scholarly journals Prokaryotic ribosomal RNA stimulates zebrafish embryonic innate immune system

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Abhishikta Basu ◽  
Maki Yoshihama ◽  
Tamayo Uechi ◽  
Naoya Kenmochi
Keyword(s):  
Immune System ◽  
Innate Immune System ◽  
18S Rrna Gene ◽  
Positive Control ◽  
Innate Immune ◽  
Rrna Gene ◽  
Immune Recognition ◽  
E Coli

Abstract Objectives Cell-culture studies reported that prokaryotic RNA molecules among the various microbe-associated molecular patterns (MAMPs) were uniquely present in live bacteria and were categorized as viability-associated MAMPs. They also reported that specific nucleotide modifications are instrumental in the discrimination between self and nonself RNAs. The aim of this study was to characterize the in vivo immune induction potential of prokaryotic and eukaryotic ribosomal RNAs (rRNAs) using zebrafish embryos as novel whole animal model system. Additionally, we aimed to test the possible role of rRNA modifications in immune recognition. Results We used three immune markers to evaluate the induction potential of prokaryotic rRNA derived from Escherichia coli and eukaryotic rRNAs from chicken (nonself) and zebrafish (self). Lipopolysaccharide (LPS) of Pseudomonas aeruginosa served as a positive control. E. coli rRNA had an induction potential equivalent to that of LPS. The zebrafish innate immune system could discriminate between self and nonself rRNAs. Between the nonself rRNAs, E. coli rRNA was more immunogenic than chicken rRNA. The in vitro transcript of zebrafish 18S rRNA gene without the nucleotide modifications was not recognized by its own immune system. Our data suggested that prokaryotic rRNA is immunostimulatory in vivo and could be useful as an adjuvant.

scholarly journals Interaction of Antibiotics with Innate Host Defense Factors against Salmonella enterica Serotype Newport

mSphere ◽  
2017 ◽  
Vol 2 (6) ◽  
Author(s):  
George Sakoulas ◽  
Monika Kumaraswamy ◽  
Armin Kousha ◽  
Victor Nizet
Keyword(s):  
Innate Immunity ◽  
Immune System ◽  
Innate Immune System ◽  
Salmonella Enterica ◽  
Clinical Performance ◽  
Innate Immune ◽  
Content Type ◽  
Antimicrobial Assay

ABSTRACT It is becoming increasingly understood that the current paradigms of in vitro antimicrobial susceptibility testing may have significant shortcomings in predicting activity in vivo. This study evaluated the activity of several antibiotics alone and in combination against clinical isolates of Salmonella enterica serotype Newport (meningitis case) utilizing both conventional and physiological media. In addition, the interactions of these antibiotics with components of the innate immune system were evaluated. Azithromycin, which has performed quite well clinically despite high MICs in conventional media, was shown to be more active in physiological media and to enhance innate immune system killing. Alternatively, chloramphenicol did not show enhanced immune system killing, paralleling its inferior clinical performance to other antibiotics that have been used to treat Salmonella meningitis. These findings are important additions to the building understanding of current in vitro antimicrobial assay limitations that hopefully will amount to future improvements in these assays to better predict clinical efficacy and activity in vivo. This study examines the pharmacodynamics of antimicrobials that are used to treat Salmonella with each other and with key components of the innate immune system. Antimicrobial synergy was assessed using time-kill and checkerboard assays. Antimicrobial interactions with innate immunity were studied by employing cathelicidin LL-37, whole-blood, and neutrophil killing assays. Ceftriaxone and ciprofloxacin were found to be synergistic in vitro against Salmonella enterica serotype Newport. Ceftriaxone, ciprofloxacin, and azithromycin each demonstrated synergy with the human cathelicidin defense peptide LL-37 in killing Salmonella. Exposure of Salmonella to sub-MICs of ceftriaxone resulted in enhanced susceptibility to LL-37, whole blood, and neutrophil killing. The activity of antibiotics in vivo against Salmonella may be underestimated in bacteriologic media lacking components of innate immunity. The pharmacodynamic interactions of antibiotics used to treat Salmonella with each other and with components of innate immunity warrant further study in light of recent findings showing in vivo selection of antimicrobial resistance by single agents in this pathogen. IMPORTANCE It is becoming increasingly understood that the current paradigms of in vitro antimicrobial susceptibility testing may have significant shortcomings in predicting activity in vivo. This study evaluated the activity of several antibiotics alone and in combination against clinical isolates of Salmonella enterica serotype Newport (meningitis case) utilizing both conventional and physiological media. In addition, the interactions of these antibiotics with components of the innate immune system were evaluated. Azithromycin, which has performed quite well clinically despite high MICs in conventional media, was shown to be more active in physiological media and to enhance innate immune system killing. Alternatively, chloramphenicol did not show enhanced immune system killing, paralleling its inferior clinical performance to other antibiotics that have been used to treat Salmonella meningitis. These findings are important additions to the building understanding of current in vitro antimicrobial assay limitations that hopefully will amount to future improvements in these assays to better predict clinical efficacy and activity in vivo.

