Activation Of Host Defense Mechanisms Research Articles

Modulating the Cellular Immune Response of Oligonucleotides by Brush Polymer-Assisted Compaction.

Unwanted stimulation of the innate immune system by foreign nucleic acids has been one of the major barriers preventing bioactive sequences from reaching market. Foreign nucleic acids can be recognized by multiple pattern recognition receptors (PRRs), which trigger a signaling cascade to activate host defense systems, leading to a range of side effects. This study demonstrates that polyethylene glycol (PEG)-modified DNA strands can greatly reduce the activation of the innate immune system, and the extent of reduction is dependent upon polymer architecture. Highly branched brushes with long PEG side chains achieve the best suppression by blocking PRR interactions via a local steric effect. Interestingly, the brush polymer creates little barrier toward DNA-DNA interaction. Quantification of inflammatory cytokines in both mRNA and protein levels as well as the extent of cellular uptake shows a direct correlation between steric congestion and reduction of cellular immune response. These results suggest that the brush architecture offers unique advantages for PEGylating oligonucleotides in the context of minimizing unwanted immune system activation.

Immunostimulatory effect of aqueous extract of Coriandrum sativum L. seed on macrophages.

Coriandrum sativum L. seed is generally used as a spice and crude drug. Although many functions of the various components in C. sativum L. seed have been reported, the immunostimulatory effect of water-soluble components in C. sativum L. seed has not been studied. In the present study, we focused on the immunostimulatory effect of C. sativum L. seed aqueous extract (CAE) on macrophages as a novel health function of C. sativum L. seed components. CAE significantly enhanced the production of tumor necrosis factor (TNF)-α and interleukin (IL)-6 in both RAW264.7 cells and peritoneal macrophages by enhancing the expression levels of these cytokine genes. CAE also stimulated nitric oxide (NO) production and the phagocytosis activity in RAW264.7 cells. We suggest that the activity of CAE is a result of the upregulation of mitogen-activated protein kinase and nuclear factor-κB cascades via TLR4. In addition, IL-6 production by peritoneal macrophages collected from CAE-administered mice was significantly enhanced, suggesting that CAE could stimulate macrophage activity in vivo. The findings of the present study suggest that CAE contains a novel water-soluble component with an immunostimulatory effect on macrophages. CAE would contribute to activating host defense against pathogens by stimulating the innate immunity. © 2017 Society of Chemical Industry.

Single-step purification of cyclotides using affinity chromatography.

Cyclotides are considered promising scaffolds for drug development owing to their inherent host defence activities and highly stable structure, defined by the cyclic cystine knot. These proteins are expressed as complex mixtures in plants. Although several methods have been developed for their isolation and analysis, purification of cyclotides is still a lengthy process. Here, we describe the use of affinity chromatography for the purification of cyclotides using polyclonal IgG antibodies raised in rabbits against cycloviolacin O2 and immobilized on NHS-activated Sepharose columns. Cycloviolacin O2 was used as a model substance to evaluate the chromatographic principle, first as a pure compound and then in combination with other cyclotides, that is, bracelet cyclotide cycloviolacin O19 and Möbius cyclotide kalata B1, and in a plant extract. We demonstrate that single-step purification of cyclotides by affinity chromatography is possible but cross reactivity may occur between homologue cyclotides of the bracelet subfamily.

Contribution of leukocytes to the induction and resolution of the acute inflammatory response in chickens

A successful immune response against invading pathogens relies on the efficient activation of host defense mechanisms and a timely return to immune homeostasis. Despite their importance, these mechanisms remain ill-defined in most animal groups. This study focuses on the acute inflammatory response of chickens, important both as an avian model with a unique position in evolution as well as an increasingly notable target of infectious zoonotic diseases. We took advantage of an in vivo self-resolving intra-abdominal challenge model to provide an integrative view of leukocyte responses during the induction and resolution phases of acute inflammation. Our results showed rapid leukocyte infiltration into the abdominal cavity post zymosan challenge (significant increase as early as 4 h), which was dominated by heterophils. Peak leukocyte infiltration and ROS production reached maximum levels at 12 h post challenge, which was significantly earlier than comparative studies in teleost fish and mice. Both heterophils and monocyte/macrophages contributed to ROS production. Local leukocyte infiltration was preceded by an increase in peripheral leukocytes and a drop in the number of bone marrow leukocytes. The proportion of apoptotic leukocytes increased following peak of acute inflammation, rising to significant levels within the abdominal cavity by 48 h, consistent with other indicators for the resolution of inflammation. Importantly, comparison of chicken phagocytic responses with those previously shown in agnathan, teleost and murine models suggested a progressive evolutionary shift towards an increased sensitivity to pro-inflammatory pathogen-derived particles and decreased sensitivity towards homeostatic stimuli. Thus, while significant conservation can be noted across the immune systems of endotherms, this study highlights additional unique features that govern the induction and resolution of acute inflammation in the avian system, which may be relevant to disease susceptibility and performance.

