Antimicrobial Peptides in Human Health and Disease Chapter Abstracts
Introduction
General Concepts of Antimicrobial Peptides, Past and Future
Michael Zasloff
Microbes outnumber us in our world and we are composed of materials that can readily serve as nutrients for microbial growth. Indeed, man, an organism of some 1015 cells, harbors in excess of 1013 bacteria, comprising hundreds or perhaps thousands of different species, not to speak of the untold numbers of fungi and viruses that call us their home. Yet most of the time, our bodies do not exhibit visible signs of any battle taking place between our immune system and these microbes that have taken up residence in and on our bodies. Thus, despite the presence of microbes in our oral cavity, on our skin, in our gut, on our conjunctivae, and so on, these surfaces do not normally exhibit the classic signs associated with what we have come to recognize as evidence of a response to microbial infection, namely, inflammation. In "health", our epithelial surfaces are free of redness, warmth, swelling, and pus. Why? Is the absence of signs of "battle" due to the failure of microbes to "physically" access our tissues, or to the engagement of host defense mechanisms that are not associated with signs of "inflammation"? This state of wellness is so free of visible signs that we have to a large extent failed to recognize and better understand the complex machinery required for its maintenance. Much of classical immunology has focused on better understanding the original signs and symptoms of infection and unraveling the natural history of the medical conditions in which signs of disease are evident. In addition, the success of vaccination and the obvious therapeutic importance of transplantation have stimulated studies of mechanisms engaged by mammals in these settings. We have taken for granted and, by and large, overlooked the defensive mechanisms that have permitted us to live in harmony with our microbial neighbors.
Section 1: Select Diverse Peptides with Antimicrobial Action in Humans
Chapter 1
The Role of Cathelicidins in the Innate Host Defences of Mammals
Margherita Zanetti
The cathelicidin peptides comprise one of several families of antimicrobial peptides that are found in neutrophils and epithelia as components of the early host defenses of mammals against infection. All cathelicidin family members are synthesized and stored in cells as two-domain proteins. These are split on demand to produce a cathelin protein and an antimicrobial peptide. Accumulating evidence indicates that both the cathelin portion and the C-terminal peptide exert biological activities connected with host protection. This review presents an overview of the structure and biology of cathelicidins and discusses recent progress in cathelicidin research- with emphasis on the functional properties and role in host defense of the human cathelicidin hCAP18/LL-37 Although investigators initially concentrated their attention on antibiotic activity, it is becoming clear now that the LL-37 is a multifunctional molecule that may mediate various host responses, and thus represents an essential component of the innate immune system in humans.
Chapter 2
The Role of Alpha and Beta Defensins in Human Defense
Jens-Michael Schrder
Defensins are cysteine-rich antimicrobial peptides made by plants, insects and mammals. They vary in length and number of disulfide bonds and have a complex three-dimensional folding pattern. In humans, four a-defensins (HNP-1, -2, -3 and -4) are stored in neutrophil granules. Two a-defensins (HD-5 and HD-6) are secreted as inactive precursor peptides by specialized gut epithelial cells (Paneth cells) and converted into antimicrobial peptides by proteolytic processing. Four human b-defensins (hBD-1, -2, -3 and -4) are predominantly produced by various epithelial cells in a relatively organ-specific manner and are often focally expressed in the uppermost, highly differentiated cell layers. hBD-1 is constitutively produced, while hBD-2, -3 and -4 are inducible by cytokines and/or contact with bacteria. Defensins show antimicrobial activity against various Gram-negative and Gram-positive bacteria, many fungi and some viruses. Human genome screening has identified many other human defensin genes that show a rather restricted tissue expression. Future research will focus on defensin gene transfer, large scale production and identification of selective inducers.
Chapter 3 Granulysin is a low molecular weight protein located in the granules of human natural killer cells and T-lymphocytes. The 9kDa form is processed from a larger 15kDa precursor and has lytic activity against a broad spectrum of microbes and tumor cells. Granulysin contributes to immune responses in infectious diseases, malignancies and graft rejection, where it is increased in peripheral blood cells and affected tissue. Towards the establishment of granulysin as a novel therapy in infectious diseases, peptides have been developed that maintain antibacterial activity, but have lost the ability to lyse eukaryotic cells. Several studies have measured granulysin expression in various diseases and suggest it could develop as a powerful marker to monitor cellular immunity.
