intestinal epithelial cell-specific deletion of SCF highlight the importance of immune cell-produced SCF in driving the pathogenesis of colitis. Importantly, antibody-mediated neutralization of total SCF or the specific SCF248 isoform decreased immune cell infiltration and enhanced mucosal wound repair following biopsy-induced colonic injury or DSS-induced colitis., Conclusions: These data demonstrate that SCF functions as a pro-inflammatory mediator in mucosal tissues and that specific neutralization of SCF248 could be a viable therapeutic option to reduce intestinal inflammation and promote mucosal wound repair in individuals with IBD., (© The Author(s) 2023. Published by Oxford University Press on behalf of Crohn’s & Colitis Foundation. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Dysregulated neutrophil (PMN) transmigration across epithelial surfaces (TEpM) significantly contributes to chronic inflammatory diseases, yet mechanisms defining this process remain poorly understood. In the intestine, uncontrolled PMN TEpM is a hallmark of disease flares in ulcerative colitis. Previous in vitro studies directed at identifying molecular determinants that mediate TEpM have shown that plasma membrane proteins including CD47 and CD11b/CD18 play key roles in regulating PMN TEpM across monolayers of intestinal epithelial cells. Here, we show that CD47 modulates PMN TEpM in vivo using an ileal loop assay. Importantly, using novel tissue-specific CD47 knockout mice and in vitro approaches, we report that PMN-expressed, but not epithelial-expressed CD47 plays a major role in regulating PMN TEpM. We show that CD47 associates with CD11b/CD18 in the plasma membrane of PMN, and that loss of CD47 results in impaired CD11b/CD18 activation. In addition, in vitro and in vivo studies using function blocking antibodies support a role of CD47 in regulating CD11b-dependent PMN TEpM and chemotaxis. Taken together, these findings provide new insights for developing approaches to target dysregulated PMN infiltration in the intestine. Moreover, tissue-specific CD47 knockout mice constitute an important new tool to study contributions of cells expressing CD47 to inflammation in vivo.

Polymorphonuclear neutrophils (PMNs) play a critical role in clearing invading microbes and promoting tissue repair following infection/injury. However, dysregulated PMN trafficking and associated tissue damage is pathognomonic of numerous inflammatory mucosal diseases. The final step in PMN influx into mucosal lined organs (including the lungs, kidneys, skin, and gut) involves transepithelial migration (TEpM). The β2-integrin CD11b/CD18 plays an important role in mediating PMN intestinal trafficking, with recent studies highlighting that terminal fucose and GlcNAc glycans on CD11b/CD18 can be targeted to reduce TEpM. However, the role of the most abundant terminal glycan, sialic acid (Sia), in regulating PMN epithelial influx and mucosal inflammatory function is not well understood. Here we demonstrate that inhibiting sialidase-mediated removal of α2-3-linked Sia from CD11b/CD18 inhibits PMN migration across intestinal epithelium in vitro and in vivo. Sialylation was also found to regulate critical PMN inflammatory effector functions, including degranulation and superoxide release. Finally, we demonstrate that sialidase inhibition reduces bacterial peptide-mediated CD11b/CD18 activation in PMN and blocks downstream intracellular signaling mediated by spleen tyrosine kinase (Syk) and p38 MAPK. These findings suggest that sialylated glycans on CD11b/CD18 represent potentially novel targets for ameliorating PMN-mediated tissue destruction in inflammatory mucosal diseases.

Clinical symptoms in many inflammatory diseases of the intestine are directly related to neutrophil (PMN) migration across colonic mucosa and into the intestinal lumen, yet in-vivo studies detailing this process are lacking. Using real-time intravital microscopy and a new distal colon loop model, we report distinct PMN migratory dynamics in response to several models of acute colonic injury. PMNs exhibited rapid swarming responses after mechanically induced intestinal wounds. Similar numbers of PMNs infiltrated colonic mucosa after wounding in germ-free mice, suggesting microbiota-independent mechanisms. By contrast, acute mucosal injury secondary to either a treatment of mice with dextran sodium sulfate or an IL-10 receptor blockade model of colitis resulted in lamina propria infiltration with PMNs that were largely immotile. Biopsy wounding of colonic mucosa in DSS-treated mice did not result in enhanced PMN swarming however, intraluminal application of the neutrophil chemoattractant LTB 4 under such conditions resulted in enhanced transepithelial migration of PMNs. Analyses of PMNs that had migrated into the colonic lumen revealed that the majority of PMNs were directly recruited from the circulation and not from the immotile pool in the mucosa. Decreased PMN motility parallels upregulation of the receptor CXCR4 and apoptosis. Similarly, increased expression of CXCR4 on human PMNs was observed in colonic biopsies from people with active ulcerative colitis. This new approach adds an important tool to investigate mechanisms regulating PMN migration across mucosa within the distal intestine and will provide new insights for developing future anti-inflammatory and pro-repair therapies., (© The Author(s) 2022. Published by Oxford University Press on behalf of National Academy of Sciences.)

Dysregulated healing of injured mucosa is a hallmark of many pathological conditions, including inflammatory bowel disease. Mucosal injury and chronic intestinal inflammation are also associated with alterations in epithelial glycosylation. Previous studies have revealed that inflammation-induced glycan sialyl Lewis A on epithelial CD44v6 acts as a ligand for transmigrating PMNs. Here we report that robust sialylated Lewis glycan expression was induced in colonic mucosa from individuals with ulcerative colitis and Crohn disease as well as in the colonic epithelium of mice with colitis induced by dextran sodium sulfate (DSS). Targeting of sialylated epithelial Lewis glycans with mAb GM35 reduced disease activity and improved mucosal integrity during DSS-induced colitis in mice. Wound healing studies revealed increased epithelial proliferation and migration responses as well as improved mucosal repair after ligation of epithelial sialyl Lewis glycans. Finally, we showed that GM35-mediated increases in epithelial proliferation and migration were mediated through activation of kinases that signal downstream of CD44v6 (Src, FAK, Akt). These findings suggest that sialylated Lewis glycans on CD44v6 represent epithelial targets for improved recovery of intestinal barrier function and restitution of mucosal homeostasis after inflammation or injury.

Polymorphonuclear neutrophils (PMNs) play a critical role in the innate immune response to invading pathogens. However, dysregulated mucosal trafficking of PMNs and associated epithelial tissue damage is a pathological hallmark of numerous inflammatory conditions including inflammatory bowel disease. The glycoprotein CD11b/CD18 plays a well-described role in regulating PMN transepithelial migration and PMN inflammatory functions. Previous studies have demonstrated that targeting of the N-linked glycan Lewis X on CD11b blocks PMN transepithelial migration (TEpM). Given evidence of glycosylation-dependent regulation of CD11b/CD18 function, we performed MALDI TOF Mass Spectrometry (MS) analyses on CD11b/CD18 purified from human PMNs. Unusual glycan epitopes identified on CD11b/CD18 included high Mannose oligosaccharides recognized by the Galanthus Nivalis lectin and biantennary galactosylated N-glycans recognized by the Phaseolus Vulgaris erythroagglutinin lectin. Importantly, we show that selective targeting of glycans on CD11b with such lectins results in altered intracellular signaling events that inhibit TEpM and differentially affect key PMN inflammatory functions including phagocytosis, superoxide release and apoptosis. Taken together, these data demonstrate that discrete glycan motifs expressed on CD11b/CD18 such as biantennary galactose could represent novel targets for selective manipulation of CD11b function and reduction of PMN-associated tissue damage in chronic inflammatory diseases., (© 2019 Federation of American Societies for Experimental Biology.)

