20 ELUCIDATING THE ROLE OF FUSOBACTERIUM NUCLEATUM IN INTESTINAL INFLAMMATION

2020 ◽  
Vol 26 (Supplement_1) ◽  
pp. S29-S29
Author(s):  
Melinda Engevik ◽  
Heather Danhof ◽  
Robert Britton ◽  
James Versalovic
Keyword(s):  
Immune Cell ◽  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Bacterial Species ◽  
Fusobacterium Nucleatum ◽  
Outer Membrane Vesicles ◽  
Patient Treatment ◽  
Mucus Layer

Abstract Background While the direct cause of inflammatory bowel disease (IBD) is unknown, the gut microbiota is speculated to play a key role. The complexity of the microbiome has made it difficult to pinpoint whether bacterial species are specifically associated with IBD exacerbations; although select microbes have emerged as compelling candidates. Several groups have identified increased abundance in Fusobacterium in IBD patients. Fusobacterium nucleatum drives inflammation in the oral cavity, but few studies have examined the potential for F. nucleatum to promote intestinal inflammation. We hypothesize that F. nucleatum secretes Outer Membrane Vesicles (OMVs) that activate epithelial Toll-like receptor 4 (TLR4) to drive inflammation. Methods & Results Given the prevalence of F. nucleatum in IBD specimens, we sought to determine if this pathobiont could promote pro-inflammatory responses in human epithelial cultures. Using fluorescently tagged F. nucleatum, we demonstrate that F. nucleatum subspecies polymorphum adheres to the mucus layer of human colonic HT29-MTX cells. Application of F. nucleatum metabolites to HT29-MTX cells resulted in upregulation of pro-inflammatory cytokines IL-8 and TNF by qPCR and ELISA. Purified OMVs from F. nucleatum alone were able to stimulate IL-8 and TNF production. This demonstrates the robust response of colonic epithelial cells to F. nucleatum. Additionally, we used human jejunum and colon enteroid monolayers treated with F. nucleatum metabolites in an anti-oxidant free enteroid media and found that F. nucleatum secreted products promoted TNF secretion by ELISA. Using the Luminex Magpix Platform we further queried the enteroid system to assess which pathways were activated. Enteroid monolayers treated with F. nucleatum metabolites exhibited increased phosphorylated ERK and CREB, downstream effectors of TLRs. We next sought to address whether F. nucleatum alone could elicit pro-inflammatory responses in a mouse model. Mice harboring a human microbiota, or humanized mice, were treated for 5 days with a cocktail of antibiotics and treated with F. nucleatum (108 CFU) by oral gavage; a regimen designed to mimic IBD patient treatment. Compared to control mice that received antibiotics and PBS vehicle control, mice treated with F. nucleatum exhibited disruption of the colonic architecture, with increased immune infiltrate and depleted mucus layer, resulting in a closer proximity of luminal contents to the epithelium. Analysis of mucosal gene expression revealed increased levels of epithelial TNF and KC (mouse homolog of IL-8), in addition to immune cell derived IL-6 in F. nucleatum-treated mice compared for controls. Conclusions These data provide evidence that F. nucleatum is capable of driving a pro-inflammatory signaling cascade in vitro and in vivo and F. nucleatum may represent a specific target for drug therapy.

scholarly journals Fusobacterium nucleatum Secretes Outer Membrane Vesicles and Promotes Intestinal Inflammation

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Melinda A. Engevik ◽  
Heather A. Danhof ◽  
Wenly Ruan ◽  
Amy C. Engevik ◽  
Alexandra L. Chang-Graham ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Proinflammatory Cytokines ◽  
Immune Cell ◽  
Intestinal Inflammation ◽  
Membrane Vesicles ◽  
Fusobacterium Nucleatum ◽  
Outer Membrane Vesicles ◽  
Intestinal Microbiome ◽  
Content Type