Chapter 4 Hepcidin is a four-disulfide amphipathic cationic peptide made in the liver, distributed in plasma and excreted in urine. Although it is antimicrobial in vitro, its main function in mammals is the homeostatic feedback inhibition of intestinal iron uptake and iron recycling by macrophages. In agreement with its function as an iron-regulatory hormone, hepcidin synthesis is increased by iron loading, and decreased by anemia and hypoxia. However, like some antimicrobial peptides, hepcidin is markedly induced during infection and inflammation. There increased hepcidin levels in plasma result in decreased plasma iron levels, and eventually in the characteristic anemia associated with infection and inflammation. The specific molecular mechanisms of hepcidin effect on iron transport remain to be identified. Hepcidin-mediated alterations in iron metabolism are thought to contribute to host defense, perhaps by limiting iron availability to invading microorganisms.
Section 2: Tissues and their use of Antimicrobial Peptides
Chapter 5 Antimicrobial peptides are essential molecules in the microbicidal armament of certain blood cells. Killing of bacteria was the originally proposed function of antimicrobial peptides and is still considered as their basic function. In addition, antiviral and antifungal activities have been demonstrated for the peptides, but the physiological relevance of these activities remains to be confirmed. Recently, other functions of antimicrobial peptides have been emphasized, such as chemotaxis, stimulation of cytokine release, enhancement of immunoglobulin production, stimulation of angiogenesis, activation of cell proliferation and interaction with T-cells. Some of these activities indicate regulatory functions in the adaptive immune system and the peptides contribute to the link between innate and adaptive immunity. Thus, antimicrobial peptides are not evolutionary leftovers! Instead, they are emerging as key components in innate immunity by killing microbial intruders and by being signaling components. In this chapter, the main antimicrobial peptides in blood, i.e. defensins, cathelicidins and granulysin are discussed, with reference to their activities and function. However, many questions remain to be solved in relation to health and disease.
Chapter 6 The mammalian small intestinal epithelium has a large surface area that is potentially vulnerable to infection by lumenal microbes. Host defense of the small intestine is mediated in part by antimicrobial peptides and proteins secreted by Paneth cells, epithelial cells located at the base of the crypts of Lieberkhn. Among the secretory products, lysozyme, sPLA2, and defensins are the best-characterized antimicrobials. Studies of transgenic and knockout mice have supported a pivotal role of Paneth cell defensins in protection from oral bacterial pathogens. Recent studies have both explored structure-function analysis of Paneth cell defensins and begun to elucidate regulatory pathways of Paneth cell function. Other studies suggest that Paneth cells may be the targets of virulence mechanisms of bacterial pathogens. It is perhaps likely that the antimicrobial activity of Paneth cell secretory products on transient and resident microbes in the intestinal lumen will prove to have a profound influence on health. A better understanding of innate immunity mediated by Paneth cells may lead to therapeutic strategies to augment host defense of the digestive tract.
Chapter 7 Antimicrobial peptides are expressed in the respiratory tract and act as effector substances of the innate immune system. A variety of cells synthesize and secrete antimicrobial peptides including epithelial and professional host defense cells such as neutrophils, macrophages, and NK cells. In the human lung, b-defensins originate from epithelial cells, macrophages and lymphocytes. a-defensins are made by neutrophils. LL-37/hCAP-18 is produced by epithelial cells, neutrophils, lymphocytes, and macrophages. Antimicrobial peptides act as endogenous antibiotics by direct destruction of microorganisms. Furthermore, they bind to cellular receptors and are involved in a variety of processes ranging from inflammation and chemoattraction to wound healing and angiogenesis. Concentrations of antimicrobial peptides in lung secretions are altered in several pulmonary diseases. Based on their biological functions, antimicrobial peptides contribute to the pathogenesis of lung disease. This chapter describes basic and applied biology of antimicrobial peptides in the human lung.
Chapter 8 The oral cavity is a unique environment in which antimicrobial peptides play a key role in maintaining health and may have future therapeutic applications. Present evidence suggests that a-defensins, b-defensins, LL-37, histatin, and other antimicrobial peptides and proteins have distinct but overlapping roles in maintaining oral health and preventing bacterial, fungal, and viral adherence and infection. The expression of the inducible hBD-2 in normal oral epithelium, in contrast to other epithelia, and the apparent differential signaling in response to commensal and pathogenic organisms, provides new insights into innate immunity in this body site. Commensal bacteria are excellent inducers of hBD-2 in oral epithelial cells, suggesting that the commensal bacterial community acts in a manner to benefit the overall innate immune readiness of oral epithelia. This may have major significance for understanding host defense in the complex oral environment.