CNBP acts as a key transcriptional regulator of sustained expression of interleukin-6.

The transcription of inflammatory genes is an essential step in host defense activation. Here, we show that cellular nucleic acid-binding protein (CNBP) acts as a transcription regulator that is required for activating the innate immune response. We identified specific CNBP-binding motifs present in the promoter region of sustained inflammatory cytokines, thus, directly inducing the expression of target genes. In particular, lipopolysaccharide (LPS) induced cnbp expression through an NF-κB-dependent manner and a positive autoregulatory mechanism, which enables prolonged il-6 gene expression. This event depends strictly on LPS-induced CNBP nuclear translocation through phosphorylation-mediated dimerization. Consequently, cnbp-depleted zebrafish are highly susceptible to Shigella flexneri infection in vivo. Collectively, these observations identify CNBP as a key transcriptional regulator required for activating and maintaining the immune response.

Immunostimulatory effect of dried bonito extract on mouse macrophage cell lines and mouse primary peritoneal macrophages.

Dried bonito is a preserved food used in Japan, which contains abundant flavor ingredients and functional substances. We focused on the immunostimulatory effect of dried bonito extract (DBE) on mouse macrophage-like J774.1 cells, RAW264.7 cells, and mouse primary peritoneal macrophages. DBE significantly stimulated the production of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) by both J774.1 cells and peritoneal macrophages by enhancing the cytokine gene expression levels. In addition, DBE stimulated nitric oxide production by enhancing the expression of inducible nitric oxide synthase in RAW264.7 cells. DBE also increased the phagocytosis activity of J774.1 cells. Immunoblot analysis revealed that DBE has an immunostimulatory effect on macrophages through activation of mitogen-activated protein kinase and nuclear factor-κB cascades. TNF-α production enhanced by DBE was partially inhibited by treatment with TLR4 inhibitor TAK-242, whereas IL-6 production enhanced by DBE was almost inhibited. These results suggested that DBE is thought to strongly stimulate the TLR4 signaling pathway for macrophage activation, and its activation is also involved in other signaling. Finally, the phagocytosis activity of peritoneal macrophages from DBE-administered BALB/c mice increased significantly, suggesting that DBE has the potential to stimulate macrophage activity in vivo. In conclusion, these data indicated that DBE contributes to activating host defense against pathogens by activating innate immunity.

A few sequence polymorphisms among isolates of Maize bushy stunt phytoplasma associate with organ proliferation symptoms of infected maize plants.

Background and Aims Maize bushy stunt phytoplasma (MBSP) is a bacterial pathogen of maize (Zea mays L.) across Latin America. MBSP belongs to the 16SrI-B sub-group within the genus ‘Candidatus Phytoplasma’. MBSP and its insect vector Dalbulus maidis (Hemiptera: Cicadellidae) are restricted to maize; both are thought to have coevolved with maize during its domestication from a teosinte-like ancestor. MBSP-infected maize plants show a diversity of symptoms. and it is likely that MBSP is under strong selection for increased virulence and insect transmission on maize hybrids that are widely grown in Brazil. In this study it was investigated whether the differences in genome sequences of MBSP isolates from two maize-growing regions in South-east Brazil explain variations in symptom severity of the MBSP isolates on various maize genotypes. Methods MBSP isolates were collected from maize production fields in Guaíra and Piracicaba in South-east Brazil for infection assays. One representative isolate was chosen for de novo whole-genome assembly and for the alignment of sequence reads from the genomes of other phytoplasma isolates to detect polymorphisms. Statistical methods were applied to investigate the correlation between variations in disease symptoms of infected maize plants and MBSP sequence polymorphisms. Key Results MBSP isolates contributed consistently to organ proliferation symptoms and maize genotype to leaf necrosis, reddening and yellowing of infected maize plants. The symptom differences are associated with polymorphisms in a phase-variable lipoprotein, which is a candidate effector, and an ATP-dependent lipoprotein ABC export protein, whereas no polymorphisms were observed in other candidate effector genes. Lipoproteins and ABC export proteins activate host defence responses, regulate pathogen attachment to host cells and activate effector secretion systems in other pathogens. Conclusions Polymorphisms in two putative virulence genes among MBSP isolates from maize-growing regions in South-east Brazil are associated with variations in organ proliferation symptoms of MBSP-infected maize plants.

