Activation Of Host Defense Mechanisms Research Articles

β 1-4 mannobiose enhances Salmonella-killing activity and activates innate immune responses in chicken macrophages

Salmonella spp. is one of the major causes of food-borne illness in humans, and Salmonella enteritidis (SE) infection in commercial poultry is a world-wide problem. Here we have investigated the in vitro immune-modulating effects of β 1-4 mannobiose (MNB), which was previously found to prevent SE infection in vivo in chickens, using chicken macrophage (MQ-MCSU) cells. Treatment of MQ-NCSU cells with MNB dose-dependently increased both phagocytic activity and Salmonella-killing activity of macrophages, with the highest reduction in SE viability observed at a concentration of 40 μg/ml at 48 h post-infection. Likewise, both hydrogen peroxide (H 2O 2) and nitric oxide (NO) production were increased in a dose-dependent manner by MNB. Gene expression analysis of MNB-treated macrophages revealed significant increases in the expression of iNOS, NOX-1, IFN-γ, NRAMP1, and LITAF, genes critical for host defense and antimicrobial activity, when compared to untreated cells. This data confirms that MNB possesses potent innate immune-modulating activities and can up-regulate antibacterial defenses in chicken macrophages.

Short Communication: Quantitative Proteomic Plasma Profiling Reveals Activation of Host Defense to Oxidative Stress in Chronic SIV and Methamphetamine Comorbidity

The double epidemic of substance abuse and HIV infection is a multifaceted problem To investigate mechanistic clues to the effects of substance abuse on infected individuals we preformed quantitative proteomic profiling of plasma in a methamphetamine treated nonhuman primate model for AIDS. A nontargeted quantitative approach identified extracellular superoxide dismutase to be significantly upregulated by SIV and methamphetamine treatment, and targeted studies revealed an increase in expression in the antioxidant glutathione S-transferase, thus pointing to a compensatory response to increased oxidative stress in methamphetamine-treated animals.

Inheritance of hyper sensitive response to Striga parasitism in sorghum [Sorghum bicolor (L.) Moench

Striga parasitism involves a series of events occurring beneath the soil surface. The complex nature of host/parasite interactions and the influence of environmental factors have limited our knowledge of host plant resistant mechanisms and their genetics and slowed the breeding for resistance efforts. However, the development of the new laboratory screening assays in our laboratory, have improved our understanding of host parasite interactions. Using these assays, two sorghum lines, CK32 and KP33, that posses a strong HR response, a potent active host defense response that discourage Striga parasitic establishment, were identified from progenies derived from a wild sorghum line, P47121. The aim of this study was to elucidate the inheritance of the HR response, using the Extended Agar Gel Assay. CK32 and KP33 were crossed to two sorghum cultivars, TX430 and TX2737, which possess no HR response. F1, F2, and BC1 populations were generated from each cross. To evaluate the HR response, parents, F1, F2, BC1 seedlings were infected with Striga and the roots of each seedling were examined for Striga attachment, necrosis, parasitic penetration, or discouragement. CK32, KP33 and their F1 seedling all developed necrosis with about 80% of attached Striga seedlings discouraged from establishment, whereas TX430 and TX 2737 seedlings had no necrosis around the infection areas and the attached Striga plants were penetrated and fully developed. The F2 populations from all crosses fit a segregation ratio for 15:1 (HR: no HR) response. Backcross populations segregated 3:1 (HR: no HR). The results of this segregation suggest that the HR to Striga infection in sorghum is conditioned by two nuclear genes with dominant gene action. The gene symbolsHrs1 and Hrs2 (hypersensitive response to Striga) were proposed for this trait. Key word: Striga, sorghum, parasitism, hypersensitive response, host parasite interactions.

