scholarly journals Inhibition of Complement and CD14 Attenuates the Escherichia coli-Induced Inflammatory Response in Porcine Whole Blood

2008 ◽  
Vol 77 (2) ◽  
pp. 725-732 ◽  
Author(s):  
Ebbe Billmann Thorgersen ◽  
Anne Pharo ◽  
Karin Haverson ◽  
Anne K. Axelsen ◽  
Peter Gaustad ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Inflammatory Response ◽  
Proinflammatory Cytokines ◽  
Whole Blood ◽  
Innate Immune ◽  
Dependent Manner ◽  
Bacterial Clearance ◽  
E Coli ◽  
Tnf Α ◽  
Dose Dependent

ABSTRACT The innate immune response is a double-edged sword in systemic inflammation and sepsis. Uncontrolled or inappropriate activation can damage and be lethal to the host. Several studies have investigated inhibition of downstream mediators, including tumor necrosis factor alpha (TNF-α) and interleukin-1β (IL-1β). Emerging evidence indicates that upstream inhibition is a better therapeutic approach for attenuating damaging immune activation. Therefore, we investigated inhibition of two central innate immune pathways, those of complement and CD14/Toll-like receptor 4 (TLR4)/myeloid differentiation protein 2 (MD-2), in a porcine in vitro model of Escherichia coli-induced inflammation. Porcine whole blood anticoagulated with lepuridin, which did not interfere with the complement system, was incubated with E. coli lipopolysaccharide (LPS) or whole bacteria. Inhibitors of complement and CD14 and thus the LPS CD14/TLR4/MD-2 receptor complex were tested to investigate the effect on the inflammatory response. A broad range of inflammatory readouts were used to monitor the effect. Anti-CD14 was found to saturate the CD14 molecule on granulocytes and completely inhibited LPS-induced proinflammatory cytokines in a dose-dependent manner. Anti-CD14 significantly reduced the levels of the E. coli-induced proinflammatory cytokines TNF-α and IL-1β, but not IL-8, in a dose-dependent manner. No effect on bacterial clearance was seen. Vaccinia complement control protein and smallpox inhibitor of complement enzymes, two Orthopoxvirus-encoded complement inhibitors, completely inhibited complement activation. Furthermore, these agents almost completely inhibited the expression of wCD11R3, which is associated with CD18 as a β2 integrin, on porcine granulocytes and decreased IL-8 levels significantly in a dose-dependent manner. As expected, complement inhibition reduced bacterial clearance. We conclude that inhibition of complement and CD14 attenuates E. coli-induced inflammation and might be used as a therapeutic regimen in gram-negative sepsis along with appropriate treatment with antibiotics.

scholarly journals Intracellular Free Iron and Its Potential Role in Ultrahigh-Pressure-Induced Inactivation of Escherichia coli

2012 ◽  
Vol 79 (2) ◽  
pp. 722-724 ◽  
Author(s):  
Yuan Yan ◽  
Joy G. Waite-Cusic ◽  
Periannan Kuppusamy ◽  
Ahmed E. Yousef
Keyword(s):  
Escherichia Coli ◽  
Iron Chelator ◽  
Ultrahigh Pressure ◽  
Dependent Manner ◽  
Paramagnetic Resonance ◽  
Free Iron ◽  
Content Type ◽  
E Coli ◽  
Electron Paramagnetic ◽  
Dose Dependent

ABSTRACTIntracellular free iron ofEscherichia coliwas determined by whole-cell electron paramagnetic resonance spectrometry. Ultrahigh pressure (UHP) increased both intracellular free iron and cell lethality in a pressure-dose-dependent manner. The iron chelator 2,2′-dipyridyl protected cells against UHP treatments. A mutation that produced iron overload conditions sensitizedE. colito UHP treatment.