scholarly journals Mycobacterium avium Biofilm Attenuates Mononuclear Phagocyte Function by Triggering Hyperstimulation and Apoptosis during Early Infection

2013 ◽  
Vol 82 (1) ◽  
pp. 405-412 ◽  
Author(s):  
Sasha J. Rose ◽  
Luiz E. Bermudez
Keyword(s):  
Immune System ◽  
Mycobacterium Avium ◽  
Innate Immune System ◽  
Innate Immune ◽  
Mononuclear Phagocytes ◽  
Bacterial Killing ◽  
Content Type ◽  
Tnf Α

ABSTRACTMycobacterium aviumsubsp.hominissuisis an opportunistic human pathogen that has been shown to form biofilmin vitroandin vivo. Biofilm formationin vivoappears to be associated with infections in the respiratory tract of the host. The reasoning behind howM. aviumsubsp.hominissuisbiofilm is allowed to establish and persist without being cleared by the innate immune system is currently unknown. To identify the mechanism responsible for this, we developed anin vitromodel using THP-1 human mononuclear phagocytes cocultured with establishedM. aviumsubsp.hominissuisbiofilm and surveyed various aspects of the interaction, including phagocyte stimulation and response, bacterial killing, and apoptosis.M. aviumsubsp.hominissuisbiofilm triggered robust tumor necrosis factor alpha (TNF-α) release from THP-1 cells as well as superoxide and nitric oxide production. Surprisingly, the hyperstimulated phagocytes did not effectively eliminate the cells of the biofilm, even when prestimulated with gamma interferon (IFN-γ) or TNF-α or cocultured with natural killer cells (which have been shown to induce anti-M. aviumsubsp.hominissuisactivity when added to THP-1 cells infected with planktonicM. aviumsubsp.hominissuis). Time-lapse microscopy and the TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) assay determined that contact with theM. aviumsubsp.hominissuisbiofilm led to early, widespread onset of apoptosis, which is not seen until much later in planktonicM. aviumsubsp.hominissuisinfection. Blocking TNF-α or TNF-R1 during interaction with the biofilm significantly reduced THP-1 apoptosis but did not lead to elimination ofM. aviumsubsp.hominissuis. Our data collectively indicate thatM. aviumsubsp.hominissuisbiofilm induces TNF-α-driven hyperstimulation and apoptosis of surveilling phagocytes, which prevents clearance of the biofilm by cells of the innate immune system and allows the biofilm-associated infection to persist.

scholarly journals Redundant Catalases Detoxify Phagocyte Reactive Oxygen and Facilitate Histoplasma capsulatum Pathogenesis

2013 ◽  
Vol 81 (7) ◽  
pp. 2334-2346 ◽  
Author(s):  
Eric D. Holbrook ◽  
Katherine A. Smolnycki ◽  
Brian H. Youseff ◽  
Chad A. Rappleye
Keyword(s):  
Immune System ◽  
Innate Immune System ◽  
Histoplasma Capsulatum ◽  
Reactive Oxygen ◽  
Innate Immune ◽  
Respiratory Pathogen ◽  
Human Neutrophils ◽  
Content Type