Moxibustion activates host defense against herpes simplex virus type I through augmentation of cytokine production

Moxibustion is a technique used in traditional oriental medicine, the aim of which is to cure and/or prevent illness by activating a person's ability for self-healing. In this study, we assessed how moxibustion would affect the immune system and whether it would augment protective immunity. Mice were treated with moxibustion at Zusanli (ST36) acupoints; we analyzed mortality and cytokine activity in sera after infection with herpes simplex virus type 1 (HSV-1), and cytokine gene expression in the skin and the spleen without a virus challenge. Our study demonstrates that pretreatment of BALB/c mice with moxibustion resulted in a marked increase in the survival rate after infection with lethal doses of HSV-1, and elevated serum levels of IL-1β and IFN-γ on days 1 and 6 post-infection with HSV-1. Semi-quantitative RT-PCR assay showed that moxibustion treatment augmented the expression of IL-1α, IL-1β, IL-6, universal-IFN-α, MIP-1α, and TNF-α mRNA in the skin, and IL-1α, IL-1β, IL-12p40, IL-15, u-IFN-α, MIP-1α, and TNF-α mRNA in the spleen. Moreover, moxibustion induces augmentation of natural killer cell activity. Collectively, our study demonstrates that moxibustion activates protective responses against HSV-1 infection through the activation of cytokine production including IFN, and of NK cells.

Behavioural fever in infected honeybees: parasitic manipulation or coincidental benefit?

Infection by a parasite often induces behavioural changes in the host and these changes may benefit either the host or the parasite. However, whether these changes are active host defence mechanisms or parasitic manipulations or simply incidental byproducts of the infection is not always clear. It has been suggested that understanding the proximate mechanisms of these changes as well as comparative studies could help distinguish these alternatives better. Behavioural fever is a common response to an infection in many animals and we investigated the phenomenon in the novel host-parasite relationship between the honeybee and the temperature-sensitive microsporidian Nosema ceranae. Our results show that infected bees prefer higher temperatures and even though this seems to benefit the pathogen, the proximate mechanism underlying this change is the pathological stress underlying the infection. Especially because it is a new host-parasite relationship, it is best to label the observed behavioural change as a case of incidental benefit although this does not rule out selection acting on it. We discuss the importance of looking at the behavioural outcomes of host-parasite relationships and the importance of studying them at multiple levels for understanding their origin and maintenance.

Altered Linkage of Hydroxyacyl Chains in Lipid A of Campylobacter jejuni Reduces TLR4 Activation and Antimicrobial Resistance

Modification of the lipid A moiety of bacterial lipopolysaccharide influences cell wall properties, endotoxic activity, and bacterial resistance to antimicrobial peptides. Known modifications are variation in the number or length of acyl chains and/or attached phosphoryl groups. Here we identified two genes (gnnA and gnnB) in the major foodborne pathogen Campylobacter jejuni that enable the synthesis of a GlcN3N precursor UDP 2-acetamido-3-amino-2,3-dideoxy-alpha-D-glucopyranose (UDP-GlcNAc3N) in the lipid A backbone. Mass spectrometry of purified lipooligosaccharide verified that the gene products facilitate the formation of a 2,3-diamino-2,3-dideoxy-D-glucose (GlcN3N) disaccharide lipid A backbone when compared with the beta-1'-6-linked D-glucosamine (GlcN) disaccharide observed in Escherichia coli lipid A. Functional assays showed that inactivation of the gnnA or gnnB gene enhanced the TLR4-MD2-mediated NF-kappaB activation. The mutants also displayed increased susceptibility to killing by the antimicrobial peptides polymyxin B, colistin and the chicken cathelicidin-1. The gnnA and gnnB genes are organized in one operon with hemH, encoding a ferrochelatase catalyzing the last step in heme biosynthesis. These results indicate that lipid A modification resulting in amide-linked acyl chains in the lipid A is an effective mechanism to evade activation of the innate host defense and killing by antimicrobial peptides.

