Characterization and gene expression analysis of the two main Th17 cytokines (IL-17A/F and IL-22) in turbot,Scophthalmus maximus

This report describes the cloning, characterization and gene expression pattern of two Th17 cytokines, interleukin (IL)-17A/F and -22, in turbot Scophthalmus maximus. The turbot IL-17A/F cDNA contains a 516 bp open reading frame encoding a deduced IL-17A/F protein of 171 amino acid (aa) residues, containing a predicted signal peptide of 31 aa. Turbot IL-22 had a 564 bp ORF coding for a 187 aa protein with a 33 aa signal peptide. The turbot IL-22 protein contained a typical IL-10 family signature. Both cytokines had highest expression levels in the intestine followed by head kidney and gills. Stimulation with the Gram negative bacterium Aeromonas salmonicida was able to modulate IL-17A/F and IL-22 expression in head kidney, spleen and liver but not the intestine. PMA and PHA were also able to induce the expression of both cytokines, suggesting that, as expected, T-cells are likely the main producers of these molecules in turbot as in mammals.     

RACK-1, a multifaceted regulator is required for C. elegans innate immunity against S. flexneri M9OT infection

The nematode C. elegans has the ability to clear off bacterial colonization in the intestine using pathogen specific innate immune response. Here, we show that C. elegans RACK-1 has been vital in determining the survival of worms under specific pathogenic infection. Among various pathogens tested, S. flexneri M9OT (SF) exhibited highest pathogenicity by killing rack-1 mutant worm-VC3013 earlier when compared to WT. The expression level of rack-1 mRNA was found to be decreased and it further indicated that the host translational event appeared to be affected during SF infection. Hence, inhibition of translational machinery was the foremost reason for the early mortality in C. elegans. Apparently, variation in the expression of RACK-1 affects the activation of p38 and JNK-MAPK pathway which consequently triggered expression of nlp-29 and longevity, respectively. The study unveils novel defense mechanisms exist for C. elegans in facilitating enhanced immunity by RACK-1 against SF infection.     

Identification and involvement of ferritin in the response to pathogen challenge in the abalone,Haliotis diversicolor

Accumulating data has demonstrated that ferritin plays an important role in host defense responses against infection by pathogens in many organisms. In this study, ultracentrifugation was used to isolate ferritin from abalone, Haliotis diversicolor, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis revealed that this ferritin consisted of two subunits (designated as HdFer1 and HdFer2). There are no disulfide bonds between the HdFer1 and HdFer2 subunits; however, these subunits co-assemble to form heteropolymers. A novel ferritin subunit (HdFer2) was cloned from H. diversicolor by 5ʹ and 3ʹ RACE (rapid amplification of cDNA ends) approach. The full-length HdFer2 cDNA sequence consists of 878 bp with an open reading frame of 513 bp that encodes a protein that is 170 amino acids in length. Quantitative real-time PCR analysis revealed that HdFer1 and HdFer2 were transcribed in various tissues, such as the mantle, gill and hepatopancreas, with the highest levels of expression in the hepatopancreas. Following a challenge with the pathogen, Vibrio harveyi, the expression of HdFer1 and HdFer2 were markedly induced at different times. This study has identified a novel ferritin subunit in H. diversicolor which will contribute to further exploration of the role of ferritin in mollusk innate immune defense against invading pathogens.     

HLA‐G 14‐bp Ins/Ins Genotype in Patients Harbouring Helicobacter pylori Infection: A Potential Risk Factor?

H. pylori is a potent pathogen due to its capacity to successfully evade host defence mechanisms. Despite inducing immune responses in infected individuals, sometimes these responses fail to clear the infection and the bacterium establishes a persistent infection leading to chronic inflammation. In this context, we hypothesized that human leucocyte antigen G (HLA‐G), a non‐classical major histocompatibility complex molecule that has the ability to regulate immune responses both in physiological and in pathological conditions, may play an important role in promoting tolerance and helping H. pylori to subvert host defence and consequently establish a chronic infection. Therefore, we evaluated the expression of HLA‐G 14‐bp Ins/Del polymorphism in patients harbouring H. pylori infection, as well as their relationship with histological and demographic variables, to gain a better understanding of the actual role of HLA‐G and its genetic polymorphisms in bacterial infection. Sixty‐eight patients with clinical symptoms suggestive of H. pylori infection were enrolled to assess HLA‐G 14‐bp Ins/Del polymorphism allele and genotype frequencies. After adjustment for covariates (age and gender), the odds of having the genotype Ins/Ins, compared to Del/Del, were 3.77 times greater among HP+ cases than among controls. These findings suggest that the 14‐bp Ins/Ins genotype, already associated with inflammatory and autoimmune diseases as well as some viral and parasitic infections, could confer a greater risk of developing H. pylori infection.     