Differential modulation of innate immune response by epinephrine and estradiol

2017 ◽  
Vol 30 (3) ◽  
Author(s):  
Sona Margaryan ◽  
Armenuhi Hyusyan ◽  
Anush Martirosyan ◽  
Shushan Sargsian ◽  
Gayane Manukyan
Keyword(s):  
Immune Response ◽  
Innate Immune Response ◽  
Inflammatory Responses ◽  
Innate Immune ◽  
Dependent Manner ◽  
Induced Expression ◽  
Short Term Exposure ◽  
Tnf Α ◽  
Vitro Effect ◽  
Dose Dependent

AbstractBackgroundAlthough it is widely accepted that catecholamines and estrogens influence immunity and have consequences for health, their effect on innate immunity (e.g. monocytes and neutrophils) is still not fully investigated.Materials and methodsOur study aimed to analyze the production of tumor necrosis factor (TNF)-α, interleukin (IL)-1β, monocyte chemoattractant protein (MCP)-1 and IL-8 by whole blood cells following short-term exposure to epinephrine (Epi) and 17β-estradiol (E2) in the presence or absence of lipopolysaccharide (LPS). We also evaluated the in vitro effect of these hormones on expression of β2 integrin (CD11b/CD18) and L-selectin (CD62L) by circulating neutrophils and monocytes in the blood of healthy subjects.ResultsEpi has shown a potential to modulate the production of pro-inflammatory mediators. Its exposure resulted in significantly increased production of IL-8 in a dose-dependent manner. On the contrary, a dose-dependent suppression of LPS-induced production of IL-1β, IL-8, and MCP-1 by Epi was observed. In neutrophils, a modest rise in CD11b expression was observed after Epi exposure. Simultaneously, Epi suppressed LPS-induced expression of CD11b and CD18. In monocytes, Epi suppressed LPS-induced expression of C11b. E2 inhibited LPS-induced TNF-α production and caused a significant decrease in CD62L expression in both cell populations. No significant changes were observed after double exposure of cells with Epi and E2.ConclusionsThus, our results show that Epi and E2 differentially modulate the innate immune response and have a dual effect on cytokine modulation. The findings suggest that the observed immunoregulatory role of Epi and E2 may influence the outcome in endotoxin responses and can be critical in the regulation of inflammatory responses.

The Zinc Finger Protein Gfi1 Controls TLR4-Mediated Inflammatory Response by Directly Antagonizing NF-κB Transcription Factor

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 469-469
Author(s):  
Ehssan Sharif-Askari ◽  
Hui Zeng ◽  
Lothar Vassen ◽  
Christian Kosan ◽  
Cyrus Khandanpour ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Direct Interaction ◽  
Innate Immune ◽  
Dependent Manner ◽  
Negative Regulators ◽  
Tlr Signaling ◽  
Tnf Α ◽  
Dose Dependent ◽  
Lps Treatment ◽  
Stimulation Of