Mucosal wound repair is coordinated by dynamic crosstalk between endogenous and exogenous mediators and specific receptors on epithelial cells and infiltrating immune cells. One class of such receptor-ligand pairs involves formyl peptide receptors (FPRs) that have been shown to influence inflammatory response and repair. Here we explored the role of murine Fpr2/3, an ortholog of human FPR2/receptor for lipoxin A4 (ALX), in orchestrating intestinal mucosal repair. Compared with wild-type (WT) mice, Fpr2/3 -/- mice exhibited delayed recovery from acute experimental colitis and perturbed repair after biopsy-induced colonic mucosal injury. Decreased numbers of infiltrating monocytes were observed in healing wounds from Fpr2/3 -/- mice compared with WT animals. Bone marrow transplant experiments revealed that Fpr2/3 -/- monocytes showed a competitive disadvantage when infiltrating colonic wounds. Moreover, Fpr2/3 -/- monocytes were defective in chemotactic responses to the chemokine CC chemokine ligand (CCL)20, which is up-regulated during early phases of inflammation. Analysis of Fpr2/3 -/- monocytes revealed altered expression of the CCL20 receptor CC chemokine receptor (CCR)6, suggesting that Fpr2/3 regulates CCL20-CCR6-mediated monocyte chemotaxis to sites of mucosal injury in the gut. These findings demonstrate an important contribution of Fpr2/3 in facilitating monocyte recruitment to sites of mucosal injury to influence wound repair.-Birkl, D., O'Leary, M. N., Quiros, M., Azcutia, V., Schaller, M., Reed, M., Nishio, H., Keeney, J., Neish, A. S., Lukacs, N. W., Parkos, C. A., Nusrat, A. Formyl peptide receptor 2 regulates monocyte recruitment to promote intestinal mucosal wound repair.

CD47 is a ubiquitously expressed transmembrane glycoprotein that regulates inflammatory responses and tissue repair. Here, we show that normal mice treated with anti-CD47 antibodies, and Cd47-null mice have impaired intestinal mucosal wound healing. Furthermore, intestinal epithelial cell (IEC)-specific loss of CD47 does not induce spontaneous immune-mediated intestinal barrier disruption but results in defective mucosal repair after biopsy-induced colonic wounding or Dextran Sulfate Sodium (DSS)-induced mucosal damage. In vitro analyses using primary cultures of CD47-deficient murine colonic IEC or human colonoid-derived IEC treated with CD47-blocking antibodies demonstrate impaired epithelial cell migration in wound healing assays. Defective wound repair after CD47 loss is linked to decreased epithelial β1 integrin and focal adhesion signaling, as well as reduced thrombospondin-1 and TGF-β1. These results demonstrate a critical role for IEC-expressed CD47 in regulating mucosal repair and raise important considerations for possible alterations in wound healing secondary to therapeutic targeting of CD47.

Junctional adhesion molecule-A (JAM-A) is a transmembrane glycoprotein expressed on leukocytes, endothelia, and epithelia that regulates biological processes including barrier function and immune responses. While JAM-A has been reported to facilitate tissue infiltration of leukocytes under inflammatory conditions, the contributions of leukocyte-expressed JAM-A in vivo remain unresolved. We investigated the role of leukocyte-expressed JAM-A in acute peritonitis induced by zymosan, lipopolysaccharide (LPS), or TNFα using mice with selective loss of JAM-A in myelomonocytic cells (LysM-Cre;Jam-a fl/fl ). Surprisingly, in LysM-Cre;Jam-a fl/fl mice, loss of JAM-A did not affect neutrophil (PMN) recruitment into the peritoneum in response to zymosan, LPS, or TNFα although it was significantly reduced in Jam-a KO mice. In parallel, Jam-a KO peritoneal macrophages exhibited diminished CXCL1 chemokine production and decreased activation of NF-kB, whereas those from LysM-Cre;Jam-a fl/fl mice were unaffected. Using Villin-Cre;Jam-a fl/fl mice, targeted loss of JAM-A on intestinal epithelial cells resulted in increased intestinal permeability along with reduced peritoneal PMN migration as well as lower levels of CXCL1 and active NF-kB similar to that observed in Jam-a KO animals. Interestingly, in germ-free Villin-Cre;Jam-a fl/fl mice, PMN recruitment was unaffected suggesting dependence on gut microbiota. Such observations highlight the functional link between a leaky gut and regulation of innate immune responses.

Polymorphonuclear neutrophils (PMNs) play a critical role in host defense against infection and in the resolution of inflammation. However, immune responses mediated by PMN must be tightly regulated to facilitate elimination of invading pathogens without inducing detrimental inflammation and host tissue damage. Specific engagement of cell surface immunoreceptors by a diverse range of extracellular signals regulates PMN effector functions through differential activation of intracellular signaling cascades. Although mechanisms of PMN activation mediated via cell signaling pathways have been well described, less is known about negative regulation of PMN function by immune signaling cascades. Here, we provide an overview of immunoreceptor-mediated negative regulation of key PMN effector functions including maturation, migration, phagocytosis, reactive oxygen species release, degranulation, apoptosis, and NET formation. Increased understanding of mechanisms of suppression of PMN effector functions may point to possible future therapeutic targets for the amelioration of PMN-mediated autoimmune and inflammatory diseases., (©2017 Society for Leukocyte Biology.)

In response to injury, epithelial cells migrate and proliferate to cover denuded mucosal surfaces and repair the barrier defect. This process is orchestrated by dynamic crosstalk between immune cells and the epithelium; however, the mechanisms involved remain incompletely understood. Here, we report that IL-10 was rapidly induced following intestinal mucosal injury and was required for optimal intestinal mucosal wound closure. Conditional deletion of IL-10 specifically in CD11c-expressing cells in vivo implicated macrophages as a critical innate immune contributor to IL-10-induced wound closure. Consistent with these findings, wound closure in T cell- and B cell-deficient Rag1-/- mice was unimpaired, demonstrating that adaptive immune cells are not absolutely required for this process. Further, following mucosal injury, macrophage-derived IL-10 resulted in epithelial cAMP response element-binding protein (CREB) activation and subsequent synthesis and secretion of the pro-repair WNT1-inducible signaling protein 1 (WISP-1). WISP-1 induced epithelial cell proliferation and wound closure by activating epithelial pro-proliferative pathways. These findings define the involvement of macrophages in regulating an IL-10/CREB/WISP-1 signaling axis, with broad implications in linking innate immune activation to mucosal wound repair.

CD47 is known to play an important role in CD4 + T cell homeostasis. We recently reported a reduction in mice deficient in the Cd47 gene (Cd47 -/- ) CD4 + T cell adhesion and transendothelial migration (TEM) in vivo and in vitro as a result of impaired expression of high-affinity forms of LFA-1 and VLA-4 integrins. A prior study concluded that Cd47 -/- mice were resistant to experimental autoimmune encephalomyelitis (EAE) as a result of complete failure in CD4 + T cell activation after myelin oligodendrocyte glycoprotein peptide 35-55 aa (MOG 35-55 ) immunization. As the prior EAE study was published before our report, authors could not have accounted for defects in T cell integrin function as a mechanism to protect Cd47 -/- in EAE. Thus, we hypothesized that failure of T cell activation involved defects in LFA-1 and VLA-4 integrins. We confirmed that Cd47 -/- mice were resistant to MOG 35-55 -induced EAE. Our data, however, supported a different mechanism that was not a result of failure of CD4 + T cell activation. Instead, we found that CD4 + T cells in MOG 35-55 -immunized Cd47 -/- mice were activated, but clonal expansion contracted within 72 h after immunization. We used TCR crosslinking and mitogen activation in vitro to investigate the underlying mechanism. We found that naïve Cd47 -/- CD4 + T cells exhibited a premature block in proliferation and survival because of impaired activation of LFA-1, despite effective TCR-induced activation. These results identify CD47 as an important regulator of LFA-1 and VLA-4 integrin-adhesive functions in T cell proliferation, as well as recruitment, and clarify the roles played by CD47 in MOG 35-55 -induced EAE., (© Society for Leukocyte Biology.)