Innate immune defence in the human gastrointestinal tract

The mucosal surface of the gastrointestinal tract represents a major entry point and ecological niche for many microbes. It forms an important immune barrier, absorbing nutrients, whilst preventing invasion by organisms. Of the extra-ordinarily diverse species that comprise the microbial world, relatively few organisms are able to succeed in breaching this barrier in an otherwise healthy host. The production and secretion of antimicrobial peptides (AMPs) from surface epithelia and circulating immune cells are likely to play a key role in host protection and homeostasis. A number of these peptides are constitutively produced providing resident protection, whereas others are induced during infection and inflammation. In addition to directly eradicating microorganisms, it is becoming increasingly apparent that AMPs are multi-functional with diverse immuno-modulatory properties. This review focuses on three families of AMPs, defensins, cathelicidins, and lysozyme, and discusses their role in mucosal defence.     

Border maneuvers: deployment of mucosal immune defenses against Toxoplasma gondii

Toxoplasma gondii is a highly prevalent protozoan pathogen that is transmitted through oral ingestion of infectious cysts. As such, mucosal immune defenses in the intestine constitute the first and arguably most important line of resistance against the parasite. The response to infection is now understood to involve complex three-way interactions between Toxoplasma, the mucosal immune system, and the host intestinal microbiota. Productive outcome of these interactions ensures resolution of infection in the intestinal mucosa. Nonsuccessful outcome may result in emergence of proinflammatory damage that can spell death for the host. Here, we discuss new advances in our understanding of the mechanisms underpinning these disparate outcomes, with particular reference to initiators, effectors, and regulators of mucosal immunity stimulated by Toxoplasma in the intestine.     

Regulation of epithelial immunity by IL-17 family cytokines

Cutaneous and mucosal epithelial cells function as both a physical barrier and as immune sentinels against environmental challenges, such as microbial pathogens, allergens and stress. The crosstalk between epithelial cells and leukocytes is essential for orchestrating proper immune responses during host defense. Interleukin (IL)-17 family cytokines are important players in regulating innate epithelial immune responses. Although IL-17A and IL-17F promote antibacterial and antifungal responses, IL-17E is essential for defense against parasitic infections. Emerging data indicate that another member of this family, IL-17C, specifically regulates epithelial immunity. IL-17C production serves as an immediate defense mechanism by epithelial cells, utilizing an autocrine mechanism to promote antibacterial responses at barrier surfaces.     

Intestinal innate immunity to Campylobacter jejuni results in induction of bactericidal human beta-defensins 2 and 3

Campylobacter jejuni is the most prevalent cause of bacterial diarrhea worldwide. Despite the serious health problems caused by this bacterium, human innate immune responses to C. jejuni infection remain poorly defined. Human beta-defensins, a family of epithelial antimicrobial peptides, are a major component of host innate defense at the gastrointestinal mucosal surface. In this study, the effect of two different C. jejuni wild-type strains on human intestinal epithelial innate responses was investigated. Up-regulation of beta-defensin gene and peptide expression during infection was observed and recombinant beta-defensins were shown to have a direct bactericidal effect against C. jejuni through disruption of cell wall integrity. Further studies using an isogenic capsule-deficient mutant showed that, surprisingly, the absence of the bacterial polysaccharide capsule did not change the innate immune responses induced by C. jejuni or the ability of C. jejuni to survive exposure to recombinant beta-defensins. This study suggests a major role for this family of antimicrobial peptides in the innate immune defense against this human pathogen.     