Abstract Inflammatory responses are complex and comprise multiple mediators including cytokines such as TNF-alpha (TNF-α) and IL-1beta. These cytokines are synthesized and secreted in response to signaling by plasma membrane receptors of the Toll-like receptor (TLR) family. A central downstream element of TLR-dependent signaling is the transcription factor NF-kappaB (NF-κB), which plays a pivotal role in controlling the proper sequence of events during an inflammatory response. In unstimulated cells, NF-κB is bound to inhibitory IkappaB (IκB) proteins and remains sequestered in the cytoplasm. Stimulation of TLRs triggers a signaling cascade that leads to phosphorylation and proteasomal degradation of IκB, resulting in the translocation of NF-κB to the nucleus, where it acts as a transcriptional activator of target genes. To keep the innate immune system under control, the TLR signaling cascade is under a tight control of many positive and negative regulators. We have previously shown that the transcription factor Growth Factor Independence 1 (Gfi1) represents a novel factor limiting the inflammatory immune response including TNF-α. Gfi1-deficient (Gfi1−/−) mice show a very strong systemic response to the TLR4 ligand and endotoxin LPS and die rapidly within 36 h with symptoms of septic shock. Here, we investigated the molecular mechanism of this exaggerated TNF-α production in the absence of Gfi1. It is known that endotoxin stimulation results in the activation of the transcription factor NF-κB through TLR4, leading to TNF-α production. This activation also resulted in rapid and de novo expression of Gfi1 in the nucleus in a time- and dose-dependent manner. The expression of Gfi1 was not due to feedback regulation from secreted TNF, since TNF-deficient macrophages were also able to upregulate Gfi1 mRNA following LPS stimulation. As expected, LPS stimulation of Gfi1−/− macrophages resulted in significantly higher levels of TNF-α mRNA, and secreted TNF-α cytokine. Strikingly and in contrast to most known negative regulators of TLRs, Gfi1 did not affect the activity or the expression levels of the cytoplasmic components of TLR signaling pathway. Additionally, NF-κB phosphorylation and nuclear translocation post- LPS treatment were intact in both Gfi1−/− and Gfi1+/+ macrophages. Immunoprecipitation analysis from cells endogenously expressing Gfi1 and NF-κB or over-expressing these two proteins post transfection, clearly revealed a direct interaction between Gfi1 and the p65 subunit of NF-κB. Immunofluorescence staining of macrophages post-LPS treatment confirmed direct interaction of these two proteins in the nucleus at the endogenous level. Gfi1 represses transcription by binding to DNA recognition sequences in target gene promoters. Thus, aiming to investigate the effect of Gfi1 expression on NF-κB nuclear signaling, we found that LPS treatment enhances NF-κB DNA binding activity in Gfi1−/− macrophages as compared to Gfi1+/+ cells. Furthermore, over expression of Gfi1 protein resulted in negative regulation of NF-κB mediated gene activation in a dose-dependent manner. Chromatin immune precipitation with anti-p65 antibodies from LPS stimulated Gfi1+/+ and Gfi1−/− macrophages revealed enhanced NF-κB promoter occupancy at the TNF gene in Gfi1−/− macrophages as compared to Gfi1+/+ cells. In conclusion, our findings reveal a novel function for Gfi1 in the innate immune response by directly antagonizing NF-κB function. This molecular perceptive of TNF-α regulation during inflammation may provide an attractive strategy for therapeutic intervention in chronic inflammatory diseases and certain cancers.

Aqueous extracts ofCimicifuga racemosaand phenolcarboxylic constituents inhibit production of proinflammatory cytokines in LPS-stimulated human whole blood

2009 ◽  
Vol 87 (11) ◽  
pp. 963-972 ◽  
Author(s):  
Diethart Schmid ◽  
Florian Woehs ◽  
Martin Svoboda ◽  
Theresia Thalhammer ◽  
Peter Chiba ◽  
...  
Keyword(s):  
Proinflammatory Cytokines ◽  
Whole Blood ◽  
Black Cohosh ◽  
Active Principle ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Antiinflammatory Action ◽  
Tnf Α ◽  
Isoferulic Acid ◽  
Ifn Γ

Cimicifuga racemosa (black cohosh) is commonly used in traditional medicines as treatment for menopausal symptoms and as an antiinflammatory remedy. To clarify the mechanism of action and active principle for the antiinflammatory action, the effects of aqueous C. racemosa root extracts (CRE) and its major constituents on the release of the proinflammatory cytokines IL-6, TNF-α, IFN-γ, and the chemokine IL-8 were investigated in lipopolysaccharide (LPS)-stimulated whole blood of healthy volunteers. CRE (3 µg/µL and 6 µg/µL) reduced LPS-induced release of IL-6 and TNF-α in a concentration- and time-dependent manner and almost completely blocked release of IFN-γ into the plasma supernatant. Except for IFN-γ, these effects were attenuated at longer incubation periods. IL-8 secretion was stimulated by CRE. As shown by quantitative real-time RT-PCR, effects on cytokines were based on preceding changes in mRNA levels except for IL-8. According to their content in CRE, the phenolcarboxylic compounds caffeic acid, ferulic acid, and isoferulic acid, as well as the triterpene glycosides 23-epi-26-deoxyactein and cimigenol-3-O-xyloside, were tested at representative concentrations. Among these, isoferulic acid was the prominent active principle in CRE, responsible for the observed inhibition of IL-6, TNF-α, and IFN-γ, but not for IL-8 stimulation. The effect of this compound may explain the antiinflammatory activities of CRE and its beneficial actions in rheumatism and other inflammatory diseases.