Candida glabrata tryptophan‐based pigment production via the Ehrlich pathway

Pigments contribute to the pathogenicity of many fungi, mainly by protecting fungal cells from host defence activities. Here, we have dissected the biosynthetic pathway of a tryptophan-derived pigment of the human pathogen Candida glabrata, identified key genes involved in pigment production and have begun to elucidate the possible biological function of the pigment. Using transcriptional analyses and a transposon insertion library, we have identified genes associated with pigment production. Targeted deletion mutants revealed that the pigment is a by-product of the Ehrlich pathway of tryptophan degradation: a mutant lacking a tryptophan-upregulated aromatic aminotransferase (Aro8) displayed significantly reduced pigmentation and a recombinantly expressed version of this protein was sufficient for pigment production in vitro. Pigment production is tightly regulated as the synthesis is affected by the presence of alternative nitrogen sources, carbon sources, cyclic AMP and oxygen. Growth of C. glabrata on pigment inducing medium leads to an increased resistance to hydrogen peroxide, an effect which was not observed with a mutant defective in pigmentation. Furthermore, pigmented yeast cells had a higher survival rate when exposed to human neutrophils and caused increased damage in a monolayer model of human epithelia, indicating a possible role of pigmentation during interactions with host cells.

Impact of Interleukin-17 on Macrophage Phagocytosis of Apoptotic Neutrophils and Particles

There is now substantial evidence that the cytokine interleukin-17 orchestrates the accumulation of neutrophils in mammals and thereby contributes to host defense. However, the role of IL-17 in controlling neutrophil turnover is not fully understood. Here, we demonstrate that IL-17 stimulates the apoptosis of mouse neutrophils and, simultaneously, the release of the microbicidal compound, myeloperoxidase. IL-17 also stimulates mouse macrophages to phagocytose aged neutrophils and latex beads, and it induces an increase in a soluble form of the phagocytic receptor, lectin-like oxidized low-density lipoprotein receptor-1 as well. In contrast, IL-17 does not markedly increase the release of the archetype neutrophil-recruiting cytokine, macrophage inflammatory protein-2 in mouse macrophages. Importantly, IL-17 also stimulates the phagocytosis of latex beads in human monocyte-derived macrophages. Thus, IL-17 bears the potential to control both phagocytosis and neutrophil turnover during activation of host defense.

IL-1β Processing in Host Defense: Beyond the Inflammasomes

Stimulation and release of proinflammatory cytokines is an essential step for the activation of an effective innate host defense, and subsequently for the modulation of adaptive immune responses. Interleukin-1β (IL-1β) and IL-18 are important proinflammatory cytokines that on the one hand activate monocytes, macropages, and neutrophils, and on the other hand induce Th1 and Th17 adaptive cellular responses. They are secreted as inactive precursors, and the processing of pro-IL-1β and pro-IL-18 depends on cleavage by proteases. One of the most important of these enzymes is caspase-1, which in turn is activated by several protein platforms called the inflammasomes. Inflammasome activation differs in various cell types, and knock-out mice defective in either caspase-1 or inflammasome components have an increased susceptibility to several types of infections. However, in other infections and in models of sterile inflammation, caspase-1 seems to be less important, and alternative mechanisms such as neutrophil-derived serine proteases or proteases released from microbial pathogens can process and activate IL-1β. In conclusion, IL-1β/IL-18 processing during infection is a complex process in which the inflammasomes are only one of several activation mechanisms.

Pathogenesis of Steroid-Resistant Airway Hyperresponsiveness: Interaction between IFN-γ and TLR4/MyD88 Pathways

Chronic inflammation and airway hyperresponsiveness (AHR) in asthmatics are usually managed effectively by treatment with glucocorticoids. However, a subset of patients remains refractory to therapy. The underlying mechanisms are largely unknown, although recruitment of neutrophils (rather than eosinophils) is strongly correlated, suggesting a role for nonallergic host defense responses. Increased levels of IFN-gamma and endotoxins are also related to severe asthma and because these activate host defense pathways, we investigated a possible common etiologic link to steroid-resistant disease. To be able to unravel the complexity of asthmatic inflammation, we used two model systems which permitted dissection of the relevant molecular pathways. In the first of these, we transferred transgenic OVA(323-339) peptide-specific IFN-gamma-producing Th1 cells into mice. These animals were subsequently challenge via the airways with OVA(323-339) peptide and/or LPS. Challenge with both components, but not with either one individually, induced AHR. Importantly, AHR was resistant to treatment with dexamethasone. Development of AHR was dependent on IFN-gamma, inhibited by depletion of pulmonary macrophages (but not neutrophils) and abrogated in TLR4- or MyD88-deficient mice. In contrast, in the second model in which we transferred OVA(323-339) peptide-activated Th2 cells, eosinophilic inflammation and AHR were induced, and both were suppressed by steroid treatment. We conclude that cooperative signaling between IFN-gamma and TLR4/MyD88 constitutes a previously unrecognized pathway that regulates macrophage-dependent steroid-resistant AHR.

