Function and therapeutic potential of host defence peptides.

Cationic host defence (antimicrobial) peptides are an important component of the innate immune systems of a wide variety of plants, animals, and bacteria. Although most of these compounds have direct antimicrobial activities under specific conditions, a greater appreciation for the diversity of functions of these molecules is beginning to develop in the field. In addition to their directly antimicrobial activities, they also have a broad spectrum of activity on the host immune system, with both pro-inflammatory and anti-inflammatory effects being invoked. Increasingly sophisticated approaches to understand the role of host defence peptides in modulating innate immunity are already serving to guide the development of novel therapeutics.     

Defensins: antimicrobial peptides of innate immunity

The production of natural antibiotic peptides has emerged as an important mechanism of innate immunity in plants and animals. Defensins are diverse members of a large family of antimicrobial peptides, contributing to the antimicrobial action of granulocytes, mucosal host defence in the small intestine and epithelial host defence in the skin and elsewhere. This review, inspired by a spate of recent studies of defensins in human diseases and animal models, focuses on the biological function of defensins.     

Drosophila melanogaster embryonic haemocytes: masters of multitasking

Drosophila melanogaster haemocytes constitute the cellular arm of a robust innate immune system in flies. In the adult and larva, these cells operate as the first line of defence against invading microorganisms: they phagocytose pathogens and produce antimicrobial peptides. However, in the sterile environment of the embryo, these important immune functions are largely redundant. Instead, throughout development, embryonic haemocytes are occupied with other tasks: they undergo complex migrations and carry out several non-immune functions that are crucial for successful embryogenesis.     

Confronting physiology: how do infected flies die?

Fruit fly immunology is on the verge of an exciting new path. The fruit fly has served as a strong model for innate immune responses; the field is now expanding to use the fruit fly to study pathogenesis. We argue here that, to understand pathogenesis in the fly, we need to understand pathology - and to understand pathology, we need to confront physiology with molecular tools. When flies are infected with a pathogen, they get sick. We group the events following infection into three categories: innate immune responses (defence mechanisms by which the fly attempts to kill or neutralize the microbe, some of which can themselves cause harm to the fly); microbial virulence (mechanisms by which the microbe evades the immune response); and host pathology (physiologies adversely affected by either the immune response or microbial virulence). We divide this review into sections mirroring these categories. The molecular study of infection in the fruit fly has focused on the first category, has begun to explore the second, and has yet to tap the full potential of the fly regarding the third.     

Invertebrates as animal models for Staphylococcus aureus pathogenesis: a window into host-pathogen interaction

Recently, the use of invertebrate models of infection has given exciting insights into host-pathogen interaction for a number of bacteria. In particular, this has revealed important factors of the host response with remarkable parallels in higher organisms. Here, we review the advances attained in the elucidation of virulence determinants of a major human pathogen, Staphylococcus aureus, in relation to the invertebrate models thus far applied, the silkworm (Bombyx mori), the fruit fly (Drosophila melanogaster) and the roundworm (Caenorhabditis elegans). Also, the major pathways of host defence are covered in light of the response to S. aureus and the similarities and divergences in innate immunity of vertebrates and invertebrates. Consequently, we comparatively consider pathogen recognition receptors, signal transduction pathways (including Toll, Imd and others), and the humoral and cellular antimicrobial effectors. The technically convenient and ethically acceptable invertebrates appear as a valuable first tool to discriminate molecules participating from both sides of the host-S. aureus interaction as well as a high throughput method for antimicrobial screening.     

A family of helminth molecules that modulate innate cell responses via molecular mimicry of host antimicrobial peptides

Over the last decade a significant number of studies have highlighted the central role of host antimicrobial (or defence) peptides in modulating the response of innate immune cells to pathogen-associated ligands. In humans, the most widely studied antimicrobial peptide is LL-37, a 37-residue peptide containing an amphipathic helix that is released via proteolytic cleavage of the precursor protein CAP18. Owing to its ability to protect against lethal endotoxaemia and clinically-relevant bacterial infections, LL-37 and its derivatives are seen as attractive candidates for anti-sepsis therapies. We have identified a novel family of molecules secreted by parasitic helminths (helminth defence molecules; HDMs) that exhibit similar biochemical and functional characteristics to human defence peptides, particularly CAP18. The HDM secreted by Fasciola hepatica (FhHDM-1) adopts a predominantly α-helical structure in solution. Processing of FhHDM-1 by F. hepatica cathepsin L1 releases a 34-residue C-terminal fragment containing a conserved amphipathic helix. This is analogous to the proteolytic processing of CAP18 to release LL-37, which modulates innate cell activation by classical toll-like receptor (TLR) ligands such as lipopolysaccharide (LPS). We show that full-length recombinant FhHDM-1 and a peptide analogue of the amphipathic C-terminus bind directly to LPS in a concentration-dependent manner, reducing its interaction with both LPS-binding protein (LBP) and the surface of macrophages. Furthermore, FhHDM-1 and the amphipathic C-terminal peptide protect mice against LPS-induced inflammation by significantly reducing the release of inflammatory mediators from macrophages. We propose that HDMs, by mimicking the function of host defence peptides, represent a novel family of innate cell modulators with therapeutic potential in anti-sepsis treatments and prevention of inflammation.     