Cloning and expression of a functional rat liver D-β-hydroxybutyrate dehydrogenase-β-galactosidase fusion protein in Escherichia coli

1991 ◽  
Vol 69 (9) ◽  
pp. 670-673
Author(s):  
Sharon Churchill ◽  
Perry Churchill
Keyword(s):  
Escherichia Coli ◽  
Rat Liver ◽  
Polyclonal Antibodies ◽  
Cloning And Expression ◽  
Dependent Manner ◽  
Bacteriophage Λ ◽  
Expression Library ◽  
Expression Plasmid ◽  
E Coli ◽  
Dose Dependent

A rat liver bacteriophage λ expression library was probed using polyclonal antibodies raised to purified rat liver D-β-hydroxybutyrate dehydrogenase (BDH). A clone was selected that contained a 1.2-kb insert. The insert placed in an expression plasmid was utilized to transform Escherichia coli. These cells were shown to possess phosphatidylcholine-dependent BDH activity. Cells transformed with only the plasmid had no detectable BDH activity in the presence of phosphatidylcholine. The expressed activity in E. coli could be inhibited in a dose-dependent manner by BDH antiserum.Key words: D-β-hydroxybutyrate dehydrogenase, cloning, expression.

scholarly journals The Escherichia coli Common Pilus and the Bundle-Forming Pilus Act in Concert during the Formation of Localized Adherence by Enteropathogenic E. coli

2009 ◽  
Vol 191 (11) ◽  
pp. 3451-3461 ◽  
Author(s):  
Zeus Saldaña ◽  
Ayşen L. Erdem ◽  
Stephanie Schüller ◽  
Iruka N. Okeke ◽  
Mark Lucas ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epithelial Cells ◽  
Cultured Cells ◽  
Dependent Manner ◽  
Triple Mutant ◽  
E Coli ◽  
Accessory Factor ◽  
Isogenic Mutants ◽  
Dose Dependent ◽  
Background Expression

ABSTRACT Although the bundle-forming pilus (BFP) of enteropathogenic Escherichia coli (EPEC) mediates microcolony formation on epithelial cells, the adherence of BFP-deficient mutants is significantly abrogated, but the mutants are still adherent due to the presence of intimin and possibly other adhesins. In this study we investigated the contribution of the recently described E. coli common pilus (ECP) to the overall adherence properties of EPEC. We found that ECP and BFP structures can be simultaneously observed in the course (between zero time and 7 h during infection) of formation of localized adherence on cultured epithelial cells. These two pilus types colocalized at different levels of the microcolony topology, tethering the adhering bacteria. No evidence of BFP disappearance was found after prolonged infection. When expressed from a plasmid present in nonadherent E. coli HB101, ECP rendered this organism highly adherent at levels comparable to those of HB101 expressing the BFP. Purified ECP bound in a dose-dependent manner to epithelial cells, and the binding was blocked with anti-ECP antibodies, confirming that the pili possess adhesin properties. An ECP mutant showed only a modest reduction in adherence to cultured cells due to background expression levels of BFP and intimin. However, isogenic mutants not expressing EspA or BFP were significantly less adherent when the ecpA gene was also deleted. Furthermore, a ΔespA ΔecpA double mutant (unable to translocate Tir and to establish intimate adhesion) was at least 10-fold less adherent than the ΔespA and ΔecpA single mutants, even in the presence of BFP. A Δbfp ΔespA ΔecpA triple mutant showed the least adherence compared to the wild type and all the isogenic mutant strains tested, suggesting that ECP plays a synergistic role in adherence. Our data indicate that ECP is an accessory factor that, in association with BFP and other adhesins, contributes to the multifactorial complex interaction of EPEC with host epithelial cells.