ABSTRACTHistoplasma capsulatumis a respiratory pathogen that infects phagocytic cells. The mechanisms allowingHistoplasmato overcome toxic reactive oxygen molecules produced by the innate immune system are an integral part ofHistoplasma's ability to survive during infection. To probe the contribution ofHistoplasmacatalases in oxidative stress defense, we created and analyzed the virulence defects of mutants lacking CatB and CatP, which are responsible for extracellular and intracellular catalase activities, respectively. Both CatB and CatP protectedHistoplasmafrom peroxide challengein vitroand from antimicrobial reactive oxygen produced by human neutrophils and activated macrophages. Optimal protection required both catalases, as the survival of a double mutant lacking both CatB and CatP was lower than that of single-catalase-deficient cells. Although CatB contributed to reactive oxygen species defensesin vitro, CatB was dispensable for lung infection and extrapulmonary disseminationin vivo. Loss of CatB from a strain also lacking superoxide dismutase (Sod3) did not further reduce the survival ofHistoplasmayeasts. Nevertheless, some catalase function was required for pathogenesis since simultaneous loss of both CatB and CatP attenuatedHistoplasmavirulencein vivo. These results demonstrate thatHistoplasma's dual catalases comprise a system that enablesHistoplasmato efficiently overcome the reactive oxygen produced by the innate immune system.

In Vitro Modulation of E. coli Community Behavior and Human Innate Immune System by Lantibiotic Nisin

2012 ◽  
Vol 18 (3) ◽  
pp. 171-183 ◽  
Author(s):  
Deovrat N. Begde ◽  
Sunita B. Bundale ◽  
Mashitha V. Pise ◽  
Jaishree A. Rudra ◽  
Nandita A. Nashikkar ◽  
...  
Keyword(s):  
Immune System ◽  
Innate Immune System ◽  
Innate Immune ◽  
E Coli ◽  
Community Behavior

scholarly journals Flagellar Motility Is a Key Determinant of the Magnitude of the Inflammasome Response to Pseudomonas aeruginosa

2013 ◽  
Vol 81 (6) ◽  
pp. 2043-2052 ◽  
Author(s):  
Yash R. Patankar ◽  
Rustin R. Lovewell ◽  
Matthew E. Poynter ◽  
Jeevan Jyot ◽  
Barbara I. Kazmierczak ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Immune System ◽  
Innate Immune System ◽  
Innate Immune ◽  
Bacterial Motility ◽  
Inflammasome Activation ◽  
Flagellar Motility ◽  
Content Type

ABSTRACTWe previously demonstrated that bacterial flagellar motility is a fundamental mechanism by which host phagocytes bind and ingest bacteria. Correspondingly, loss of bacterial motility, consistently observed in clinical isolates from chronicPseudomonas aeruginosainfections, enables bacteria to evade association and ingestion ofP. aeruginosaby phagocytes bothin vitroandin vivo. Since bacterial interactions with the phagocyte cell surface are required for type three secretion system-dependent NLRC4 inflammasome activation byP. aeruginosa, we hypothesized that reduced bacterial association with phagocytes due to loss of bacterial motility, independent of flagellar expression, will lead to reduced inflammasome activation. Here we report that inflammasome activation is reduced in response to nonmotileP. aeruginosa. NonmotileP. aeruginosaelicits reduced IL-1β production as well as caspase-1 activation by peritoneal macrophages and bone marrow-derived dendritic cellsin vitro. Importantly, nonmotileP. aeruginosaalso elicits reduced IL-1β levelsin vivoin comparison to those elicited by wild-typeP. aeruginosa. This is the first demonstration that loss of bacterial motility results in reduced inflammasome activation and antibacterial IL-1β host response. These results provide a critical insight into how the innate immune system responds to bacterial motility and, correspondingly, how pathogens have evolved mechanisms to evade the innate immune system.

216 Anti-tumor activity of iosH2 by blocking LILRB2 receptor signalling

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A235-A235
Author(s):  
Osiris Marroquin Belaunzaran ◽  
Anahita Rafiei ◽  
Anil Kumar ◽  
Julia Kolibaba ◽  
Lorenz Vogt ◽  
...  
Keyword(s):  
T Cells ◽  
Immune System ◽  
T Cell ◽  
Innate Immune System ◽  
Cell Activation ◽  
Innate Immune ◽  
High Affinity ◽  
Anti Tumor Activity