Macrophage regulation & function in helminth infection

Macrophages are innate immune cells with essential roles in host defense, inflammation, immune regulation and repair. During infection with multicellular helminth parasites, macrophages contribute to pathogen trapping and killing as well as to tissue repair and the resolution of type 2 inflammation. Macrophages produce a broad repertoire of effector molecules, including enzymes, cytokines, chemokines and growth factors that govern anti-helminth immunity and repair of parasite-induced tissue damage. Helminth infection and the associated type 2 immune response induces an alternatively activated macrophage (AAM) phenotype that – beyond driving host defense - prevents aberrant Th2 cell activation and type 2 immunopathology. The immune regulatory potential of macrophages is exploited by helminth parasites that induce the production of anti-inflammatory mediators such as interleukin 10 or prostaglandin E₂ to evade host immunity. Here, we summarize current insights into the mechanisms of macrophage-mediated host defense and repair during helminth infection and highlight recent progress on the immune regulatory crosstalk between macrophages and helminth parasites. We also point out important remaining questions such as the translation of findings from murine models to human settings of helminth infection as well as long-term consequences of helminth-induced macrophage reprogramming for subsequent host immunity.     

Evaluation of immune response in turbot (Psetta maxima L.) tenacibaculosis: Haematological and immunohistochemical studies

Tenacibaculum maritimum is a Gram-negative bacterium that causes large losses in farmed turbot, Psetta maxima (L.). Main lesions included erosive and ulcerative skin injuries in different locations in the body of the fish. This study was set up to gain insights into the immune response of the turbot against this bacterial infection. To determine the variations in the peripheral blood leukocytes a minimum of 200 leukocytes in blood smears were counted. The presence of immunoglobulin positive (Ig⁺) cells in spleen, kidney, intestine, thymus and skin from turbot experimentally infected with T. maritimum were studied using an immunohistochemical method. In fish challenged with the bacterium the percentage of circulating granulocytes, lymphocytes and trombocytes showed significant differences at different points of sampling. Moreover, the number of Ig⁺ cells significantly increased in the spleen, kidney, intestine and thymus. In the skin a strong immunoreactivity was observed in the interstitial liquid in damaged areas. The results obtained in this study indicated that the infection by T. maritimum in turbot induced an immune response that involved changes in the number of peripheral blood leukocytes and in the number and distribution of Ig⁺ cells.     

Innate lymphoid cells: critical regulators of allergic inflammation and tissue repair in the lung

Maintenance of epithelial barrier function in the skin, respiratory tract and intestine is critical to limit exposure to commensal and pathogenic microbes and to maintain tissue homeostasis. Innate lymphoid cells (ILCs) are a recently recognized innate immune cell population that plays critical roles in host defense, regulation of inflammation and promotion of wound healing and tissue repair at barrier surfaces. In this review we discuss recent advances in the understanding of how ILC populations in the respiratory tract impact allergic airway inflammation and lung epithelial repair.     

Clearance of Citrobacter rodentium requires B cells but not secretory immunoglobulin A (IgA) or IgM antibodies

Citrobacter rodentium, a murine model pathogen for human enteropathogenic Escherichia coli, predominantly colonizes the lumen and mucosal surface of the colon and cecum and causes crypt hyperplasia and mucosal inflammation. Mice infected with C. rodentium develop a secretory immunoglobulin A (IgA) response, but the role of B cells or secretory antibodies in host defense is unknown. To address this question, we conducted oral C. rodentium infections in mice lacking B cells, IgA, secreted IgM, polymeric Ig receptor (pIgR), or J chain. Normal mice showed peak bacterial numbers in colon and feces at 1 week and bacterial eradication after 3 to 4 weeks. B-cell-deficient mice were equally susceptible initially but could not control infection subsequently. Tissue responses showed marked differences, as infection of normal mice was accompanied by transient crypt hyperplasia and mucosal inflammation in the colon and cecum at 2 but not 6 weeks, whereas B-cell-deficient mice had few mucosal changes at 2 weeks but severe epithelial hyperplasia with ulcerations and mucosal inflammation at 6 weeks. The functions of B cells were not mediated by secretory antibodies, since mice lacking IgA or secreted IgM or proteins required for their transport into the lumen, pIgR or J chain, cleared C. rodentium normally. Nonetheless, systemic administration of immune sera reduced bacterial numbers significantly in normal and pIgR-deficient mice, and depletion of IgG abrogated this effect. These results indicate that host defense against C. rodentium depends on B cells and IgG antibodies but does not require production or transepithelial transport of IgA or secreted IgM.     