scholarly journals Lactobacillus casei Zhang Counteracts Blood-Milk Barrier Disruption and Moderates the Inflammatory Response in Escherichia coli-Induced Mastitis

2021 ◽  
Vol 12 ◽  
Author(s):  
Yuhui Zheng ◽  
Gang Liu ◽  
Wei Wang ◽  
Yajing Wang ◽  
Zhijun Cao ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Inflammatory Response ◽  
Tight Junction ◽  
Lactobacillus Casei ◽  
Mammary Tissue ◽  
Tight Junction Proteins ◽  
E Coli ◽  
Tnf Α ◽  
Injury Models ◽  
Junction Proteins

Escherichia coli is a common mastitis-causing pathogen that can disrupt the blood-milk barrier of mammals. Although Lactobacillus casei Zhang (LCZ) can alleviate mice mastitis, whether it has a prophylactic effect on E. coli-induced mastitis through intramammary infusion, as well as its underlying mechanism, remains unclear. In this study, E. coli-induced injury models of bovine mammary epithelial cells (BMECs) and mice in lactation were used to fill this research gap. In vitro tests of BMECs revealed that LCZ significantly inhibited the E. coli adhesion (p < 0.01); reduced the cell desmosome damage; increased the expression of the tight junction proteins claudin-1, claudin-4, occludin, and zonula occludens-1 (ZO-1; p < 0.01); and decreased the expression of the inflammatory cytokines tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 (p < 0.01), thereby increasing trans-epithelial electric resistance (p < 0.01) and attenuating the lactate dehydrogenase release induced by E. coli (p < 0.01). In vivo tests indicated that LCZ significantly reduced the injury and histological score of mice mammary tissues in E. coli-induced mastitis (p < 0.01) by significantly promoting the expression of the tight junction proteins claudin-3, occludin, and ZO-1 (p < 0.01), which ameliorated blood-milk barrier disruption, and decreasing the expression of the inflammatory cytokines (TNF-α, IL-1β, and IL-6) in mice mammary tissue (p < 0.01). Our study suggested that LCZ counteracted the disrupted blood-milk barrier and moderated the inflammatory response in E. coli-induced injury models, indicating that LCZ can ameliorate the injury of mammary tissue in mastitis.

scholarly journals Secretion of proinflammatory cytokines by normal human melanocytes in response to lipopolysaccharide.

2011 ◽  
Vol 58 (4) ◽  
Author(s):  
Irena Tam ◽  
Krystyna Stępień
Keyword(s):  
Immune System ◽  
Inflammatory Cytokines ◽  
Proinflammatory Cytokines ◽  
Large Body ◽  
Dependent Manner ◽  
Skin Immune System ◽  
Human Melanocytes ◽  
Tnf Α ◽  
Normal Human ◽  
Dose Dependent

A large body of evidence suggests that epidermal melanocytes are an integral part of the skin immune system and can be considered immunocompetent cells. Recently, it has been reported that human melanocytes constitutively express Toll-like receptors and may be involved in the induction of several inflammatory cytokines. In the study the secretion of IL-1β, IL-6 and TNF-α by cultured normal melanocytes was investigated after stimulation with lipopolysaccharide. LPS increased the secretion of IL-1β in a dose-dependent manner. IL-1β stimulated release of IL-6 and TNF-α by melanocytes, whereas LPS activated production of TNF-α, but not of IL-6. These observations indicate that LPS can participate in the regulation of cytokine activity in normal human melanocytes and suggest that cytokines released by melanocytes could affect melanocytes themselves or/and other cells of the epidermis.