Coordinated inhibition of C/EBP by Tribbles in multiple tissues is essential for Caenorhabditis elegans development.

BackgroundTribbles proteins are conserved pseudokinases that function to control kinase signalling and transcription in diverse biological processes. Abnormal function in human Tribbles has been implicated in a number of diseases including leukaemia, metabolic syndromes and cardiovascular diseases. Caenorhabditis elegans Tribbles NIPI-3 was previously shown to activate host defense upon infection by promoting the conserved PMK-1/p38 mitogen-activated protein kinase (MAPK) signalling pathway. Despite the prominent role of Tribbles proteins in many species, our knowledge of their mechanism of action is fragmented, and the in vivo functional relevance of their interactions with other proteins remains largely unknown.ResultsHere, by characterizing nipi-3 null mutants, we show that nipi-3 is essential for larval development and viability. Through analyses of genetic suppressors of nipi-3 null mutant lethality, we show that NIPI-3 negatively controls PMK-1/p38 signalling via transcriptional repression of the C/EBP transcription factor CEBP-1. We identified CEBP-1’s transcriptional targets by ChIP-seq analyses and found them to be enriched in genes involved in development and stress responses. Unlike its cell-autonomous role in innate immunity, NIPI-3 is required in multiple tissues to control organismal development.ConclusionsTogether, our data uncover an unprecedented crosstalk involving multiple tissues, in which NIPI-3 acts as a master regulator to inhibit CEBP-1 and the PMK-1/p38 MAPK pathway. In doing so, it keeps innate immunity in check and ensures proper organismal development.Electronic supplementary materialThe online version of this article (doi:10.1186/s12915-016-0320-z) contains supplementary material, which is available to authorized users.

Obligate Biotroph Pathogens of the Genus Albugo Are Better Adapted to Active Host Defense Compared to Niche Competitors.

Recent research suggested that plants behave differently under combined versus single abiotic and biotic stress conditions in controlled environments. While this work has provided a glimpse into how plants might behave under complex natural conditions, it also highlights the need for field experiments using established model systems. In nature, diverse microbes colonize the phyllosphere of Arabidopsis thaliana, including the obligate biotroph oomycete genus Albugo, causal agent of the common disease white rust. Biotrophic, as well as hemibiotrophic plant pathogens are characterized by efficient suppression of host defense responses. Lab experiments have even shown that Albugo sp. can suppress non-host resistance, thereby enabling otherwise avirulent pathogen growth. We asked how a pathogen that is vitally dependent on a living host can compete in nature for limited niche space while paradoxically enabling colonization of its host plant for competitors? To address this question, we used a proteomics approach to identify differences and similarities between lab and field samples of Albugo sp.-infected and -uninfected A. thaliana plants. We could identify highly similar apoplastic proteomic profiles in both infected and uninfected plants. In wild plants, however, a broad range of defense-related proteins were detected in the apoplast regardless of infection status, while no or low levels of defense-related proteins were detected in lab samples. These results indicate that Albugo sp. do not strongly affect immune responses and leave distinct branches of the immune signaling network intact. To validate our findings and to get mechanistic insights, we tested a panel of A. thaliana mutant plants with induced or compromised immunity for susceptibility to different biotrophic pathogens. Our findings suggest that the biotroph pathogen Albugo selectively interferes with host defense under different environmental and competitive pressures to maintain its ecological niche dominance. Adaptation to host immune responses while maintaining a partially active host immunity seems advantageous against competitors. We suggest a model for future research that considers not only host–microbe but in addition microbe–microbe and microbe–host environment factors.