Wiskott-Aldrich syndrome (WAS) is associated with mutations in the WAS protein (WASp), which plays a critical role in the initiation of T cell receptor-driven (TCR-driven) actin polymerization. The clinical phenotype of WAS includes susceptibility to infection, allergy, autoimmunity, and malignancy and overlaps with the symptoms of dedicator of cytokinesis 8 (DOCK8) deficiency, suggesting that the 2 syndromes share common pathogenic mechanisms. Here, we demonstrated that the WASp-interacting protein (WIP) bridges DOCK8 to WASp and actin in T cells. We determined that the guanine nucleotide exchange factor activity of DOCK8 is essential for the integrity of the subcortical actin cytoskeleton as well as for TCR-driven WASp activation, F-actin assembly, immune synapse formation, actin foci formation, mechanotransduction, T cell transendothelial migration, and homing to lymph nodes, all of which also depend on WASp. These results indicate that DOCK8 and WASp are in the same signaling pathway that links TCRs to the actin cytoskeleton in TCR-driven actin assembly. Further, they provide an explanation for similarities in the clinical phenotypes of WAS and DOCK8 deficiency.

Background: Hyperglycemia is acknowledged as a pro-inflammatory condition and a major cause of vascular damage. Nevertheless, we have previously described that high glucose only promotes inflammation in human vascular cells previously primed with pro-inflammatory stimuli, such as the cytokine interleukin (IL)1β. Here, we aimed to identify the cellular mechanisms by which high glucose exacerbates the vascular inflammation induced by IL1β., Methods: Cultured human aortic smooth muscle cells (HASMC) and isolated rat mesenteric microvessels were treated with IL1β in medium containing 5.5-22 mmol/L glucose. Glucose uptake and consumption, lactate production, GLUT1 levels, NADPH oxidase activity and inflammatory signalling (nuclear factor-κB activation and inducible nitric oxide synthase expression) were measured in HASMC, while endothelium-dependent relaxations to acetylcholine were determined in rat microvessels. Pharmacological inhibition of IL1 receptors, NADPH oxidase and glucose-6-phosphate dehydrogenase (G6PD), as well as silencing of G6PD, were also performed. Moreover, the pentose phosphate pathway (PPP) activity and the levels of reduced glutathione were determined., Results: We found that excess glucose uptake in HASMC cultured in 22 mM glucose only occurred following activation with IL1β. However, the simple entry of glucose was not enough to be deleterious since over-expression of the glucose transporter GLUT1 or increased glucose uptake following inhibition of mitochondrial respiration by sodium azide was not sufficient to trigger inflammatory mechanisms. In fact, besides allowing glucose entry, IL1β activated the PPP, thus permitting some of the excess glucose to be metabolized via this route. This in turn led to an over-activation NADPH oxidase, resulting in increased generation of free radicals and the subsequent downstream pro-inflammatory signalling. Moreover, in rat mesenteric microvessels high glucose incubation enhanced the endothelial dysfunction induced by IL1β by a mechanism which was abrogated by the inhibition of the PPP., Conclusions: A pro-inflammatory stimulus like IL1β transforms excess glucose into a vascular deleterious agent by causing an increase in glucose uptake and its subsequent diversion into the PPP, promoting the pro-oxidant conditions required for the exacerbation of pro-oxidant and pro-inflammatory pathways. We propose that over-activation of the PPP is a crucial mechanism for the vascular damage associated to hyperglycemia.

The mechanisms driving T cell homing to lymph nodes and migration to tissue are well described but little is known about factors that affect T cell egress from tissues. Here, we generate mice with a T cell-specific deletion of the scaffold protein A kinase anchoring protein 9 (AKAP9) and use models of inflammatory disease to demonstrate that AKAP9 is dispensable for T cell priming and migration into tissues and lymph nodes, but is required for T cell retention in tissues. AKAP9 deficiency results in increased T cell egress to draining lymph nodes, which is associated with impaired T cell re-activation in tissues and protection from organ damage. AKAP9-deficient T cells exhibit reduced microtubule-dependent recycling of TCRs back to the cell surface and this affects antigen-dependent activation, primarily by non-classical antigen-presenting cells. Thus, AKAP9-dependent TCR trafficking drives efficient T cell re-activation and extends their retention at sites of inflammation with implications for disease pathogenesis.

Previous studies have demonstrated that the cell surface receptor Slamf1 (CD150) is requisite for optimal NADPH-oxidase (Nox2) dependent reactive oxygen species (ROS) production by phagocytes in response to Gram- bacteria. By contrast, Slamf8 (CD353) is a negative regulator of ROS in response to Gram+ and Gram- bacteria. Employing in vivo migration after skin sensitization, induction of peritonitis, and repopulation of the small intestine demonstrates that in vivo migration of Slamf1-/- dendritic cells and macrophages is reduced, as compared to wt mice. By contrast, in vivo migration of Slamf8-/- dendritic cells, macrophages and neutrophils is accelerated. These opposing effects of Slamf1 and Slamf8 are cell-intrinsic as judged by in vitro migration in transwell chambers in response to CCL19, CCL21 or CSF-1. Importantly, inhibiting ROS production of Slamf8-/- macrophages by diphenyleneiodonium chloride blocks this in vitro migration. We conclude that Slamf1 and Slamf8 govern ROS-dependent innate immune responses of myeloid cells, thus modulating migration of these cells during inflammation in an opposing manner.

Leukocyte CD18 integrins increase their affinity for ligand by transmitting allosteric signals to and from their ligand-binding αI domain. Mechanical forces induce allosteric changes that paradoxically slow dissociation by increasing the integrin/ligand bond lifetimes, referred to as catch bonds. Mac-1 formed catch bonds with its ligands. However, a Mac-1 gene (ITGAM) coding variant (rs1143679, R77H), which is located in the β-propeller domain and is significantly associated with systemic lupus erythematosus risk, exhibits a marked impairment in 2D ligand affinity and affinity maturation under mechanical force. Targeted mutations and activating antibodies reveal that the failure in Mac-1 R77H allostery is rescued by induction of cytoplasmic tail separation and full integrin extension. These findings demonstrate roles for R77, and the β-propeller in which it resides, in force-induced allostery relay and integrin bond stabilization. Defects in these processes may have pathological consequences, as the Mac-1 R77H variant is associated with increased susceptibility to lupus., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