BackgroundThe human leukocyte immunoglobulin-like receptor family B (LILR B) acts as check point blockade of the innate immune system by inhibiting leukocyte activation through SHP phosphatase recruitment. Some of the physiological ligands include classical HLA class I molecules, including beta-2-microglobulin (B2M) free open conformers (OC). Natural HLA-OC expression is known from autoimmune disease leading to immune activation by pleiotropic effects since they bind to LILRB and KIR family members reducing Treg and MDSC numbers and increased effector T-cell and NK-cell activation, respectively. We have generated an IgG4-HLA-57 open conformer (OC) molecule (iosH2) with high affinity for LILRB molecules and demonstrate its anti-cancer activity in vitro and in vivo.Methods iosH2 was produced by transient gene expression in CHO cells and purified by standard chromatography. Affinity of iosH2 binding was quantified by ELISA and SPR analysis. HLA-G mediated signaling and competition was assessed using functional cell lines. Effect of iosH2 on activation of SHP1/2 was assessed using Western Blot. Functional assays including in vitro polarization and phagocytosis potential of primary macrophages was assessed by flow cytometry in the presence of iosH2 or isotype control. Effect of iosH2 on T cell activation was evaluated in co-cultures of cancer and T cells. Mouse models were used to assess in vivo activity.Results iosH2 binds to LILRB2 with high affinity and blocks the activation of HLA-G. In addition, iosH2 blocks receptor-mediated activation of SHP1/2. iosH2 promotes a shift from M2 to M1 macrophages with enhanced tumor cell phagocytosis in vitro. iosH2 enhances activation and killing potential of T cells in cancer cells and T cells co-culture assay. iosH2 exerts therapeutic efficacy in mouse transgenic (melanoma) and different syngeneic tumor models (e.g. pancreatic, colon and breast cancer) as monotherapy. Moreover, it acts synergistically in vivo with PD1 blocking antibodies achieving long-term tumor control. Ex vivo tumor sample analysis demonstrates a significant reduction of MDSC and Tregs and a shift towards an activated inflammatory M1 macrophage phenotype. Loss of MDSC functionality was paralleled by enhanced CD8+ T cell expansion and activity.Conclusions iosH2 binds to LILRB2 with high affinity, restores immune cell function in vitro and demonstrates anti-tumor activity in different in vivo mouse models. In addition, it acts synergistically in vivo with PD1. iosH2 is a first-in-class OC therapeutic with robust anti-tumor activity by promoting key components of the innate immune system. Clinical development is under way and phase I trial in preparation.

scholarly journals CD46 Plays a Key Role in Tailoring Innate Immune Recognition of Apoptotic and Necrotic Cells

2005 ◽  
Vol 280 (43) ◽  
pp. 36342-36354 ◽  
Author(s):  
Kristina Elward ◽  
Mark Griffiths ◽  
Masashi Mizuno ◽  
Claire L. Harris ◽  
Jim W. Neal ◽  
...  
Keyword(s):  
Immune System ◽  
Innate Immune System ◽  
Apoptotic Cell ◽  
Membrane Attack Complex ◽  
Factor H ◽  
Innate Immune ◽  
Immune Recognition ◽  
Alternative Pathways ◽  
Complement Regulator ◽  
Complement C1q

Complement is the canonical innate immune system involved in host defense and tissue repair with the clearance of cell debris. In contrast to the robust armory mounted against microbial nonself-pathogens, complement is selectively activated on altered self (i.e. apoptotic and necrotic cells) to instruct the safe demise by poorly characterized mechanisms. Our data shed new light on the role of complement C1q in sensing nucleic acids (NA) rapidly exposed on apoptotic Jurkat T cell membranes and in driving C3 opsonization but without the lytic membrane attack complex. DNA/RNase-treated apoptotic cells failed to activate complement. We found that several other apoptotic cell models, including senescent keratinocytes, ionophore-treated sperm cells, and CMK-derived platelets, stained for cleaved caspase 3 were rapidly losing the key complement regulator CD46. CD46 from nuclear and membrane stores was found to cluster into blebs and shed into microparticles together with NA, phosphatidylserine, C1q, and factor H. Classical and alternative pathways of complement were involved in the recognition of H2O2-treated necrotic cells. Membrane attack complex was detected on necrotic cells possibly as a result of CD46 and CD59 shedding into soluble forms. Our data highlight a novel and universal paradigm whereby the complement innate immune system is using two synergistic strategies with the recognition of altered self-NA and missing self-CD46 signals to instruct and tailor the efficient removal of apoptotic and necrotic cells in immunoprivileged sites.

scholarly journals Effect of Hypoglycemia on Inflammatory Responses and the Response to Low-Dose Endotoxemia in Humans