Metabolites Associated With Circulating Interleukin-6 in Older Adults.

Circulating levels of the pro-inflammatory cytokine interleukin-6 (IL-6) levels are elevated in older adults, but mechanisms are unclear. In the current study, we used an untargeted metabolomic approach to develop an improved understanding about mechanisms related to circulating IL-6 in older adults. Serum IL-6 values were log-transformed to normalize its distribution. Multivariable-adjusted linear regression was used to examine the association between 324 serum metabolites with log IL-6. Backward elimination linear regression was used to develop a metabolite predictor set representative of log IL-6. Thirty-six metabolites were significantly associated (p < 0.05 and q < 0.30) with log IL-6 in 73 older adults (average age, 78 years). Metabolites related to tryptophan metabolism (kynurenine, 3-indoxyl sulfate, indoleacetate, indolepropionate, C-glycosyltryptophan), infectious burden (C-glycosyltryptophan, N6-carbamoylthreonyladenosine, 1-methylurate, N-formylmethionine, N1-methyladenosine, 3-indoxyl sulfate, bilirubin (E,E), indoleacetate, γ-CEHC, N-acetylneuraminate), aryl hydrocarbon receptor activation and cytochrome P450 (CYP) 1A expression (kynurenine, 3-indoxyl sulfate, indoleacetate, N6-carbamoylthreonyladenosine, bilirubin, 1-methylurate) were positively associated, whereas metabolites related to CYP-mediated ω-oxidation (adipate, 8-hydroxyoctanoate, azelate, sebacate, undecanedioate, γ-CEHC), and peroxisome proliferator activated receptor-alpha (PPAR-α) activation (13 + 9-HODE, bilirubin, 5-oxoproline, cholesterol, glycerate, uridine) were negatively associated with log IL-6. The use of backward elimination regression identified tyrosine, cysteine, uridine, bilirubin, N-formylmethionine, indoleacetate, and 3-indoxyl sulfate to collectively explain 51% of the variance inherent in log IL-6. These data suggest roles for tryptophan metabolism, infectious burden, activation of host defense, and detoxification through CYP1A-mediated pathways in mechanisms related to elevated inflammation, whereas CYP-mediated ω-oxidation and PPAR-α activation may be related to decreased inflammation in older adults.

Broad defects in the energy metabolism of leukocytes underlie immunoparalysis in sepsis.

The acute phase of sepsis is characterized by a strong inflammatory reaction. At later stages in some patients, immunoparalysis may be encountered, which is associated with a poor outcome. By transcriptional and metabolic profiling of human patients with sepsis, we found that a shift from oxidative phosphorylation to aerobic glycolysis was an important component of initial activation of host defense. Blocking metabolic pathways with metformin diminished cytokine production and increased mortality in systemic fungal infection in mice. In contrast, in leukocytes rendered tolerant by exposure to lipopolysaccharide or after isolation from patients with sepsis and immunoparalysis, a generalized metabolic defect at the level of both glycolysis and oxidative metabolism was apparent, which was restored after recovery of the patients. Finally, the immunometabolic defects in humans were partially restored by therapy with recombinant interferon-γ, which suggested that metabolic processes might represent a therapeutic target in sepsis.

Editorial: Salicylic Acid Signaling Networks

Editorial: Salicylic Acid Signaling Networks

Activity of Medicinal Plant Extracts on Multiplication of Mycobacterium tuberculosis under Reduced Oxygen Conditions Using Intracellular and Axenic Assays.

Aim. Test the activity of selected medicinal plant extracts on multiplication of Mycobacterium tuberculosis under reduced oxygen concentration which represents nonreplicating conditions. Material and Methods. Acetone, ethanol and aqueous extracts of the plants Acorus calamus L. (rhizome), Ocimum sanctum L. (leaf), Piper nigrum L. (seed), and Pueraria tuberosa DC. (tuber) were tested on Mycobacterium tuberculosis H37Rv intracellularly using an epithelial cell (A549) infection model. The extracts found to be active intracellularly were further studied axenically under reducing oxygen concentrations. Results and Conclusions. Intracellular multiplication was inhibited ≥60% by five of the twelve extracts. Amongst these 5 extracts, in axenic culture, P. nigrum (acetone) was active under aerobic, microaerophilic, and anaerobic conditions indicating presence of multiple components acting at different levels and P. tuberosa (aqueous) showed bactericidal activity under microaerophilic and anaerobic conditions implying the influence of anaerobiosis on its efficacy. P. nigrum (aqueous) and A. calamus (aqueous and ethanol) extracts were not active under axenic conditions but only inhibited intracellular growth of Mycobacterium tuberculosis, suggesting activation of host defense mechanisms to mediate bacterial killing rather than direct bactericidal activity.