Inflammatory bowel disease (IBD) represents a group of recurrent chronic inflammatory disorders associated with autoimmune dysregulation, typically characterized by neutrophil infiltration and mucosal inflammatory lesions. Neutrophils, as the earliest immune cells to arrive at inflamed tissues, play a dual role in the onset and progression of mucosal inflammation in IBD. Most of these cells specifically express CD177, a molecule increasingly recognized for its critical role in the pathogenesis of IBD. Under IBD-related inflammatory stimuli, CD177 is highly expressed on neutrophils and promotes their migration. CD177 + neutrophils activate bactericidal and barrier-protective functions at IBD mucosal inflammation sites and regulate the release of inflammatory mediators highly correlated with the severity of inflammation in IBD patients, thus playing a dual role. However, mitigating the detrimental effects of neutrophils in inflammatory bowel disease remains a challenge. Based on these data, we have summarized recent articles on the role of neutrophils in intestinal inflammation, with a particular emphasis on CD177, which mediates the recruitment, transepithelial migration, and activation of neutrophils, as well as their functional consequences. A better understanding of CD177 + neutrophils may contribute to the development of novel therapeutic targets to selectively modulate the protective role of this class of cells in IBD. [ABSTRACT FROM AUTHOR]

Cell activation and nutrient dysregulation are common consequences of atherosclerosis and its preceding risk factors, such as hypertension, dyslipidemia, and diabetes. These diseases may also impact cellular metabolism and consequently cell function, and the other way around, altered cellular metabolism can impact disease development and progression through altered cell function. Understanding the contribution of altered cellular metabolism to atherosclerosis and how cellular metabolism may be altered by co-morbidities and atherosclerosis risk factors could support the development of novel strategies to lower the risk of CVD. Therefore, we briefly review disease pathogenesis and the principles of cell metabolic pathways, before detailing changes in cellular metabolism in the context of atherosclerosis and comorbidities. In the hypoxic, inflammatory and hyperlipidemic milieu of the atherosclerotic plaque riddled with oxidative stress, metabolism shifts to increase anaerobic glycolysis, the pentose-phosphate pathway and amino acid use. We elaborate on metabolic changes for macrophages, neutrophils, vascular endothelial cells, vascular smooth muscle cells and lymphocytes in the context of atherosclerosis and its co-morbidities hypertension, dyslipidemia, and diabetes. Since causal relationships of specific key genes in a metabolic pathway can be cell type-specific and comorbidity-dependent, the impact of cell-specific metabolic changes must be thoroughly explored in vivo, with a focus on also systemic effects. When cell-specific treatments become feasible, this information will be crucial for determining the best metabolic intervention to improve atherosclerosis and its interplay with co-morbidities. [ABSTRACT FROM AUTHOR]

Endothelial cells (ECs) are vital structural units of the cardiovascular system possessing two principal distinctive properties: heterogeneity and plasticity. Endothelial heterogeneity is defined by differences in tissue-specific endothelial phenotypes and their high predisposition to modification along the length of the vascular bed. This aspect of heterogeneity is closely associated with plasticity, the ability of ECs to adapt to environmental cues through the mobilization of genetic, molecular, and structural alterations. The specific endothelial cytoarchitectonics facilitate a quick structural cell reorganization and, furthermore, easy adaptation to the extrinsic and intrinsic environmental stimuli, known as the epigenetic landscape. ECs, as universally distributed and ubiquitous cells of the human body, play a role that extends far beyond their structural function in the cardiovascular system. They play a crucial role in terms of barrier function, cell-to-cell communication, and a myriad of physiological and pathologic processes. These include development, ontogenesis, disease initiation, and progression, as well as growth, regeneration, and repair. Despite substantial progress in the understanding of endothelial cell biology, the role of ECs in healthy conditions and pathologies remains a fascinating area of exploration. This review aims to summarize knowledge and concepts in endothelial biology. It focuses on the development and functional characteristics of endothelial cells in health and pathological conditions, with a particular emphasis on endothelial phenotypic and functional heterogeneity. [ABSTRACT FROM AUTHOR]

Background: Monocytes and macrophages play an indispensable role in maintaining intestinal homeostasis and modulating mucosal immune responses in inflammatory bowel disease (IBD). Although numerous studies have described macrophage properties in IBD, the underlying mechanisms whereby the monocyte-macrophage lineage modulates intestinal homeostasis during gut inflammation remain elusive. Main body: In this review, we decipher the cellular and molecular mechanisms governing the generation of intestinal mucosal macrophages and fill the knowledge gap in understanding the origin, maturation, classification, and functions of mucosal macrophages in intestinal niches, particularly the phagocytosis and bactericidal effects involved in the elimination of cell debris and pathogens. We delineate macrophage-mediated immunoregulation in the context of producing pro-inflammatory and anti-inflammatory cytokines, chemokines, toxic mediators, and macrophage extracellular traps (METs), and participating in the modulation of epithelial cell proliferation, angiogenesis, and fibrosis in the intestine and its accessory tissues. Moreover, we emphasize that the maturation of intestinal macrophages is arrested at immature stage during IBD, and the deficiency of MCPIP1 involves in the process via ATF3-AP1S2 signature. In addition, we confirmed the origin potential of IL-1B+ macrophages and defined C1QB+ macrophages as mature macrophages. The interaction crosstalk between the intestine and the mesentery has been described in this review, and the expression of mesentery-derived SAA2 is upregulated during IBD, which contributes to immunoregulation of macrophage. Moreover, we also highlight IBD-related susceptibility genes (e.g., RUNX3, IL21R, GTF2I, and LILRB3) associated with the maturation and functions of macrophage, which provide promising therapeutic opportunities for treating human IBD. Conclusion: In summary, this review provides a comprehensive, comprehensive, in-depth and novel description of the characteristics and functions of macrophages in IBD, and highlights the important role of macrophages in the molecular and cellular process during IBD. [ABSTRACT FROM AUTHOR]

Accumulating literature suggests that the farnesoid-X receptor (FXR), a nuclear bile acid receptor best known for its role in bile acid homeostasis, is also a potent context-dependent regulator of inflammation. FXR may thus be relevant to several intestinal disease states including inflammatory bowel disease, necrotizing enterocolitis, and sepsis. In this study, we tested the effects of FXR deletion on acute murine intestinal inflammation. We found that FXR knockout (KO) mice were protected from intestinal injury and barrier dysfunction induced by lipopolysaccharide (LPS) injection, dithizone (DI)/Klebsiella, and cecal ligation/puncture models. In the LPS model, RNA sequencing and qPCR analysis showed that this protection correlated with substantial reduction in LPSinduced proinflammatory gene expression, including lower tissue levels of Il1a, Il1b, and Tnf. Examining functional effects on the epithelium, we found that LPS-induced tight junctional disruption as assessed by internalization of ZO-1 and occludin was ameliorated in FXR KO animals. Taken together, these data suggest a role for FXR in the intestinal barrier during inflammatory injury. NEW & NOTEWORTHY Intestinal barrier failure is a hallmark in gut-origin sepsis. We demonstrate that the intestinal barriers of farnesoid-X receptor (FXR) knockout (KO) animals are protected from inflammatory insult using multiple models of acute intestinal inflammation. This protection is due to decreased inflammatory cytokine production and maintenance of tight junctional architecture seen within the KO animals. This is the first report of FXR deletion being protective to the intestinal barrier. [ABSTRACT FROM AUTHOR]