scholarly journals The Antimalarial Artemisinin Synergizes with Antibiotics To Protect against Lethal Live Escherichia coli Challenge by Decreasing Proinflammatory Cytokine Release

2006 ◽  
Vol 50 (7) ◽  
pp. 2420-2427 ◽  
Author(s):  
Jun Wang ◽  
Hong Zhou ◽  
Jiang Zheng ◽  
Juan Cheng ◽  
Wei Liu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Sensor Technology ◽  
Dependent Manner ◽  
Cpg Odn ◽  
Lethal Challenge ◽  
Necrosis Factor Alpha ◽  
E Coli ◽  
Tlr4 Mrna ◽  
Tnf Α ◽  
Anti Inflammatory

ABSTRACT In the present study artemisinin (ART) was found to have potent anti-inflammatory effects in animal models of sepsis induced by CpG-containing oligodeoxy-nucleotides (CpG ODN), lipopolysaccharide (LPS), heat-killed Escherichia coli 35218 or live E. coli. Furthermore, we found that ART protected mice from a lethal challenge by CpG ODN, LPS, or heat-killed E. coli in a dose-dependent manner and that the protection was related to a reduction in serum tumor necrosis factor alpha (TNF-α). More significantly, the administration of ART together with ampicillin or unasyn (a complex of ampicillin and sulbactam) decreased mortality from 100 to 66.7% or 33.3%, respectively, in mice subjected to a lethal live E. coli challenge. Together with the observation that ART alone does not inhibit bacterial growth, this result suggests that ART protection is achieved as a result of its anti-inflammatory activity rather than an antimicrobial effect. In RAW264.7 cells, pretreatment with ART potently inhibited TNF-α and interleukin-6 release induced by CpG ODN, LPS, or heat-killed E. coli in a dose- and time-dependent manner. Experiments utilizing affinity sensor technology revealed no direct binding of ART with CpG ODN or LPS. Flow cytometry further showed that ART did not alter binding of CpG ODN to cell surfaces or the internalization of CpG ODN. In addition, upregulated levels of TLR9 and TLR4 mRNA were not attenuated by ART treatment. ART treatment did, however, block the NF-κB activation induced by CpG ODN, LPS, or heat-killed E. coli. These findings provide compelling evidence that ART may be an important potential drug for sepsis treatment.

scholarly journals Inhibition of Escherichia coli-Induced Meningitis by Carboxyfullerence

1999 ◽  
Vol 43 (9) ◽  
pp. 2273-2277 ◽  
Author(s):  
Nina Tsao ◽  
Puthuparampil P. Kanakamma ◽  
Tien-Yau Luh ◽  
Chen-Kung Chou ◽  
Huan-Yao Lei
Keyword(s):  
Escherichia Coli ◽  
Water Soluble ◽  
Dependent Manner ◽  
Neutrophilic Infiltration ◽  
Necrosis Factor Alpha ◽  
E Coli ◽  
Brain Barrier Permeability ◽  
Factor Alpha ◽  
Dose Dependent ◽  
Necrosis Factor

ABSTRACT The effect of a water-soluble malonic acid derivative of carboxyfullerence (C60) against Escherichia coli-induced meningitis was tested. C60 can protect the mice from E. coli-induced death in a dose-dependent manner. C60 administered intraperitoneally as late as 9 h after E. coliinjection was still protective. The C60-treated mice had less tumor necrosis factor alpha and interleukin-1β production by staining of brain tissue compared to the levels of production for nontreated mice. The E. coli-induced increases in blood-brain barrier permeability and inflammatory neutrophilic infiltration were also inhibited. These data suggest that C60 is a potentially therapeutic agent for bacterial meningitis.

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