ABSTRACT Multiple studies have implicated microbes in the development of inflammation, but the mechanisms remain unknown. Bacteria in the genus Fusobacterium have been identified in the intestinal mucosa of patients with digestive diseases; thus, we hypothesized that Fusobacterium nucleatum promotes intestinal inflammation. The addition of >50 kDa F. nucleatum conditioned media, which contain outer membrane vesicles (OMVs), to colonic epithelial cells stimulated secretion of the proinflammatory cytokines interleukin-8 (IL-8) and tumor necrosis factor (TNF). In addition, purified F. nucleatum OMVs, but not compounds <50 kDa, stimulated IL-8 and TNF production; which was decreased by pharmacological inhibition of Toll-like receptor 4 (TLR4). These effects were linked to downstream effectors p-ERK, p-CREB, and NF-κB. F. nucleatum >50-kDa compounds also stimulated TNF secretion, p-ERK, p-CREB, and NF-κB activation in human colonoid monolayers. In mice harboring a human microbiota, pretreatment with antibiotics and a single oral gavage of F. nucleatum resulted in inflammation. Compared to mice receiving vehicle control, mice treated with F. nucleatum showed disruption of the colonic architecture, with increased immune cell infiltration and depleted mucus layers. Analysis of mucosal gene expression revealed increased levels of proinflammatory cytokines (KC, TNF, IL-6, IFN-γ, and MCP-1) at day 3 and day 5 in F. nucleatum-treated mice compared to controls. These proinflammatory effects were absent in mice who received F. nucleatum without pretreatment with antibiotics, suggesting that an intact microbiome is protective against F. nucleatum-mediated immune responses. These data provide evidence that F. nucleatum promotes proinflammatory signaling cascades in the context of a depleted intestinal microbiome. IMPORTANCE Several studies have identified an increased abundance of Fusobacterium in the intestinal tracts of patients with colon cancer, liver cirrhosis, primary sclerosing cholangitis, gastroesophageal reflux disease, HIV infection, and alcoholism. However, the direct mechanism(s) of action of Fusobacterium on pathophysiological within the gastrointestinal tract is unclear. These studies have identified that F. nucleatum subsp. polymorphum releases outer membrane vesicles which activate TLR4 and NF-κB to stimulate proinflammatory signals in vitro. Using mice harboring a human microbiome, we demonstrate that F. nucleatum can promote inflammation, an effect which required antibiotic-mediated alterations in the gut microbiome. Collectively, these results suggest a mechanism by which F. nucleatum may contribute to intestinal inflammation.

scholarly journals Viperin is anti-viral in vitro but is dispensable for restricting dengue virus replication or induction of innate and inflammatory responses in vivo

2021 ◽  
Vol 102 (10) ◽  
Author(s):  
Wisam-Hamzah Al Shujairi ◽  
Luke P. Kris ◽  
Kylie van der Hoek ◽  
Evangeline Cowell ◽  
Gustavo Bracho-Granado ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Immune Cell ◽  
Inflammatory Responses ◽  
Denv Infection ◽  
Brain Morphology ◽  
Moderate Increase ◽  
Tnf Α ◽  
Significant Difference

Viperin has antiviral function against many viruses, including dengue virus (DENV), when studied in cells in culture. Here, the antiviral actions of viperin were defined both in vitro and in a mouse in vivo model of DENV infection. Murine embryonic fibroblasts (MEFs) derived from mice lacking viperin (vip−/−) showed enhanced DENV infection, accompanied by increased IFN-β and induction of ISGs; IFIT1 and CXCL-10 but not IRF7, when compared to wild-type (WT) MEFs. In contrast, subcutaneous challenge of immunocompetent WT and vip−/− mice with DENV did not result in enhanced infection. Intracranial infection with DENV resulted in body weight loss and neurological disease with a moderate increase in mortality in vip−/− compared with WT mice, although this was not accompanied by altered brain morphology, immune cell infiltration or DENV RNA level in the brain. Similarly, DENV induction of IFN-β, IFIT1, CXCL-10, IRF7 and TNF-α was not significantly different in WT and vip−/− mouse brain, although there was a modest but significant increase in DENV induction of IL-6 and IfI27la in the absence of viperin. NanoString nCounter analysis confirmed no significant difference in induction of a panel of inflammatory genes in WT compared to vip−/− DENV-infected mouse brains. Further, polyI:C stimulation of bone marrow-derived macrophages (BMDMs) induced TNF-α, IFN-β, IL-6 and Nos-2, but responses were not different in BMDMs generated from WT or vip−/− mice. Thus, while there is significant evidence of anti-DENV actions of viperin in some cell types in vitro, for DENV infection in vivo a lack of viperin does not affect systemic or brain susceptibility to DENV or induction of innate and inflammatory responses.

scholarly journals Resolvins as Regulators of the Immune System

2010 ◽  
Vol 10 ◽  
pp. 818-831 ◽  
Author(s):  
Hiroyuki Seki ◽  
Takaharu Sasaki ◽  
Tomomi Ueda ◽  
Makoto Arita
Keyword(s):  
Immune System ◽  
Acute Inflammation ◽  
Immune Cell ◽  
Inflammatory Responses ◽  
Bioactive Metabolites ◽  
Cell Functions ◽  
First Response ◽  
Beneficial Impact