Treatments of colitis, inflammation of the intestine, rely on induction of immune suppression associated with systemic adverse events, including recurrent infections. This treatment strategy is specifically problematic in the increasing population of patients with cancer with immune checkpoint inhibitor (ICI)-induced colitis, as immune suppression also interferes with the ICI-treatment response. Thus, there is a need for local-acting treatments that reduce inflammation and enhance intestinal healing. Here, we investigated the effect and safety of bacterial delivery of short-lived immunomodulating chemokines to the inflamed intestine in mice with colitis. Colitis was induced by dextran sulfate sodium (DSS) alone or in combination with ICI (anti-PD1 and anti-CTLA-4), and Limosilactobacillus reuteri R2LC (L. reuteri R2LC) genetically modified to express the chemokine CXCL12-1α (R2LC_CXCL12, emilimogene sigulactibac) was given perorally. In addition, the pharmacology and safety of the formulated drug candidate, ILP100-Oral, were evaluated in rabbits. Peroral CXCL12-producing L. reuteri R2LC significantly improved colitis symptoms already after 2 days in mice with overt DSS and ICI-induced colitis, which in benchmarking experiments was demonstrated to be superior to treatments with anti-TNF-α, anti-α4β7, and corticosteroids. The mechanism of action involved chemokine delivery to Peyer’s patches (PPs), confirmed by local CXCR4 signaling, and increased numbers of colonic, regulatory immune cells expressing IL-10 and TGF-β1. No systemic exposure or engraftment could be detected in mice, and product feasibility, pharmacology, and safety were confirmed in rabbits. In conclusion, peroral CXCL12-producing L. reuteri R2LC efficiently ameliorates colitis, enhances mucosal healing, and has a favorable safety profile. NEW & NOTEWORTHY Colitis symptoms are efficiently reduced by peroral administration of probiotic bacteria genetically modified to deliver CXCL12 locally to the inflamed intestine in several mouse models. [ABSTRACT FROM AUTHOR]

Increased production of advanced glycation end products (AGEs) among reducing sugars (glucose, fructose, galactose, or ribose) and amino acids/proteins via non-enzymatic Maillard reaction can be found in lifestyle-related disease (LSRD), metabolic syndrome (MetS), and obesity and immune-related diseases. Increased serum levels of AGEs may induce aging, diabetic complications, cardiovascular diseases (CVD), neurodegenerative diseases (NDD), cancer, and inflamm-aging (inflammation with immunosenescence). The Maillard reaction can also occur among reducing sugars and lipoproteins or DNAs to alter their structure and induce immunogenicity/genotoxicity for carcinogenesis. AGEs, as danger-associated molecular pattern molecules (DAMPs), operate via binding to receptor for AGE (RAGE) or other scavenger receptors on cell surface to activate PI3K-Akt-, P38-MAPK-, ERK1/2-JNK-, and MyD88-induced NF-κB signaling pathways to mediate various pathological effects. Recently, the concept of "inflamm-aging" became more defined, and we have unveiled some interesting findings in relation to it. The purpose of the present review is to dissect the potential molecular basis of inflamm-aging in patients with diabetes and immune-mediated diseases caused by different AGEs. [ABSTRACT FROM AUTHOR]

Ischemic heart disease (IHD) can trigger responses from the innate immune system, provoke aseptic inflammatory processes, and result in the recruitment and accumulation of neutrophils. Excessive recruitment of neutrophils is a potential driver of persistent cardiac inflammation. Once recruited, neutrophils are capable of secreting a plethora of inflammatory and chemotactic agents that intensify the inflammatory cascade. Additionally, neutrophils may obstruct microvasculature within the inflamed region, further augmenting myocardial injury in the context of IHD. Immune-related molecules mediate the recruitment process of neutrophils, such as immune receptors and ligands, immune active molecules, and immunocytes. Non-immune-related molecular pathways represented by pro-resolving lipid mediators are also involved in the regulation of NR. Finally, we discuss novel regulating strategies, including targeted intervention, agents, and phytochemical strategies. This review describes in as much detail as possible the upstream molecular mechanism and external intervention strategies for regulating NR, which represents a promising therapeutic avenue for IHD. [ABSTRACT FROM AUTHOR]

Introduction: As the body's first line of defense against disease and infection, neutrophils must efficiently navigate to sites of inflammation; however, neutrophil dysregulation contributes to the pathogenesis of numerous diseases that leave people susceptible to infections. Many of these diseases are also associated with changes to the protein composition of the extracellular matrix. While it is known that neutrophils and endothelial cells, which play a key role in neutrophil activation, are sensitive to the mechanical and structural properties of the extracellular matrix, our understanding of how protein composition in the matrix affects the neutrophil response to infection is incomplete. Methods: To investigate the effects of extracellular matrix composition on the neutrophil response to infection, we used an infection-on-a-chip microfluidic device that replicates a portion of a blood vessel endothelium surrounded by a model extracellular matrix. Model blood vessels were fabricated by seeding human umbilical vein endothelial cells on 2, 4, or 6 mg/mL type I collagen hydrogels. Primary human neutrophils were loaded into the endothelial lumens and stimulated by adding the bacterial pathogen Pseudomonas aeruginosa to the surrounding matrix. Results: Collagen concentration did not affect the cell density or barrier function of the endothelial lumens. Upon infectious challenge, we found greater neutrophil extravasation into the 4 mg/mL collagen gels compared to the 6 mg/mL collagen gels. We further found that extravasated neutrophils had the highest migration speed and distance in 2mg/mL gels and that these values decreased with increasing collagen concentration. However, these phenomena were not observed in the absence of an endothelial lumen. Lastly, no differences in the percent of extravasated neutrophils producing reactive oxygen species were observed across the various collagen concentrations. Discussion: Our study suggests that neutrophil extravasation and migration in response to an infectious challenge are regulated by collagen concentration in an endothelial cell-dependent manner. The results demonstrate how the mechanical and structural aspects of the tissue microenvironment affect the neutrophil response to infection. Additionally, these findings underscore the importance of developing and using microphysiological systems for studying the regulatory factors that govern the neutrophil response. [ABSTRACT FROM AUTHOR]

Puumala orthohantavirus-caused hemorrhagic fever with renal syndrome (PUUVHFRS) is characterized by strong neutrophil activation. Neutrophils are the most abundant immune cell type in the circulation and are specially equipped to rapidly respond to infections. They are more heterogenous than previously appreciated, with specific neutrophil subsets recently implicated in inflammation and immunosuppression. Furthermore, neutrophils can be divided based on their density to either low-density granulocytes (LDGs) or "normal density" polymorphonuclear cell (PMN) fractions. In the current study we aimed to identify and characterize the different neutrophil subsets in the circulation of PUUV-HFRS patients. PMNs exhibited an activation of antiviral pathways, while circulating LDGs were increased in frequency following acute PUUV-HFRS. Furthermore, cell surface marker expression analysis revealed that PUUV-associated LDGs are primarily immature and most likely reflect an increased neutrophil production from the bone marrow. Interestingly, both the frequency of LDGs and the presence of a "left shift" in blood associated with the extent of thrombocytopenia, one of the hallmarks of severe HFRS, suggesting that maturing neutrophils could play a role in disease pathogenesis. These results imply that elevated circulating LDGs might be a general finding in acute viral infections. However, in contrast to the COVID-19 associated LDGs described previously, the secretome of PUUV LDGs did not show significant immunosuppressive ability, which suggests inherent biological differences in the LDG responses that can be dependent on the causative virus or differing infection kinetics. [ABSTRACT FROM AUTHOR]