C-type lectins and phagocytosis

To recognise and respond to pathogens, germ-line encoded pattern recognition receptors (PRRs) bind to conserved microbial structures and activate host defence systems, including microbial uptake by phagocytosis. Phagocytosis is a complex process that is instrumental in the control of extracellular pathogens, and this activity is mediated by several PRRs, including a number of C-type lectins. While some of these receptors have clearly been shown to mediate or regulate the uptake of pathogens, others are more contentious and are less well understood in terms of their phagocytic potential. Furthermore, very little is known about the underlying phagocytic mechanisms. Here, we review the phagocytic roles of the mannose receptor, Dectin-1, dendritic cell-specific ICAM grabbing non-integrin (DC-SIGN), DCL-1, mannose binding lectin and surfactant proteins A and D.

Tomato Transcriptional Responses to a Foliar and a Vascular Fungal Pathogen Are Distinct

Plant activation of host defense against pathogenic microbes requires significant host transcriptional reprogramming. In this study, we compared transcriptional changes in tomato during compatible and incompatible interactions with the foliar fungal pathogen Cladosporium fulvum and the vascular fungal pathogen Verticillium dahliae. Although both pathogens colonize different host tissues, they display distinct commonalities in their infection strategy; both pathogens penetrate natural openings and grow strictly extracellular. Furthermore, resistance against both pathogens is conveyed by the same class of resistance proteins, the receptor-like proteins. For each individual pathogen, the expression profile of the compatible and incompatible interaction largely overlaps. However, when comparing between the two pathogens, the C. fulvum-induced transcriptional changes show little overlap with those induced by V. dahliae. Moreover, within the subset of genes that are regulated by both pathogens, many genes show inverse regulation. With pathway reconstruction, networks of tomato genes implicated in photorespiration, hypoxia, and glycoxylate metabolism were identified that are repressed upon infection with C. fulvum and induced by V. dahliae. Similarly, auxin signaling is differentially affected by the two pathogens. Thus, differentially regulated pathways were identified with novel strategies that allowed the use of state-of-the-art tools, even though tomato is not a genetic model organism.

Drosophila Intestinal Response to Bacterial Infection: Activation of Host Defense and Stem Cell Proliferation

Although Drosophila systemic immunity is extensively studied, little is known about the fly's intestine-specific responses to bacterial infection. Global gene expression analysis of Drosophila intestinal tissue to oral infection with the Gram-negative bacterium Erwinia carotovora revealed that immune responses in the gut are regulated by the Imd and JAK-STAT pathways, but not the Toll pathway. Ingestion of bacteria had a dramatic impact on the physiology of the gut that included modulation of stress response and increased stem cell proliferation and epithelial renewal. Our data suggest that gut homeostasis is maintained through a balance between cell damage due to the collateral effects of bacteria killing and epithelial repair by stem cell division. The Drosophila gut provides a powerful model to study the integration of stress and immunity with pathways associated with stem cell control, and this study should prove to be a useful resource for such further studies.

Host defence mechanisms against bacterial aggression in periodontal disease: basic mechanisms

Periodontal diseases are complex bacteria-induced infections characterised by an inflammatory host response to plaque microbiota and their by-products. Most of these microorganisms have virulence factors capable of causing massive tissue destruction both directly, through tissue invasion and the production of harmful substances, or indirectly, by activation of host defense mechanisms, creating an inflammatory infiltrate of potent catabolic activity that can interfere with normal host defense mechanisms. In response to the aggression, host defense mechanisms activate innate and adaptive immune responses. Our aim is to offer a general overview of the main mechanisms involved in the host response to bacterial aggression in periodontitis, such as lipopolysaccharide receptor CD14, complement system, polymorphonuclear neutrophils, antibodies and immunoglobulins.