Inflammation is the first response of the immune system to infection or injury, but excessive or inappropriate inflammatory responses contribute to a range of acute and chronic human diseases. Clinical assessment of dietary supplementation of ω-3 polyunsaturated fatty acids (i.e., eicosapentaenoic acid [EPA] and docosahexaenoic acid [DHA]) indicate that they have beneficial impact on these diseases, although the mechanisms are poorly understood at the molecular level. In this decade, it has been revealed that EPA and DHA are enzymatically converted to bioactive metabolites in the course of acute inflammation and resolution. These metabolites were shown to regulate immune cell functions and to display potent anti-inflammatory actions bothin vitroandin vivo. Because of their ability to resolve an acute inflammatory response, they are referred to as proresolving mediators, or resolvins. In this review, we provide an overview of the formation and actions of these lipid mediators.

scholarly journals Mucosa-associated microbiota drives pathogenic functions in IBD-derived intestinal iNKT cells

2019 ◽  
Vol 2 (1) ◽  
pp. e201800229 ◽  
Author(s):  
Claudia Burrello ◽  
Gabriella Pellegrino ◽  
Maria Rita Giuffrè ◽  
Giulia Lovati ◽  
Ilaria Magagna ◽  
...  
Keyword(s):  
Lamina Propria ◽  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Ex Vivo ◽  
Lymphoid Tissues ◽  
Inkt Cells ◽  
Inflammatory Processes ◽  
Inflammatory Bowel

Inflammatory bowel disease (IBD) pathogenesis has been linked to the aberrant activation of the Gut-associated lymphoid tissues against components of the intestinal microbiota. Although the contribution of CD4+ T helper cells to inflammatory processes is being increasingly acknowledged, the functional engagement of human invariant natural killer T (iNKT) cells is still poorly defined. Here, we evaluated the functional characteristics of intestinal iNKT cells during IBD pathogenesis and to exploit the role of mucosa-associated microbiota recognition in triggering iNKT cells’ pro-inflammatory responses in vivo. Lamina propria iNKT cells, isolated from surgical specimens of active ulcerative colitis and Crohn’s disease patients and non-IBD donors, were phenotypically and functionally analyzed ex vivo, and stable cell lines and clones were generated for in vitro functional assays. iNKT cells expressing a pro-inflammatory cytokine profile were enriched in the lamina propria of IBD patients, and their exposure to the mucosa-associated microbiota drives pro-inflammatory activation, inducing direct pathogenic activities against the epithelial barrier integrity. These observations suggest that iNKT cell pro-inflammatory functions may contribute to the fuelling of intestinal inflammation in IBD patients.

scholarly journals Barrier Protection and Recovery Effects of Gut Commensal Bacteria on Differentiated Intestinal Epithelial Cells In Vitro

Nutrients ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2251
Author(s):  
Nooshin Mohebali ◽  
Katharina Ekat ◽  
Bernd Kreikemeyer ◽  
Anne Breitrück
Keyword(s):  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Therapeutic Potential ◽  
Bacterial Species ◽  
Cell Monolayer ◽  
Commensal Bacteria ◽  
Epithelial Barrier ◽  
Inflammatory Stimulus ◽  
Monocyte Chemoattractant Protein 1

Alterations in the gut microbiota composition play a crucial role in the pathogenesis of inflammatory bowel disease (IBD) as specific commensal bacterial species are underrepresented in the microbiota of IBD patients. In this study, we examined the therapeutic potential of three commensal bacterial species, Faecalibacterium prausnitzii (F. prausnitzii), Roseburia intestinalis (R. intestinalis) and Bacteroides faecis (B. faecis) in an in vitro model of intestinal inflammation, by using differentiated Caco-2 and HT29-MTX cells, stimulated with a pro-inflammatory cocktail consisting of interleukin-1β (IL-1β), tumor necrosis factor-α (TNFα), interferon-γ (IFNγ), and lipopolysaccharide (LPS). Results obtained in this work demonstrated that all three bacterial species are able to recover the impairment of the epithelial barrier function induced by the inflammatory stimulus, as determined by an amelioration of the transepithelial electrical resistance (TEER) and the paracellular permeability of the cell monolayer. Moreover, inflammatory stimulus increased claudin-2 expression and decreased occludin expression were improved in the cells treated with commensal bacteria. Furthermore, the commensals were able to counteract the increased release of interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) induced by the inflammatory stimulus. These findings indicated that F. prausnitzii, R. intestinalis and B. faecis improve the epithelial barrier integrity and limit inflammatory responses.