Human immunodeficiency virus (HIV) infection remains an important global public health problem. About 40 million people are infected with HIV, and this infection caused about 630,000 deaths in 2022. The hallmark of HIV infection is the depletion of CD4+ T helper lymphocytes (Th cells). There are at least seven different Th subtypes, and not all are the main targets of HIV. Moreover, the effect of the virus in a specific subtype can be completely different from that of the others. Although the most compromised Th subtype in HIV infection is Th17, HIV can induce important dysregulations in other subtypes, such as follicular Th (Tfh) cells and regulatory Th cells (Treg cells or Tregs). Several studies have shown that HIV can induce an increase in the immunosuppressive activity of Tregs without causing a significant reduction in their numbers, at least in the early phase of infection. The increased activity of this Th subtype seems to play an important role in determining the immunodeficiency status of HIV-infected patients, and Tregs may represent a new target for innovative anti-HIV therapies, including the so-called "Kick and Kill" therapeutic method whose goal is the complete elimination of the virus and the healing of HIV infection. In this review, we report the most important findings on the effects of HIV on different CD4+ T cell subtypes, the molecular mechanisms by which the virus impairs the functions of these cells, and the implications for new anti-HIV therapeutic strategies. [ABSTRACT FROM AUTHOR]

Thousands struggle with acute and chronic intestinal injury due to various causes. Epithelial intestinal healing is dependent on phenotypic transitions to a mobile phenotype. Focal adhesion kinase (FAK) is a ubiquitous protein that is essential for cell mobility. This phenotype change is mediated by FAK activation and proves to be a promising target for pharmaceutical intervention. While FAK is crucial for intestinal healing, new evidence connects FAK with innate immunity and the importance it plays in macrophage/monocyte chemotaxis, as well as other intracellular signaling cascades. These cascades play a part in macrophage/monocyte polarization, maturation, and inflammation that is associated with intestinal injury. Colony stimulating factors (CSFs) such as macrophage colony stimulating factor (M-CSF/CSF-1) and granulocyte macrophage colony stimulating factor (GM-CSF/CSF-2) play a critical role in maintaining homeostasis within intestinal mucosa by crosstalk capabilities between macrophages and epithelial cells. The communication between these cells is imperative in orchestrating healing upon injury. Diving deeper into these connections may allow us a greater insight into the role that our immune system plays in healing, as well as a better comprehension of inflammatory diseases of the gut. [ABSTRACT FROM AUTHOR]

Immune rejection is the major limitation for porcine xenograft survival in primate recipients. Proinflammatory cytokines play important roles in immune rejection and have been found to mediate the pathological effects in various clinical and experimental transplantation trials. IL-17 and TNF-α play critical pathological roles in immune disorders, such as psoriasis and rheumatoid arthritis. However, the pathological roles of human IL-17 (hIL-17) and human TNF-α (hTNF-α) in xenotransplantation remain unclear. Here we found that hIL-17 and hTNFa additively or synergistically regulate the expression of 697 genes in porcine aortic endothelial cells (PAECs). Overall, 415 genes were found to be synergistically regulated, while 282 genes were found to be additively regulated. Among these, 315 genes were upregulated and 382 genes were downregulated in PAECs. Furthermore, we found that hIL-17 and hTNF-α additively or synergistically induced the expression of various proinflammatory cytokines and chemokines (e.g., IL1a, IL6, and CXCL8) and decreased the expression of certain antiinflammatory genes (e.g., IL10). Moreover, hIL-17 plus hTNF-α increased the expression of IL1R1 and IL6ST, receptors for IL1 and IL6, respectively, and decreased anti-inflammatory gene receptor expression (IL10R). hIL-17 and hTNF-α synergistically or additively induced CXCL8 and CCL2 expression and consequently promoted primary human neutrophil and human leukemia monocytic cell migration, respectively. In addition, hIL-17 and hTNF-α induced pro-coagulation gene (SERPINB2 and F3) expression and decreased anticoagulation gene (TFPI, THBS1, and THBD) expression. Additionally, hIL-17 and hTNF-α synergistically decreased occludin expression and consequently promoted human antibodymediated complement-dependent cytotoxicity. Interestingly, hTNF-α increased swine leukocyte antigen (SLA) class I expression; however, hIL-17 decreased TNF-α-mediated SLA-I upregulation. We concluded that hIL-17 and hTNF-α likely promote the inflammatory response, coagulation cascade, and xenoantibody-mediated cell injury. Thus, blockade of hIL-17 and hTNF-α together might be beneficial for xenograft survival in recipients. [ABSTRACT FROM AUTHOR]

Atherosclerotic cardiovascular disease poses a significant global health issue, with dyslipidemia standing out as a major risk factor. In recent decades, lipid-lowering therapies have evolved significantly, with statins emerging as the cornerstone treatment. These interventions play a crucial role in both primary and secondary prevention by effectively reducing cardiovascular risk through lipid profile enhancements. Beyond their primary lipid-lowering effects, extensive research indicates that these therapies exhibit pleiotropic actions, offering additional health benefits. These include anti-inflammatory properties, improvements in vascular health and glucose metabolism, and potential implications in cancer management. While statins and ezetimibe have been extensively studied, newer lipid-lowering agents also demonstrate similar pleiotropic effects, even in the absence of direct cardiovascular benefits. This narrative review explores the diverse pleiotropic properties of lipid-modifying therapies, emphasizing their non-lipid effects that contribute to reducing cardiovascular burden and exploring emerging benefits for non-cardiovascular conditions. Mechanistic insights into these actions are discussed alongside their potential therapeutic implications [ABSTRACT FROM AUTHOR]

The immune cell system is a critical component of host defense. Recruitment of immune cells to sites of infection, immune reaction, or injury is complex and involves coordinated adhesive interactions between the leukocyte and the endothelial cell monolayer that lines blood vessels. This article reviews basic mechanisms in the recruitment of leukocytes to tissues and then selectively reviews new concepts that are emerging based on advances in live cell imaging microscopy and mouse strains. These emerging concepts are altering the conventional paradigms of inflammatory leukocyte recruitment established in the early 1990s. Indeed, recent publications have identified previously unrecognized contributions from pericytes and interstitial leukocytes and their secreted products that guide leukocytes to their targets. Investigators have also begun to design organs on a chip. Recent reports indicate that this avenue of research holds much promise.

CD47 plays an important but incompletely understood role in the innate and adaptive immune responses. CD47, also called integrin-associated protein, has been demonstrated to associate in cis with β1 and β3 integrins. Here we test the hypothesis that CD47 regulates adhesive functions of T-cell α4β1 (VLA-4) and αLβ2 (LFA-1) in in vivo and in vitro models of inflammation. Intravital microscopy studies reveal that CD47(-/-) Th1 cells exhibit reduced interactions with wild-type (WT) inflamed cremaster muscle microvessels. Similarly, murine CD47(-/-) Th1 cells, as compared with WT, showed defects in adhesion and transmigration across tumor necrosis factor-α (TNF-α)-activated murine endothelium and in adhesion to immobilized intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion protein 1 (VCAM-1) under flow conditions. Human Jurkat T-cells lacking CD47 also showed reduced adhesion to TNF-α-activated endothelium and ICAM-1 and VCAM-1. In cis interactions between Jurkat T-cell β2 integrins and CD47 were detected by fluorescence lifetime imaging microscopy. Unexpectedly, Jurkat CD47 null cells exhibited a striking defect in β1 and β2 integrin activation in response to Mn(2+) or Mg(2+)/ethylene glycol tetraacetic acid treatment. Our results demonstrate that CD47 associates with β2 integrins and is necessary to induce high-affinity conformations of LFA-1 and VLA-4 that recognize their endothelial cell ligands and support leukocyte adhesion and transendothelial migration.