Adaptor Protein-3-Mediated Trafficking of TLR2 Ligands Controls Specificity of Inflammatory Responses but Not Adaptor Complex Assembly.

Innate immune engagement results in the activation of host defenses that produce microbe-specific inflammatory responses. A long-standing interest in the field of innate immunity is to understand how varied host responses are generated through the signaling of just a limited number of receptors. Recently, intracellular trafficking and compartmental partitioning have been identified as mechanisms that provide signaling specificity for receptors by regulating signaling platform assembly. We show that cytokine activation as a result of TLR2 stimulation occurs at different intracellular locations and is mediated by the phagosomal trafficking molecule adaptor protein-3 (AP-3). AP-3 is required for trafficking TLR2 purified ligands or the Lyme disease causing bacterium, Borrelia burgdorferi, to LAMP-1 lysosomal compartments. The presence of AP-3 is necessary for the activation of cytokines such as IL-6 but not TNF-α or type I IFNs, suggesting induction of these cytokines occurs from a different compartment. Lack of AP-3 does not interfere with the recruitment of TLR signaling adaptors TRAM and MyD88 to the phagosome, indicating that the TLR-MyD88 signaling complex is assembled at a prelysosomal stage and that IL-6 activation depends on proper localization of signaling molecules downstream of MyD88. Finally, infection of AP-3-deficient mice with B. burgdorferi resulted in altered joint inflammation during murine Lyme arthritis. Our studies further elucidate the effects of phagosomal trafficking on tailoring immune responses in vitro and in vivo.

Osteopontin binds and modulates functions of eosinophil‐recruiting chemokines

Allergic asthma is characterized by eosinophilic inflammation and airway obstruction. There is also an increased risk of pulmonary infection caused by Streptococcus pneumoniae, in particular during severe asthma where high levels of the glycoprotein, osteopontin (OPN), are present in the airways. Eosinophils can be recruited by chemokines activating the receptor CCR3 including eotaxin-1/CCL11, eotaxin-2/CCL24, eotaxin-3/CCL26, RANTES/CCL5, and MEC/CCL28. In addition to inducing chemotaxis, several of these molecules have defensin-like antibacterial properties. This study set out to elucidate the functional consequences of OPN binding to eosinophil-recruiting chemokines. Antibacterial activities of the chemokines were investigated using viable count assays and electron microscopy. Binding studies were performed by means of surface plasmon resonance. The potential interference of OPN with antibacterial, receptor-activating, and lipopolysaccharide-neutralizing abilities of these chemokines was investigated. We found that OPN bound all eosinophil-recruiting chemokines with high affinity except for CCL5. The eosinophil-recruiting chemokines all displayed bactericidal activity against S. pneumoniae, but only CCL26 and CCL28 retained high antibacterial activity in the presence of sodium chloride at physiologic concentrations. Preincubation of the chemokines with OPN strongly inhibited their antibacterial activity against S. pneumoniae but did not affect their ability to activate CCR3. All chemokines investigated showed LPS-neutralizing activity that was impaired by OPN only in the case of CCL24. The data suggest that OPN may impair host defense activities of the chemokines without affecting their eosinophil-recruiting properties. This could be one mechanism explaining the increased vulnerability to acquire pneumococcal infection in parallel with sustained allergic inflammation in asthma.

Antagonistic Interplay between MicroRNA-155 and IL-10 during Lyme Carditis and Arthritis.