scholarly journals Anti-Inflammatory and Immunosuppressive Effects of the A2AAdenosine Receptor

2011 ◽  
Vol 11 ◽  
pp. 320-339 ◽  
Author(s):  
Gillian R. Milne ◽  
Timothy M. Palmer
Keyword(s):  
Molecular Mechanisms ◽  
Immune Cell ◽  
Inflammatory Responses ◽  
Current Evidence ◽  
Receptor Subtypes ◽  
Protective Effects ◽  
Stress Injury ◽  
Anti Inflammatory

The production of adenosine represents a critical endogenous mechanism for regulating immune and inflammatory responses during conditions of stress, injury, or infection. Adenosine exerts predominantly protective effects through activation of four 7-transmembrane receptor subtypes termed A1, A2A, A2B, and A3, of which the A2Aadenosine receptor (A2AAR) is recognised as a major mediator of anti-inflammatory responses. The A2AAR is widely expressed on cells of the immune system and numerousin vitrostudies have identified its role in suppressing key stages of the inflammatory process, including leukocyte recruitment, phagocytosis, cytokine production, and immune cell proliferation. The majority of actions produced by A2AAR activation appear to be mediated by cAMP, but downstream events have not yet been well characterised. In this article, we review the current evidence for the anti-inflammatory effects of the A2AAR in different cell types and discuss possible molecular mechanisms mediating these effects, including the potential for generalised suppression of inflammatory gene expression through inhibition of the NF-κB and JAK/STAT proinflammatory signalling pathways. We also evaluate findings fromin vivostudies investigating the role of the A2AAR in different tissues in animal models of inflammatory disease and briefly discuss the potential for development of selective A2AAR agonists for use in the clinic to treat specific inflammatory conditions.

scholarly journals UniPR1331: Small Eph/Ephrin Antagonist Beneficial in Intestinal Inflammation by Interfering with Type-B Signaling

2021 ◽  
Vol 14 (6) ◽  
pp. 502
Author(s):  
Carmine Giorgio ◽  
Marika Allodi ◽  
Simone Palese ◽  
Andrea Grandi ◽  
Massimiliano Tognolini ◽  
...  
Keyword(s):  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Ischemia Reperfusion ◽  
Drug Candidate ◽  
Eph Receptors ◽  
Beneficial Effects ◽  
Bidirectional Signaling ◽  
Novel Drug

Eph receptors, comprising A and B classes, interact with cell-bound ephrins generating bidirectional signaling. Although mainly related to carcinogenesis and organogenesis, the role of Eph/ephrin system in inflammation is growingly acknowledged. Recently, we showed that EphA/ephrin-A proteins can modulate the acute inflammatory responses induced by mesenteric ischemia/reperfusion, while beneficial effects were granted by EphB4, acting as EphB/ephrin-B antagonist, in a murine model of Crohn’s disease (CD). Accordingly, we now aim to evaluate the effects of UniPR1331, a pan-Eph/ephrin antagonist, in TNBS-induced colitis and to ascertain whether UniPR1331 effects can be attributed to A- or B-type signaling interference. The potential anti-inflammatory action of UniPR1331 was compared to those of the recombinant proteins EphA2, a purported EphA/ephrin-A antagonist, and of ephrin-A1-Fc and EphA2-Fc, supposedly activating forward and reverse EphA/ephrin-A signaling, in murine TNBS-induced colitis and in stimulated cultured mononuclear splenocytes. UniPR1331 antagonized the inflammatory responses both in vivo, mimicking EphB4 protection, and in vitro; EphA/ephrin-A proteins were inactive or only weakly effective. Our findings represent a further proof-of-concept that blockade of EphB/ephrin-B signaling is a promising pharmacological strategy for CD management and highlight UniPR1331 as a novel drug candidate, seemingly working through the modulation of immune responses.

scholarly journals Cholesterol-modifying drugs in COVID-19

2020 ◽  
Vol 1 (1) ◽  
Author(s):  
Nathalie M Schmidt ◽  
Peter A C Wing ◽  
Jane A McKeating ◽  
Mala K Maini
Keyword(s):  
Immune Cell ◽  
Inflammatory Responses ◽  
The Elderly ◽  
Virus Infectivity ◽  
In Vivo Models ◽  
Virus Particles ◽  
Enveloped Virus ◽  
Poor Outcomes