The heptapeptide angiotensin-(1-7) is a biologically active metabolite of angiotensin II, the predominant peptide of the renin-angiotensin system. Recently, we have shown that the receptor Mas is associated with angiotensin-(1-7)-induced signalling and mediates, at least in part, the vasodilatory properties of angiotensin-(1-7). However, it remained controversial whether an additional receptor could account for angiotensin-(1-7)-induced vasorelaxation. Here, we used two different angiotensin-(1-7) antagonists, A779 and d-Pro-angiotensin-(1-7), to address this question and also to study their influence on the vasodilatation induced by bradykinin. Isolated mesenteric microvessels from both wild-type and Mas-deficient C57Bl/6 mice were precontracted with noradrenaline, and vascular reactivity to angiotensin-(1-7) and bradykinin was subsequently studied using a small-vessel myograph. Furthermore, mechanisms for Mas effects were investigated in primary human umbilical vein endothelial cells. Both angiotensin-(1-7) and bradykinin triggered a concentration-dependent vasodilatation in wild-type microvessels, which was absent in the presence of a nitric oxide synthase inhibitor. In these vessels, the pre-incubation with the Mas antagonists A779 or d-Pro-angiotensin-(1-7) totally abolished the vasodilatory capacity of both angiotensin-(1-7) and bradykinin, which was nitric oxide mediated. Accordingly, Mas-deficient microvessels lacked the capacity to relax in response to either angiotensin-(1-7) or bradykinin. Pre-incubation of human umbilical vein endothelial cells with A779 prevented bradykinin-mediated NO generation and NO synthase phosphorylation at serine 1177. The angiotensin-(1-7) antagonists A779 and d-Pro-angiotensin-(1-7) equally block Mas, which completely controls the angiotensin-(1-7)-induced vasodilatation in mesenteric microvessels. Importantly, Mas also appears to be a critical player in NO-mediated vasodilatation induced by renin-angiotensin system-independent agonists by altering phosphorylation of NO synthase.

Aims: Activation of vascular endothelial cells (ECs) contributes importantly to inflammation and atherogenesis. We previously reported that apolipoprotein CIII (apoCIII), found abundantly on circulating triglyceride-rich lipoproteins, enhances adhesion of human monocytes to ECs in vitro. Statins may exert lipid-independent anti-inflammatory effects. The present study examined whether statins suppress apoCIII-induced EC activation in vitro and in vivo., Methods and Results: Physiologically relevant concentrations of purified human apoCIII enhanced attachment of the monocyte-like cell line THP-1 to human saphenous vein ECs (HSVECs) or human coronary artery ECs (HCAECs) under both static and laminar shear stress conditions. This process mainly depends on vascular cell adhesion molecule-1 (VCAM-1), as a blocking VCAM-1 antibody abolished apoCIII-induced monocyte adhesion. ApoCIII significantly increased VCAM-1 expression in HSVECs and HCAECs. Pre-treatment with statins suppressed apoCIII-induced VCAM-1 expression and monocyte adhesion, with two lipophilic statins (pitavastatin and atorvastatin) exhibiting inhibitory effects at lower concentration than those of hydrophilic pravastatin. Nuclear factor κB (NF-κB) mediated apoCIII-induced VCAM-1 expression, as demonstrated via loss-of-function experiments, and pitavastatin treatment suppressed NF-κB activation. Furthermore, in the aorta of hypercholesterolaemic Ldlr(-/-) mice, pitavastatin administration in vivo suppressed VCAM-1 mRNA and protein, induced by apoCIII bolus injection. Similarly, in a subcutaneous dorsal air pouch mouse model of leucocyte recruitment, apoCIII injection induced F4/80+ monocyte and macrophage accumulation, whereas pitavastatin administration reduced this effect., Conclusions: These findings further establish the direct role of apoCIII in atherogenesis and suggest that anti-inflammatory effects of statins could improve vascular disease in the population with elevated plasma apoCIII.

Background & Aims: Signaling lymphocyte activation molecule (Slamf)1 is a co-stimulatory receptor on T cells and regulates cytokine production by macrophages and dendritic cells. Slamf1 regulates microbicidal mechanisms in macrophages, therefore we investigated whether the receptor affects development of colitis in mice., Methods: We transferred CD45RB(hi) CD4(+) T cells into Rag(-/-) or Slamf1(-/-)Rag(-/-) mice to induce colitis. We also induced colitis by injecting mice with an antibody that activates CD40. We determined the severity of enterocolitis based on disease activity index, histology scores, and levels of cytokine production, and assessed the effects of antibodies against Slamf1 on colitis induction. We quantified migration of monocytes and macrophage to inflamed tissues upon induction of colitis or thioglycollate-induced peritonitis and in response to tumor necrosis factor-α in an air-pouch model of leukocyte migration., Results: Colitis was reduced in Slamf1(-/-)Rag(-/-) mice, compared with Rag(-/-) mice, after transfer of CD45RB(hi) CD4(+) T cells or administration of the CD40 agonist. The numbers of monocytes and macrophages were reduced in inflamed tissues of Slamf1(-/-)Rag(-/-) mice, compared with Rag(-/-) mice, after induction of colitis and other inflammatory disorders. An antibody that inhibited Slamf1 reduced the level of enterocolitis in Rag(-/-) mice., Conclusions: Slamf1 contributes to the development of colitis in mice. It appears to indirectly regulate the appearance of monocytes and macrophages in inflamed intestinal tissues. Antibodies that inhibit Slamf1 reduce colitis in mice, so human SLAMF1 might be a therapeutic target for inflammatory bowel disease., (Copyright © 2012 AGA Institute. Published by Elsevier Inc. All rights reserved.)

At sites of inflammation, endothelial adhesion molecules bind leukocytes and transmit signals required for transendothelial migration (TEM). We previously reported that adhesive interactions between endothelial cell CD47 and leukocyte signal regulatory protein γ (SIRPγ) regulate human T cell TEM. The role of endothelial CD47 in T cell TEM in vivo, however, has not been explored. In this study, CD47⁻/⁻ mice showed reduced recruitment of blood T cells as well as neutrophils and monocytes in a dermal air pouch model of TNF-α-induced inflammation. Reconstitution of CD47⁻/⁻ mice with wild-type bone marrow cells did not restore leukocyte recruitment to the air pouch, indicating a role for endothelial CD47. The defect in leukocyte TEM in the CD47⁻/⁻ endothelium was corroborated by intravital microscopy of inflamed cremaster muscle microcirculation in bone marrow chimera mice. In an in vitro human system, CD47 on both HUVEC and T cells was required for TEM. Although previous studies showed CD47-dependent signaling required G(αi)-coupled pathways, this was not the case for endothelial CD47 because pertussis toxin, which inactivates G(αi), had no inhibitory effect, whereas G(αi) was required by the T cell for TEM. We next investigated the endothelial CD47-dependent signaling events that accompany leukocyte TEM. Ab-induced cross-linking of CD47 revealed robust actin cytoskeleton reorganization and Src- and Pyk-2-kinase dependent tyrosine phosphorylation of the vascular endothelial-cadherin cytoplasmic tail. This signaling was pertussis toxin insensitive, suggesting that endothelial CD47 signaling is independent of G(αi). These findings suggest that engagement of endothelial CD47 by its ligands triggers outside-in signals in endothelium that facilitate leukocyte TEM.