Inflammation and clearance of Chlamydia trachomatis in enteric and nonenteric mucosae

Immunization(s) fostering the induction of genital mucosa-targeted immune effectors is the goal of vaccines against sexually transmitted diseases. However, it is uncertain whether vaccine administration should be based on the current assumptions about the common mucosal immune system. We investigated the relationship between mucosal sites of infection, infection-induced inflammation, and immune-mediated bacterial clearance in mice using the epitheliotropic pathogen Chlamydia trachomatis. Chlamydial infection of the conjunctival, pulmonary, or genital mucosae stimulated significant changes in tissue architecture with dramatic up-regulation of the vascular addressin, VCAM, a vigorous mixed-cell inflammatory response with an influx of alpha4beta1+ T cells, and clearance of bacteria within 30 days. Conversely, intestinal mucosa infection was physiologically inapparent, with no change in expression of the local MAdCAM addressin, no VCAM induction, no histologically detectable inflammation, and no tissue pathology. Microbial clearance was complete within 60 days in the small intestine but bacterial titers remained at high levels for at least 8 months in the large intestine. These findings are compatible with the notion that VCAM plays a functional role in recruiting cells to inflammatory foci, and its absence from the intestinal mucosa contributes to immunologic homeostasis at that site. Also, expression of type 1 T cell-mediated immunity to intracellular Chlamydia may exhibit tissue-specific variation, with the rate and possibly the mechanism(s) of clearance differing between enteric and nonenteric mucosae. The implications of these data for the common mucosal immune system and the delivery of vaccines against mucosal pathogens are discussed.     

The development and function of mucosal lymphoid tissues: a balancing act with micro-organisms

Mucosal surfaces are constantly exposed to environmental antigens, colonized by commensal organisms and used by pathogens as points of entry. As a result, the immune system has devoted the bulk of its resources to mucosal sites to maintain symbiosis with commensal organisms, prevent pathogen entry, and avoid unnecessary inflammatory responses to innocuous antigens. These functions are facilitated by a variety of mucosal lymphoid organs that develop during embryogenesis in the absence of microbial stimulation as well as ectopic lymphoid tissues that develop in adults following microbial exposure or inflammation. Each of these lymphoid organs samples antigens from different mucosal sites and contributes to immune homeostasis, commensal containment, and immunity to pathogens. Here we discuss the mechanisms, mostly based on mouse studies, that control the development of mucosal lymphoid organs and how the various lymphoid tissues cooperate to maintain the integrity of the mucosal barrier.     

Necessary and sufficient role for T helper cells to prevent fungal dissemination in allergic lung disease

Mucosal immune responses to fungal infection range from T helper type 2 (Th2) cell-directed allergic inflammation to Th1-predominant neutrophilic inflammation, but the mechanisms directing these divergent mucosal immune outcomes and the role of T cells in host defense against mucosal fungal infections are not known. Here we examined the mouse mucosal immune responses to 12 filamentous environmental fungal species over a broad range of exposure doses and determined the requirement of T cells for host defense. For all tested fungi, low-grade conidium exposures induced Th2- and eosinophil-predominant allergic lung disease, whereas higher exposures led to rapid conversion to neutrophil- and Th1 cell-predominant inflammation, a phenomenon we term immune phenotype switching. All fungal exposure doses were further linked to the secretion of interleukin-17A (IL-17A). Fungal infections with Curvularia lunata and Aspergillus fumigatus were typically confined to the airway during allergic inflammation but became locally invasive and disseminated to the brain at higher conidium challenge doses, in association with predominant Th1 responses. Fungal dissemination occurred at relatively low challenge doses with the conidia of Aspergillus fumigatus administered to recombinase activating gene 1 (Rag-1)-deficient mice, which lack B and T cells, but B cell-deficient μMT mice and T helper cell-reconstituted Rag-1-deficient mice were comparable to wild-type mice in preventing fungal dissemination. Our findings demonstrate that Th2 cell-predominant allergic responses followed by immune phenotype switching and fungal dissemination are highly predictable outcomes with progressive fungal infectious burdens and that T helper cell responses are protective against lethal fungal dissemination.     