Multiple components contribute to ability of saliva to inhibit influenza viruses

Saliva is a potentially important barrier against respiratory viral infection but its mechanism of action is not well studied. We tested the antiviral activities of whole saliva, specific salivary gland secretions, and purified salivary proteins against strains of influenza A virus (IAV) in vitro. Whole saliva or parotid or submandibular/sublingual secretions from healthy donors inhibited IAV based on hemagglutination inhibition and neutralization assays. This differs from human immunodeficiency virus (HIV), for which only submandibular/sublingual secretions are reported to be inhibitory. Among purified salivary proteins, MUC5B, scavenger receptor cysteine-rich glycoprotein 340 (salivary gp-340), histatins, and human neutrophil defensins (HNPs) inhibited IAV at the concentrations present in whole saliva. In contrast, some abundant salivary proteins (acidic proline-rich proteins and amylase) had no activity, nor did several other less abundant salivary proteins with known activity against HIV (e.g. thrombospondin or serum leukocyte protease inhibitor). Whole saliva and MUC5B did not inhibit neuraminidase activity of IAV and viral neutralizing and aggregating activity of MUC5B was potentiated by the neuraminidase inhibitor oseltamivir. Hence, MUC5B inhibits IAV by presenting a sialic acid ligand for the viral hemagglutinin. The mechanism of action of histatins requires further study. These findings indicate that saliva represents an important initial barrier to IAV infection and underline the complexity of host defense activity of oral secretions. Of interest, antiviral activity of saliva against IAV and HIV differs in terms of specific glandular secretions and proteins that are inhibitory.

The Janus face of Bartonella quintana recognition by Toll-like receptors (TLRs): a review.

Bartonella quintana (B. quintana) is a facultative, intracellular bacterium, which causes trench fever, chronic bacteraemia and bacillary angiomatosis. Little is known about the recognition of B. quintana by the innate immune system. In this review, we address the impact of Toll-like receptors (TLRs) on the recognition of B. quintana and the activation of the host defense. When experimental models using human mononuclear cells, transfected CHO cells, or TLR2-/- and TLR4-/- mice were used, differential effects of TLR2 and TLR4 have been observed. B. quintana micro-organisms stimulated cytokine production through TLR2-mediated signals, whereas no role for TLR4 in the recognition of this pathogen was observed. When single, water-phenol extraction was performed, B. quintana LPS, stimulated cytokine production in a TLR2-dependent manner. However, when double extraction was performed in order to generate highly purified LPS, B. quintana LPS entirely lost its capacity to stimulate cytokines, demonstrating that non-LPS components of B. quintana are responsible for the recognition through TLR2. Moreover, B. quintana LPS was shown to be a potent antagonist of Toll-like receptor 4 (TLR4). In conclusion, B. quintana is an inducer of cytokines through TLR2-, but not TLR4-, dependent mechanisms. This stimulation is induced by bacterial components other than lipopolysaccharide. B. quintana LPS is a naturally occurring antagonist of Toll-like receptor 4 (TLR4). In view of the role played by TLR4 in inflammation, B. quintana LPS may be useful as an anti-TLR4 agent with therapeutic potential in both infections and autoimmune inflammation.

Divergent roles of endothelial NF-κB in multiple organ injury and bacterial clearance in mouse models of sepsis

To define the roles of endothelial-intrinsic nuclear factor κB (NF-κB) activity in host defense and multiple organ injury in response to sepsis, we generated double transgenic (TG) mice (EC-rtTA/I-κBαmt) that conditionally overexpress a degradation-resistant form of the NF-κB inhibitor I-κBα (I-κBαmt) selectively on vascular endothelium. The EC-rtTA/I-κBαmt mice had no basal, but a relatively high level of doxycycline-inducible, I-κBαmt expression. I-κBαmt expression was detected in endothelial cells, but not in fibroblasts, macrophages, and whole blood cells, confirming that transgene expression was restricted to the endothelium. When subjected to endotoxemia, EC-rtTA/I-κBαmt mice showed endothelial-selective blockade of NF-κB activation, repressed expression of multiple endothelial adhesion molecules, reduced neutrophil infiltration into multiple organs, decreased endothelial permeability, ameliorated multiple organ injury, reduced systemic hypotension, and abrogated intravascular coagulation. When subjected to cecal ligation and puncture–induced sepsis, the TG mice had less severe multiple organ injury and improved survival compared with wild-type (WT) mice. WT and EC-rtTA/I-κBαmt mice had comparable capacity to clear three different pathogenic bacteria. Our data demonstrate that endothelial NF-κB activity is an essential mediator of septic multiple organ inflammation and injury but plays little role in the host defense response to eradicate invading pathogenic bacteria.