IL-17A (IL-17) is the signature cytokine produced by Th17 cells and has been implicated in host defense against infection and the pathophysiology of autoimmunity and cardiovascular disease. Little is known, however, about the influence of IL-17 on endothelial activation and leukocyte influx to sites of inflammation. We hypothesized that IL-17 would induce a distinct pattern of endothelial activation and leukocyte recruitment when compared with the Th1 cytokine IFN-γ. We found that IL-17 alone had minimal activating effects on cultured endothelium, whereas the combination of TNF-α and IL-17 produced a synergistic increase in the expression of both P-selectin and E-selectin. Using intravital microscopy of the mouse cremaster muscle, we found that TNF-α and IL-17 also led to a synergistic increase in E-selectin-dependent leukocyte rolling on microvascular endothelium in vivo. In addition, TNF-α and IL-17 enhanced endothelial expression of the neutrophilic chemokines CXCL1, CXCL2, and CXCL5 and led to a functional increase in leukocyte transmigration in vivo and CXCR2-dependent neutrophil but not T cell transmigration in a parallel-plate flow chamber system. By contrast, endothelial activation with TNF-α and IFN-γ preferentially induced the expression of the integrin ligands ICAM-1 and VCAM-1, as well as the T cell chemokines CXCL9, CXCL10, and CCL5. These effects were further associated with a functional increase in T cell but not neutrophil transmigration under laminar shear flow. Overall, these data show that IL-17 and TNF-α act in a synergistic manner to induce a distinct pattern of endothelial activation that sustains and enhances neutrophil influx to sites of inflammation.

Neutrophils are the all-terrain vehicle of the innate immune system because of their ability to gain entry into tissues and organs, and thus, play an essential role in host defense. Exactly how this marvel of nature works is still incompletely understood. In the past 2-3 years, new players and processes have been identified in the endothelial-leukocyte adhesion cascade. Novel signaling pathways have been discovered in both the endothelium and the neutrophils that regulate various steps in the recruitment process. This review focuses on these emerging pathways and the mechanisms that regulate neutrophil recruitment across endothelium., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: Apoptosis and inflammatory/oxidative stress have been associated with hyperglycemia in human peritoneal mesothelial cells (HPMCs) and other cell types. We and others have highlighted the role of early products of non-enzymatic protein glycation in inducing proinflammatory conditions and increasing apoptotic rates in HPMCs. Loss of HPMCs seems to be a hallmark of complications associated with peritoneal membrane dysfunction. The aim of this work is to elucidate the mechanisms by which Amadori adducts may act upon HPMC apoptosis., Methods: HPMCs isolated from different patients were exposed to different Amadori adducts, i.e. highly glycated hemoglobin (10 nM) and glycated bovine serum albumin (250 μg/ml), to study cell death and several proapoptotic markers by different experimental approaches., Results: Amadori adducts, but not their respective controls, impaired cell proliferation and cell viability by means of apoptosis in a time-dependent manner. They regulated the intrinsic mitochondrial cell death signaling pathway and modulated activation of caspases, Bax, iNOS, p53, NF-κB, and mitogen-activated protein kinases (p38 and JNK) through different reactive oxygen and nitrosative species., Conclusions: Our data strongly support the idea that long-term hyperglycemia could act as an inducer of apoptosis in HPMCs through Amadori adducts, involving different oxidative and nitrosative reactive species., (Copyright © 2011 S. Karger AG, Basel.)

Background: Hyperglycemia is acknowledged as an independent risk factor for developing diabetes-associated atherosclerosis. At present, most therapeutic approaches are targeted at a tight glycemic control in diabetic patients, although this fails to prevent macrovascular complications of the disease. Indeed, it remains highly controversial whether or not the mere elevation of extracellular D-glucose can directly promote vascular inflammation, which favors early pro-atherosclerotic events., Methods and Findings: In the present work, increasing extracellular D-glucose from 5.5 to 22 mmol/L was neither sufficient to induce intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expression, analyzed by flow cytometry, nor to promote leukocyte adhesion to human umbilical vein endothelial cells (HUVEC) in vitro, measured by flow chamber assays. Interestingly, the elevation of D-glucose levels potentiated ICAM-1 and VCAM-1 expression and leukocyte adhesion induced by a pro-inflammatory stimulus, such as interleukin (IL)-1beta (5 ng/mL). In HUVEC, high D-glucose augmented the activation of extracellular signal-regulated kinase 1/2 (ERK 1/2) and nuclear transcription factor-kappaB (NF-kappaB) elicited by IL-1beta, measured by Western blot and electromobility shift assay (EMSA), respectively, but had no effect by itself. Both ERK 1/2 and NF-kappaB were necessary for VCAM-1 expression, but not for ICAM-1 expression. In vivo, leukocyte trafficking was evaluated in the rat mesenteric microcirculation by intravital microscopy. In accordance with the in vitro data, the acute intraperitoneal injection of D-glucose increased leukocyte rolling flux, adhesion and migration, but only when IL-1beta was co-administered., Conclusions: These results indicate that the elevation of extracellular D-glucose levels is not sufficient to promote vascular inflammation, and they highlight the pivotal role of a pro-inflammatory environment in diabetes, as a critical factor conditioning the early pro-atherosclerotic actions of hyperglycemia.

Objectives: The present study investigated whether high concentrations of D-glucose can trigger pro-inflammatory mechanisms in human aortic smooth muscle cells., Methods: The expression and/or the activity of inducible nitric oxide synthase (iNOS), the extracellular signal-regulated kinase (ERK) 1/2 and nuclear factor (NF)-kappaB were studied in cultured human aortic smooth muscle cells (HASMC) in response to increasing concentrations of D-glucose and/or the inflammatory cytokine interleukin (IL)-1beta., Results: Increasing D-glucose in the medium from 5.5 to 22 mmol/l had no effect on any of these parameters. However, the high concentration of D-glucose did increase iNOS expression in response to low concentrations of IL-1beta (2.5 and 5 ng/ml), as well as the IL-1beta-induced activation of both ERK 1/2 and NF-kappaB. D-glucose also enhanced, concentration-dependently, the expression and activity of iNOS induced by co-incubation with IL-1beta (10 ng/ml). Pretreatment with IL-1beta sensitized the cells to the subsequent effects of high D-glucose., Conclusions: The results indicate that high concentrations of D-glucose exacerbate the pro-inflammatory effects of IL-1beta. We suggest that the observed association between inflammation and diabetes is the result of elevated D-glucose enhancing a pre-existing inflammatory condition, rather than a direct effect of D-glucose on the production of inflammatory mediators.

Background: Endothelial dysfunction is an initial step in the pathogenesis of cardiovascular diseases. Since we previously identified the G protein-coupled receptor Mas as a receptor for angiotensin (Ang)-(1-7), a heptapeptide with endothelium-dependent vasorelaxant properties, we investigated whether alterations on the Ang-(1-7)/Mas axis alter endothelial function., Results: Ang-(1-7)-mediated relaxation of murine wild-type mesenteric arteries was equally impaired in both wild-type arteries pretreated with the Ang-(1-7) receptor blocker, A779, and arteries isolated from Mas-deficient mice. Importantly, the response to the endothelium-dependent vasorelaxant, bradykinin (BK), and acetylcholine (ACh) effects were comparably inhibited, while endothelium-independent vessel relaxation by sodium nitroprusside was unaltered in these vessels. Hypothesizing endothelial dysfunction, we proved the in-vivo relevance of the ex-vivo findings investigating mesenteric properties after 1 week of minipump infusion of A779 in wild-type mice. Both BK- and ACh-induced relaxation were significantly impaired in wild-type vessels of pretreated animals. A779-induced impairment of endothelial function was confirmed in vitro, since BK-mediated nitric oxide (NO) release was increased by Ang-(1-7) and blunted by A779 pretreatment in primary human endothelial cell cultures., Conclusions: Our data highlight a pivotal role for the receptor Mas in preserving normal vascular relaxation. Consequently, Mas agonists arise as a promising tool in the treatment of cardiovascular diseases characterized by endothelial dysfunction.