MicroRNA-155 has been shown to play a role in immune activation and inflammation, and is suppressed by IL-10, an important anti-inflammatory cytokine. The established involvement of IL-10 in the murine model of Borrelia burgdorferi-induced Lyme arthritis and carditis allowed us to assess the interplay between IL-10 and miR-155 in vivo. As reported previously, Mir155 was highly upregulated in joints from infected severely arthritic B6 Il10-/- mice, but not in mildly arthritic B6 mice. In infected hearts, Mir155 was upregulated in both strains, suggesting a role of miR-155 in Lyme carditis. Using B. burgdorferi-infected B6, Mir155-/-, Il10-/-, and Mir155-/- Il10-/- double-knockout (DKO) mice, we found that anti-inflammatory IL-10 and pro-inflammatory miR-155 have opposite and somewhat compensatory effects on myeloid cell activity, cytokine production, and antibody response. Both IL-10 and miR-155 were required for suppression of Lyme carditis. Infected Mir155-/- mice developed moderate/severe carditis, had higher B. burgdorferi numbers, and had reduced Th1 cytokine expression in hearts. In contrast, while Il10-/- and DKO mice also developed severe carditis, hearts had reduced bacterial numbers and elevated Th1 and innate cytokine expression. Surprisingly, miR-155 had little effect on Lyme arthritis. These results show that antagonistic interplay between IL-10 and miR-155 is required to balance host defense and immune activation in vivo, and this balance is particularly important for suppression of Lyme carditis. These results also highlight tissue-specific differences in Lyme arthritis and carditis pathogenesis, and reveal the importance of IL-10-mediated regulation of miR-155 in maintaining healthy immunity.

The ASK family kinases differentially mediate induction of type I interferon and apoptosis during the antiviral response.

Viral infection activates host defense mechanisms, including the production of type I interferon (IFN) and the apoptosis of infected cells. We investigated whether these two antiviral responses were differentially regulated in infected cells. We showed that the mitogen-activated protein kinase (MAPK) kinase kinase (MAPKKK) apoptosis signal-regulating kinase 1 (ASK1) was activated in cells by the synthetic double-stranded RNA analog polyinosinic:polycytidylic acid [poly(I:C)] and by RNA viruses, and that ASK1 played an essential role in both the induction of the gene encoding IFN-β (IFNB) and apoptotic cell death. In contrast, we found that the MAPKKK ASK2, a modulator of ASK1 signaling, was essential for ASK1-dependent apoptosis, but not for inducing IFNB expression. Furthermore, genetic deletion of either ASK1 or ASK2 in mice promoted the replication of influenza A virus in the lung. These results indicated that ASK1 and ASK2 are components of the antiviral defense mechanism and suggested that ASK2 acts as a key modulator that promotes apoptosis rather than the type I IFN response. Because ASK2 is selectively present in epithelium-rich tissues, such as the lung, ASK2-dependent apoptosis may contribute to an antiviral defense in tissues with a rapid repair rate in which cells could be readily replaced.

A serine protease isolated from the bristles of the Amazonic caterpillar, Premolis semirufa, is a potent complement system activator.

BackgroundThe caterpillar of the moth Premolis semirufa, commonly named pararama, is found in the Brazilian Amazon region. Accidental contact with the caterpillar bristles causes an intense itching sensation, followed by symptoms of an acute inflammation, which last for three to seven days after the first incident. After multiple accidents a chronic inflammatory reaction, called “Pararamose”, characterized by articular synovial membrane thickening with joint deformities common to chronic synovitis, frequently occurs. Although complement mediated inflammation may aid the host defense, inappropriate or excessive activation of the complement system and generation of anaphylatoxins can lead to inflammatory disorder and pathologies. The aim of the present study was to evaluate, in vitro, whether the Premolis semirufa’s bristles extract could interfere with the human complement system.ResultsThe bristles extract was able to inhibit the haemolytic activity of the alternative pathway, as well as the activation of the lectin pathway, but had no effect on the classical pathway, and this inhibition seemed to be caused by activation and consumption of complement components. The extract induced the production of significant amounts of all three anaphylatoxins, C3a, C4a and C5a, promoted direct cleavage of C3, C4 and C5 and induced a significant generation of terminal complement complexes in normal human serum. By using molecular exclusion chromatography, a serine protease of 82 kDa, which activates complement, was isolated from P. semirufa bristles extract. The protease, named here as Ps82, reduced the haemolytic activity of the alternative and classical pathways and inhibited the lectin pathway. In addition, Ps82 induced the cleavage of C3, C4 and C5 and the generation of C3a and C4a in normal human serum and it was capable to cleave human purified C5 and generate C5a. The use of Phenanthroline, metalloprotease inhibitor, in the reactions did not significantly interfere with the activity of the Ps82, whereas the presence of PMSF, serine protease inhibitor, totally blocked the activity.ConclusionThese data show that a serine protease present in the Premolis semirufa’s bristles extract has the ability to activate the complement system, which may contribute to the inflammatory process presented in humans after envenomation.