2018 ◽  
Vol 104 (4) ◽  
pp. 1187-1199 ◽  
Author(s):  
Ahmed Iqbal ◽  
Lynne R Prince ◽  
Peter Novodvorsky ◽  
Alan Bernjak ◽  
Mark R Thomas ◽  
...  
Keyword(s):  
Cardiovascular Risk ◽  
Immune System ◽  
Innate Immune System ◽  
Low Dose ◽  
Inflammatory Responses ◽  
Platelet Reactivity ◽  
Innate Immune ◽  
Endotoxin Challenge ◽  
Intermediate Monocytes

Abstract Context Hypoglycemia is emerging as a risk for cardiovascular events in diabetes. We hypothesized that hypoglycemia activates the innate immune system, which is known to increase cardiovascular risk. Objective To determine whether hypoglycemia modifies subsequent innate immune system responses. Design and Setting Single-blinded, prospective study of three independent parallel groups. Participants and Interventions Twenty-four healthy participants underwent either a hyperinsulinemic-hypoglycemic (2.5 mmol/L), euglycemic (6.0 mmol/L), or sham-saline clamp (n = 8 for each group). After 48 hours, all participants received low-dose (0.3 ng/kg) intravenous endotoxin. Main Outcome Measures We studied in-vivo monocyte mobilization and monocyte-platelet interactions. Results Hypoglycemia increased total leukocytes (9.98 ± 1.14 × 109/L vs euglycemia 4.38 ± 0.53 × 109/L, P < 0.001; vs sham-saline 4.76 ± 0.36 × 109/L, P < 0.001) (mean ± SEM), mobilized proinflammatory intermediate monocytes (42.20 ± 7.52/μL vs euglycemia 20.66 ± 3.43/μL, P < 0.01; vs sham-saline 26.20 ± 3.86/μL, P < 0.05), and nonclassic monocytes (36.16 ± 4.66/μL vs euglycemia 12.72 ± 2.42/μL, P < 0.001; vs sham-saline 19.05 ± 3.81/μL, P < 0.001). Following hypoglycemia vs euglycemia, platelet aggregation to agonist (area under the curve) increased (73.87 ± 7.30 vs 52.50 ± 4.04, P < 0.05) and formation of monocyte-platelet aggregates increased (96.05 ± 14.51/μL vs 49.32 ± 6.41/μL, P < 0.05). Within monocyte subsets, hypoglycemia increased aggregation of intermediate monocytes (10.51 ± 1.42/μL vs euglycemia 4.19 ± 1.08/μL, P < 0.05; vs sham-saline 3.81± 1.42/μL, P < 0.05) and nonclassic monocytes (9.53 ± 1.08/μL vs euglycemia 2.86 ± 0.72/μL, P < 0.01; vs sham-saline 3.08 ± 1.01/μL, P < 0.05), with platelets compared with controls. Hypoglycemia led to greater leukocyte mobilization in response to subsequent low-dose endotoxin challenge (10.96 ± 0.97 vs euglycemia 8.21 ± 0.85 × 109/L, P < 0.05). Conclusions Hypoglycemia mobilizes monocytes, increases platelet reactivity, promotes interaction between platelets and proinflammatory monocytes, and potentiates the subsequent immune response to endotoxin. These changes may contribute to increased cardiovascular risk observed in people with diabetes.

scholarly journals In Vivo Discrimination of Type 3 Secretion System-Positive and -Negative Pseudomonas aeruginosa via a Caspase-1-Dependent Pathway

2010 ◽  
Vol 78 (11) ◽  
pp. 4744-4753 ◽  
Author(s):  
Tamding Wangdi ◽  
Lilia A. Mijares ◽  
Barbara I. Kazmierczak
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Immune System ◽  
Innate Immune System ◽  
Interleukin 1 ◽  
Innate Immune ◽  
Secretion Systems ◽  
Caspase 1 ◽  
Molecular Patterns ◽  
Type 3

ABSTRACT Microbe-associated molecular patterns are recognized by Toll-like receptors of the innate immune system. This recognition enables a rapid response to potential pathogens but does not clearly provide a way for the innate immune system to discriminate between virulent and avirulent microbes. We find that pulmonary infection of mice with type 3 translocation-competent Pseudomonas aeruginosa triggers a rapid inflammatory response, while infection with isogenic translocation-deficient mutants does not. Discrimination between translocon-positive and -negative bacteria requires caspase-1 activity in bone marrow-derived cells and interleukin-1 receptor signaling. Thus, the activation of caspase-1 by bacteria expressing type 3 secretion systems allows for rapid recognition of bacteria expressing conserved functions associated with virulence.

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