Abstract Infection with severe acute respiratory syndrom coronavirus 2 (SARS-CoV-2) is more likely to lead to poor outcomes in the elderly and those with cardiovascular disease, obesity or metabolic syndrome. Here, we consider mechanisms by which dyslipidaemia and the use of cholesterol-modifying drugs could influence the virus–host relationship. Cholesterol is essential for the assembly, replication and infectivity of enveloped virus particles; we highlight several cholesterol-modifying drugs with the potential to alter the SARS-CoV-2 life cycle that could be tested in in vitro and in vivo models. Although cholesterol is an essential component of immune cell membranes, excess levels can dysregulate protective immunity and promote exaggerated pulmonary and systemic inflammatory responses. Statins block the production of multiple sterols, oxysterols and isoprenoids, resulting in a pleiotropic range of context-dependent effects on virus infectivity, immunity and inflammation. We highlight antiviral, immunomodulatory and anti-inflammatory effects of cholesterol-modifying drugs that merit further consideration in the management of SARS-CoV-2 infection.

scholarly journals Diflunisal targets the HMGB1/CXCL12 heterocomplex and blocks immune cell recruitment

2019 ◽  
Author(s):  
Federica De Leo ◽  
Giacomo Quilici ◽  
Mario Tirone ◽  
Valeria Mannella ◽  
Francesco De Marchis ◽  
...  
Keyword(s):  
Rational Design ◽  
Immune Cell ◽  
Inflammatory Responses ◽  
Inflammatory Cells ◽  
Cell Recruitment ◽  
Redox States ◽  
Inflammatory Drugs ◽  
Anti Inflammatory

AbstractExtracellular HMGB1 triggers inflammation following infection or injury, and supports tumorigenesis in inflammation-related malignancies. HMGB1 has several redox states: reduced HMGB1 recruits inflammatory cells to injured tissues forming a heterocomplex with CXCL12 and signaling via its receptor CXCR4; disulfide-containing HMGB1 binds to TLR4 and promotes inflammatory responses. Here we show that Diflunisal, an aspirin-like nonsteroidal anti-inflammatory drug (NSAID) that has been in clinical use for decades, specifically inhibits in vitro and in vivo the chemotactic activity of HMGB1 at nanomolar concentrations, at least in part by binding directly to both HMGB1 and CXCL12 and disrupting their heterocomplex. Importantly, Diflunisal does not inhibit TLR4-dependent responses. Our findings clarify the mode of action of Diflunisal, and open the way to the rational design of functionally specific anti-inflammatory drugs.

scholarly journals The amino acid sensor GCN2 inhibits inflammatory responses to apoptotic cells promoting tolerance and suppressing systemic autoimmunity

2015 ◽  
Vol 112 (34) ◽  
pp. 10774-10779 ◽  
Author(s):  
Buvana Ravishankar ◽  
Haiyun Liu ◽  
Rahul Shinde ◽  
Kapil Chaudhary ◽  
Wei Xiao ◽  
...  
Keyword(s):  
Cell Activation ◽  
Immune Cell ◽  
Inflammatory Responses ◽  
Apoptotic Cell ◽  
Metabolic Stress ◽  
Tolerance Induction ◽  
Apoptotic Cells ◽  
Cell Clearance

Efficient apoptotic cell clearance and induction of immunologic tolerance is a critical mechanism preventing autoimmunity and associated pathology. Our laboratory has reported that apoptotic cells induce tolerance by a mechanism dependent on the tryptophan catabolizing enzyme indoleamine 2,3 dioxygenase 1 (IDO1) in splenic macrophages (MΦ). The metabolic-stress sensing protein kinase GCN2 is a primary downstream effector of IDO1; thus, we tested its role in apoptotic cell-driven immune suppression. In vitro, expression of IDO1 in MΦs significantly enhanced apoptotic cell-driven IL-10 and suppressed IL-12 production in a GCN2-dependent mechanism. Suppression of IL-12 protein production was due to attenuation of IL-12 mRNA association with polyribosomes inhibiting translation while IL-10 mRNA association with polyribosomes was not affected. In vivo, apoptotic cell challenge drove a rapid, GCN2-dependent stress response in splenic MΦs with increased IL-10 and TGF-β production, whereas myeloid-specific deletion of GCN2 abrogated regulatory cytokine production with provocation of inflammatory T-cell responses to apoptotic cell antigens and failure of long-tolerance induction. Consistent with a role in prevention of apoptotic cell driven autoreactivity, myeloid deletion of GCN2 in lupus-prone mice resulted in increased immune cell activation, humoral autoimmunity, renal pathology, and mortality. In contrast, activation of GCN2 with an agonist significantly reduced anti-DNA autoantibodies and protected mice from disease. Thus, this study implicates a key role for GCN2 signals in regulating the tolerogenic response to apoptotic cells and limiting autoimmunity.

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