An alloy of zinc and innate immunity: Galvanising host defence against infection

Innate immune cells such as macrophages and neutrophils initiate protective inflammatory responses and engage antimicrobial responses to provide frontline defence against invading pathogens. These cells can both restrict the availability of certain transition metals that are essential for microbial growth and direct toxic concentrations of metals towards pathogens as antimicrobial responses. Zinc is important for the structure and function of many proteins, however excess zinc can be cytotoxic. In recent years, several studies have revealed that innate immune cells can deliver toxic concentrations of zinc to intracellular pathogens. In this review, we discuss the importance of zinc status during infectious disease and the evidence for zinc intoxication as an innate immune antimicrobial response. Evidence for pathogen subversion of this response is also examined. The likely mechanisms, including the involvement of specific zinc transporters that facilitate delivery of zinc by innate immune cells for metal ion poisoning of pathogens are also considered. Precise mechanisms by which excess levels of zinc can be toxic to microorganisms are then discussed, particularly in the context of synergy with other antimicrobial responses. Finally, we highlight key unanswered questions in this emerging field, which may offer new opportunities for exploiting innate immune responses for anti‐infective development.     

Metal ions in macrophage antimicrobial pathways: emerging roles for zinc and copper

The immunomodulatory and antimicrobial properties of zinc and copper have long been appreciated. In addition, these metal ions are also essential for microbial growth and survival. This presents opportunities for the host to either harness their antimicrobial properties or limit their availability as defence strategies. Recent studies have shed some light on mechanisms by which copper and zinc regulation contribute to host defence, but there remain many unanswered questions at the cellular and molecular levels. Here we review the roles of these two metal ions in providing protection against infectious diseases in vivo, and in regulating innate immune responses. In particular, we focus on studies implicating zinc and copper in macrophage antimicrobial pathways, as well as the specific host genes encoding zinc transporters (SLC30A, SLC39A family members) and CTRs (copper transporters, ATP7 family members) that may contribute to pathogen control by these cells.     

Intracellular innate resistance to bacterial pathogens

Mammalian innate immunity stimulates antigen-specific immune responses and acts to control infection prior to the onset of adaptive immunity. Some bacterial pathogens replicate within the host cell and are therefore sheltered from some protective aspects of innate immunity such as complement. Here we focus on mechanisms of innate intracellular resistance encountered by bacterial pathogens and how some bacteria can evade destruction by the innate immune system. Major strategies of intracellular antibacterial defence include pathogen compartmentalization and iron limitation. Compartmentalization of pathogens within the host endocytic pathway is critical for generating high local concentrations of antimicrobial molecules, such as reactive oxygen species, and regulating concentrations of divalent cations that are essential for microbial growth. Cytosolic sensing, autophagy, sequestration of essential nutrients and membrane attack by antimicrobial peptides are also discussed.     

The imd gene is required for local Cecropin expression in Drosophila barrier epithelia

Surfaces of higher eukaryotes are normally covered with microorganisms but are usually not infected by them. Innate immunity and the expression of gene-encoded antimicrobial peptides play important roles in the first line of defence in higher animals. The immune response in Drosophila promotes systemic expression of antimicrobial peptides in response to microbial infection. We now demonstrate that the epidermal cells underlying the cuticle of larvae respond to infected wounds by local expression of the genes for the antimicrobial peptide cecropin A. Thus, the Drosophila epidermis plays an active role in the innate defence against microorganisms. The immune deficiency (imd) gene was found to be a crucial component of the signal-induced epidermal expression in both embryos and larvae. In contrast, melanization, which is part of the wound healing process, is not dependent on the imd gene, indicating that the signalling pathways promoting melanization and antimicrobial peptide gene expression can be uncoupled.     

Modulating immunity as a therapy for bacterial infections

Despite our efforts to halt the increase and spread of antimicrobial resistance, bacteria continue to become less susceptible to antimicrobial drugs over time, and rates of discovery for new antibiotics are declining. Thus, it is essential to explore new paradigms for anti-infective therapy. One promising approach involves host-directed immunomodulatory therapies, whereby natural mechanisms in the host are exploited to enhance therapeutic benefit. The objective is to initiate or enhance protective antimicrobial immunity while limiting inflammation-induced tissue injury. A range of potential immune modulators have been proposed, including innate defence regulator peptides and agonists of innate immune components such as Toll-like receptors and NOD-like receptors.     