Cytoskeletal Regulation of Epithelial Barrier Function During Inflammation

Increased epithelial permeability is a common and important consequence of mucosal inflammation that results in perturbed body homeostasis and enhanced exposure to external pathogens. The integrity and barrier properties of epithelial layers are regulated by specialized adhesive plasma membrane structures known as intercellular junctions. It is generally believed that inflammatory stimuli increase transepithelial permeability by inducing junctional disassembly. This review highlights molecular events that lead to disruption of epithelial junctions during inflammation. We specifically focus on key mechanisms of junctional regulation that are dependent on reorganization of the perijunctional F-actin cytoskeleton. We discuss critical roles of myosin-II–dependent contractility and actin filament turnover in remodeling of the F-actin cytoskeleton that drive disruption of epithelial barriers under different inflammatory conditions. Finally, we highlight signaling pathways induced by inflammatory mediators that regulate reorganization of actin filaments and junctional disassembly in mucosal epithelia.The epithelium plays an important role in inflammation by serving as an interface between invading pathogens and the immune system of the host. Under physiological conditions, polarized epithelia form a protective barrier that allows regulated paracellular fluxes of solutes and nutrients as well as antigen sampling and surveillance by mucosal immune cells. However, during inflammation, this protective mechanism becomes compromised by various stimuli that originate on both sides of the epithelial barrier. On the apical (luminal) side, invading pathogenic microorganisms increase epithelial permeability to gain access into host tissue. The pathogens release a variety of epithelial barrier-disrupting agents that include pore-forming toxins, cytoskeleton-modifying proteins, and bacterial lipopolysaccharide (LPS). On the basal (tissue) side of the epithelial layer, activated immune cells also induce barrier disruption to facilitate their movement to sites of pathogen invasion. Mucosal immune cells increase epithelial permeability by secreting proinflammatory cytokines such as interferon (IFN) γ, tumor necrosis (TNF) α, and interleukin (IL)−1β or by releasing proteases and reactive oxygen species (ROS). As a result, mucosal inflammation commonly leads to sustained epithelial barrier compromise, which increases body exposure to external noxious agents, thereby further exaggerating the inflammatory response. Consequently it is believed that decreasing epithelial permeability may have beneficial effects by limiting inflammatory responses. Thus, understanding mechanisms that control the epithelial barrier disruption is important in identifying novel molecular targets for pharmacological modulation of mucosal inflammation.Properties of the epithelial barrier are regulated by specialized plasma membrane structures referred to as apical junctions. These structures are composed of adhesive and scaffolding proteins that are anchored into different cytoskeletal structures such as actin filaments, intermediate filaments, and microtubules. During inflammation, it is known that reorganizations of apical junctions mediate epithelial barrier dysfunction. Mounting evidence suggests that the actin cytoskeleton plays a pivotal role in regulating junctional integrity and remodeling under physiological and pathological states.In this review, we will discuss the role of actin filaments in the regulation of epithelial barrier integrity and its breakdown during inflammation. How inflammatory processes in different mucosal tissues induce remodeling of junction-associated filaments (F) actin and alter structure and barrier properties of epithelial cell-cell adhesions will be discussed. While epithelial junctions will be the major focus of this review, we will occasionally refer to examples of junctional regulation in the vascular endothelium. Likewise, we limit the discussion to simple columnar epithelia and exclude complex stratified epithelia such as the epidermis. Finally, we specifically focus on the role of actin filaments in maintenance and disassembly of epithelial junctions without discussing junctional reassembly during epithelial wound healing.     

The interaction of intestinal microbiota and innate lymphoid cells in health and disease throughout life

Immunity is shaped by commensal microbiota. From early life onwards, microbes colonize mucosal surfaces of the body and thereby trigger the establishment of immune homeostasis and defense mechanisms. Recent evidence reveals that the family of innate lymphoid cells (ILCs), which are mainly located in mucosal tissues, are essential in the maintenance of barrier functions as well as in the initiation of an appropriate immune response upon pathogenic infection. In this review, we summarize recent insights on the functional interaction of microbiota and ILCs at steady-state and throughout life. Furthermore, we will discuss the interplay of ILCs and the microbiota in mucosal infections focusing on intestinal immunity.