Drosomycin-Like Defensin, a Human Homologue ofDrosophila melanogasterDrosomycin with Antifungal Activity

Innate antifungal defense in Drosophila melanogaster relies on the activation of the Toll molecule and the release of drosomycin, a defensin-like molecule with antifungal properties. Ten human homologues of Toll have been described, with central roles in activation of the innate host defense. In the present study, we report a putative human homologue of the Drosophila-derived drosomycin, designated drosomycin-like defensin (DLD). Synthetic DLD displays a broad spectrum of activity against Aspergillus spp. and other clinically relevant filamentous fungi. These effects are specific for filamentous fungi; no activity has been found against yeasts or gram-positive or gram-negative bacteria. Synthetic DLD also displays immunomodulatory effects on Aspergillus-stimulated cytokine production. In addition, we show the expression of DLD mRNA in several human tissues, particularly in the skin, consistent with its putative role as a defensin against invading microorganisms. This is the first indication of an endogenous human peptide with specific antifungal activity, which is probably central in the defense against infections with molds.

Signaling by Toll-like Receptors 8 and 9 Requires Bruton's Tyrosine Kinase

Toll-like receptors (TLRs) are a primary surveillance system for the detection of pathogens and are crucial to the activation of host defense. TLR7 and TLR8 sense single-stranded RNA from viruses or host ribonucleoproteins and synthetic imidazoquinolines such as R848, whereas TLR9 senses unmethylated CpG motifs in viral and bacterial DNA and in host DNA. Here we report the endogenous interaction between Brutons's tyrosine kinase (Btk) and human TLR8 and TLR9 in the monocytic cell line THP1. We also show that R848, single-stranded RNA, and CpGB-DNA activate Btk in THP1 cells as shown by phosphorylation of the tyrosine 223 residue of Btk and also by increased autokinase activity. We demonstrate that Btk is required for NFkappaB activation, participating in the pathway to increased phosphorylation of p65 on serine 536 activated by TLR8 and TLR9. Finally we demonstrate that peripheral blood mononuclear cells from patients with X-linked agammaglobulinaemia (XLA) that have dysfunctional Btk are impaired in the induction of interleukin-6 by CpGB-DNA. This study therefore establishes Btk as a key signaling molecule that interacts with and acts downstream of TLR8 and TLR9. Lack of functioning Btk in XLA patients downstream of TLR8 and TLR9 might explain the susceptibility of XLA patients to viral infections.

Cloning, expression and characterization of ferret CXCL10

Chemokines and their receptors function in the recruitment and activation of cells of the immune system to sites of inflammation. As such, chemokines play an important role in mediating pathophysiological events during microbial infection. In particular, CXCL9, CXCL10 and CXCL11 and their cognate receptor CXCR3 have been associated with the clinical course of several infectious diseases, including severe acute respiratory syndrome (SARS) and influenza. While CXCL9, CXCL10 and CXCL11 share the same receptor and have overlapping functions, each can also have unique activity in host defense. The lack of a preferred characterized animal model for SARS has brought our attention to ferrets, which have been used for years in influenza studies. The lack of immunological reagents for ferrets prompted us to clone CXCL9, CXCL10, CXCL11 and CXCR3 and, in the case of CXCL10, to express the gene as a recombinant protein. In this study we demonstrate that endogenous ferret CXCL10 exhibits similar mRNA expression patterns in the lungs of deceased SARS patients and ferrets experimentally infected with SARS coronavirus. This study therefore represents an important step towards development of the ferret as a model for the role of CXCL9, CXCL10 and CXCL11:CXCR3 axis in severe viral infections.