Innate host responses to enteric bacterial pathogens: a balancing act between resistance and tolerance

Infection by enteric bacterial pathogens activates pathogen recognition receptors, leading to innate responses that promote host defence. While responses that promote host 'resistance' to infection, through the release of antimicrobial mediators, or the recruitment of inflammatory cells aimed at clearing the infection are best known, recent studies have begun to identify additional innate driven responses that instead promote intestinal tissue repair and host survival. Described as infection 'tolerance' responses, we and others have primarily studied these responses in the Citrobacter rodentium infection model. In this review we discuss the impact of innate resistance mechanisms on host defence, and describe how 'tolerance' responses act primarily on the intestinal epithelium, triggering epithelial cell proliferation, repair or promoting barrier function. Resistance and tolerance responses appear to work together, with tolerance repairing the tissue injury caused by resistance driven inflammation. Tolerance responses fit a pattern where innate immunity and inflammation are tightly regulated in the gastrointestinal tract. Moreover, tolerance may have developed due to the successful subversion and avoidance of host resistance by enteric bacterial pathogens. Further studies are needed to clarify the contribution of different pathogen recognition receptors to tolerance and resistance responses against bacterial pathogens, in the gut or in other host tissues.     

Isolation and characterization of antimicrobial proteins and peptide from chicken liver.

Endogenous antimicrobial peptides and proteins are crucial components of the innate immune system and play an essential role in the defense against infection. Antimicrobial activity was detected in the acid extract of livers harvested from healthy adult White Leghorn hens, Gallus gallus. Two antimicrobial proteins and one antimicrobial polypeptide were isolated from the liver extract by cation-exchange and gel filtration chromatography, followed by two-step reverse-phase high-performance liquid chromatography (RP-HPLC). These antimicrobial components were identified as histones H2A and H2B.V, and histone H2B C-terminal fragment using peptide mass fingerprinting and partial sequencing by tandem nanoelectrospray mass spectrometry. The proteins and the peptide identified in the present study, which exhibited antimicrobial activity against both Gram-positive and Gram-negative bacteria, were thermostable and showed salt-resistant activity. The antimicrobial properties of histones and histone fragment in chicken provide further evidence that histones, in addition to their role in nucleosome formation, may play an important role in innate host defense against intracellular or extracellular microbe invasion in a wide range of animal species.     

Increased internal and external bacterial load during Drosophila aging without life-span trade-off

The role of microbial load during aging of the adult fruit fly Drosophila melanogaster is incompletely understood. Here we show dramatic increases in aerobic and anaerobic bacterial load during aging, both inside the body and on the surface. Scanning electron microscopy and cell staining analyses of the surface of aged flies detected structures resembling abundant small bacteria and bacterial biofilms. Bacteria cultured from laboratory flies included aerobic species Acetobacter aceti, Acetobacter tropicalis, and Acetobacter pasteurianus and anaerobic species Lactobacillus plantarum and Lactobacillus sp. MR-2; Lactobacillus homohiochii, Lactobacillus fructivorans, and Lactobacillus brevis were identified by DNA sequencing. Reducing bacterial load and antimicrobial peptide gene expression by axenic culture or antibiotics had no effect on life span. We conclude that Drosophila can tolerate a significant bacterial load and mount a large innate immune response without a detectable trade-off with life span; furthermore, microbes do not seem to limit life span under optimized laboratory conditions.     

Defensins in innate antiviral immunity

Defensins are small antimicrobial peptides that are produced by leukocytes and epithelial cells, and that have an important role in innate immunity. Recent advances in understanding the mechanisms of the antiviral action(s) of defensins indicate that they have a dual role in antiviral defence, acting directly on the virion and on the host cell. This Review focuses on the antiviral activities and mechanisms of action of mammalian defensins, and on the clinical relevance of these activities. Understanding the complex function of defensins in innate immunity against viral infection has implications for the prevention and treatment of viral disease.     

Cytokine-mediated regulation of antimicrobial proteins

Antimicrobial proteins constitute a phylogenetically ancient form of innate immunity that provides host defence at skin and mucosal surfaces. Although some components of this system are constitutively expressed, new evidence reviewed in this Progress article shows that the production of certain antimicrobial proteins by epithelial cells can also be regulated by cytokines of the innate and adaptive immune systems. In particular, the effector cytokines interleukin-17 and interleukin-22, which are produced by the T-helper-17-cell subset, are emerging as crucial regulators of antimicrobial-peptide production in the gut and the lungs. This suggests that this T-cell lineage and its cytokines have important roles in skin and mucosal immunity.     

The bactericidal/permeability-increasing protein (BPI) in the innate defence of the lower airways

The human BPI (bactericidal/permeability-increasing protein), stored in primary azurophilic granula of neutrophil granulocytes and produced by mucosal epithelia, has been known for decades to bind LPS (lipopolysaccharide) with very high affinity and to efficiently kill Gram-negative bacteria. Thus BPI potentially represents a central component of the innate immune system to directly combat microbes and modulate subsequent adaptive immune responses. Especially in the lungs, which are frequently exposed to a variety of inhaled pathogens, antimicrobial innate defence molecules such as BPI, are of exceptional relevance. In the present review, we highlight possible functions of BPI during acute pneumonia and CF (cystic fibrosis)-associated chronic infections in the lung.