Genetic Variation in Pattern Recognition Receptors and Adaptor Proteins Associated With Development of Chronic Q Fever.

Q fever is an infection caused by Coxiella burnetii. Persistent infection (chronic Q fever) develops in 1%-5% of patients. We hypothesize that inefficient recognition of C. burnetii and/or activation of host-defense in individuals carrying genetic variants in pattern recognition receptors or adaptors would result in an increased likelihood to develop chronic Q fever. Twenty-four single-nucleotide polymorphisms in genes encoding Toll-like receptors, nucleotide-binding oligomerization domain-like receptor-2, αvβ3 integrin, CR3, and adaptors myeloid differentiation primary response protein 88 (MyD88), and Toll interleukin 1 receptor domain-containing adaptor protein (TIRAP) were genotyped in 139 patients with chronic Q fever and in 220 controls with cardiovascular risk-factors and previous exposure to C. burnetii. Associations between these single-nucleotide polymorphisms and chronic Q fever were assessed by means of univariate logistic regression models. Cytokine production in whole-blood stimulation assays was correlated with relevant genotypes. Polymorphisms in TLR1 (R80T), NOD2 (1007fsX1), and MYD88 (-938C>A) were associated with chronic Q fever. No association was observed for polymorphisms in TLR2, TLR4, TLR6, TLR8, ITGAV, ITGB3, ITGAM, and TIRAP. No correction for multiple testing was performed because only genes with a known role in initial recognition of C. burnetii were included. In the whole-blood assays, individuals carrying the TLR1 80R-allele showed increased interleukin 10 production with C. burnetii exposure. Polymorphisms in TLR1 (R80T), NOD2 (L1007fsX1), and MYD88 (-938C>A) are associated with predisposition to development of chronic Q fever. For TLR1, increased interleukin 10 responses to C. burnetii in individuals carrying the risk allele may contribute to the increased risk of chronic Q fever.

Eotaxin-3 (CCL26) exerts innate host defense activities that are modulated by mast cell proteases.

During bacterial infections of the airways, a Th1-profiled inflammation promotes the production of several host defense proteins and peptides with antibacterial activities including β-defensins, ELR-negative CXC chemokines, and the cathelicidin LL-37. These are downregulated by Th2 cytokines of the allergic response. Instead, the eosinophil-recruiting chemokines eotaxin-1/CCL11, eotaxin-2/CCL24, and eotaxin-3/CCL26 are expressed. This study set out to investigate whether these chemokines could serve as innate host defense molecules during allergic inflammation. Antibacterial activities of the eotaxins were investigated using viable count assays, electron microscopy, and methods assessing bacterial permeabilization. Fragments generated by mast cell proteases were characterized, and their potential antibacterial, receptor-activating, and lipopolysaccharide-neutralizing activities were investigated. CCL11, CCL24, and CCL26 all showed potent bactericidal activity, mediated through membrane disruption, against the airway pathogens Streptococcus pneumoniae, Staphylococcus aureus, Nontypeable Haemophilus influenzae, and Pseudomonas aeruginosa. CCL26 retained bactericidal activity in the presence of salt at physiologic concentrations, and the region holding the highest bactericidal activity was the cationic and amphipathic COOH-terminus. Proteolysis of CCL26 by chymase and tryptase, respectively, released distinct fragments of the COOH- and NH2 -terminal regions. The COOH-terminal fragment retained antibacterial activity while the NH2 -terminal had potent LPS-neutralizing properties in the order of CCL26 full-length protein. An identical fragment to NH2 -terminal fragment generated by tryptase was obtained after incubation with supernatants from activated mast cells. None of the fragments activated the CCR3-receptor. Taken together, the findings show that the eotaxins can contribute to host defense against common airway pathogens and that their activities are modulated by mast cell proteases.