Chapter 9 Many tribes in China and Africa still subscribe to myths and legends to cure skin infections. Use of extracts from plants and frog skin to promote healing of cutaneous wounds, burns and infectious diseases can be traced back for thousand of years in these regions. Recent advances in medical science have started to unlock the magic of these traditional methods of cure. Discoveries of biologically active peptides and proteins in frog skin (Zasloff, 1987) and plants (Broekaert et al., 1995) may explain the beneficial effects of these archaic treatments. These bioactive peptides are extremely effective against a range of microbes and are therefore named antimicrobial peptides. The magic of nature is also part of our own skin defense system. Mammals are born with antimicrobial peptides deployed as an important defense against microbial invasion (Gallo and Nizet, 2003). The skin is a protective interface between internal organs and the outside environment. It is exposed daily to thousands of potential pathogens, a host of toxins and physical stress. In order to face these challenges, skin epithelium functions as a physical barrier. It has an active immunological role in antigen processing and presentation as well as production of cytokines and defense molecules such as antimicrobial peptides. The number of reports demonstrating the presence and upregulation of antimicrobial peptides in human skin is increasing and reflects the significance of these peptides in cutaneous innate immunity. Therefore, this chapter summarizes recent progress pertaining to antimicrobial peptides in skin protection and their potential therapeutic applications. The reader is directed to specific chapters on defensins (Chapter 2) and cathelicidins (Chapter 1) for a more detailed discussion of these molecules. However, for purposes of this chapter's discussion of the skin a brief review of some of the most studied antimicrobial peptide families is provided.
Section 3: Clinical Concepts
Chapter 10 The critical role played by antimicrobial peptides (AMPs) in mammalian innate immunity is increasingly recognized. Bacteria differ in their intrinsic susceptibility to AMPs, and the relative resistance of some important human pathogens to these defense molecules is now appreciated as an important virulence phenotype. Experimental analysis has identified diverse mechanisms of bacterial AMP resistance including altered cell surface charge, active efflux, production of proteases or trapping proteins, and modification of host cellular processes. The contribution of these resistance mechanisms to pathogenesis is confirmed through direct comparison of wild-type bacteria and AMP-sensitive mutants using in vivo infection models. Knowledge of the molecular basis of bacterial AMP resistance may provide new targets for antimicrobial therapy of human infectious diseases.
Chapter 11 Newborns and infants are at increased risk of invasive microbial infection suggesting a functional immaturity of the immune system. Despite the derivation of the term "innate" immunity, relatively little is known about the expression and function of this aspect of host defense during gestational development, birth, and subsequent growth. Recent studies have highlighted differences in the expression of antimicrobial proteins and peptides at birth suggesting age-dependent regulation of expression and important host defense roles in development. Some infectious and inflammatory diseases of childhood are associated with altered expression of antimicrobial proteins and peptides. Clinical trials of several congeners of antimicrobial proteins and peptides have included pediatric subjects, established the safety of these agents, and have, in some instances, suggested efficacy. As progress is made in understanding how newborns and children express and deploy antimicrobial proteins and peptides, and how such expression and/or function may be dysregulated in disease, it is likely that new indications will be identified for their use in preventing and treating infections and other diseases in the young.
Chapter 12 The use of peptides of the innate immune system as anti-infective therapeutic agents has attracted attention and investment for almost twenty years. However, such peptides have failed to gain "large pharma" acceptance and to achieve FDA approval in a number of topical indications such as impetigo, diabetic foot ulcers and oral mucositis. From both this experience and advances in the field there is now renewed effort to develop one of nature's most prolific defense mechanisms into valuable preventative and therapeutic agents. There are indications that the barriers to their success may now be eroding with companies developing peptides that are more stable, cost effective and targeted at specific indications. These applications include systemic infectious disease, acne, vaginitis, wound infection and inflammation. In addition, the use of such peptides as modulators of innate immunity, to treat infectious disease and inflammation, has added a further dimension to the field. In this chapter we define the rationale for the clinical use of peptides based upon those found in the innate immune system. This rationale is centered on the natural role of these bioactive peptides in the mammalian immune system. In addition, we outline the path taken to achieve this goal by the leading peptide development companies around the world.
Current Books:
Granulysin
Steffen Stenger and Robert L. Modlin
Hepcidin
Tomas Ganz
Antimicrobial Peptides in Human Blood
Gudmundur H. Gudmundsson and Birgitta Agerberth
The Role of Paneth Cell a-Defensins in Enteric Innate Immunity
Andre J. Ouellette and Charles L. Bevins
Antimicrobial Peptides in the Lung
Christoph Beisswenger and Robert Bals
Antimicrobial Peptides in the Oral Environment: Expression and Function in Health and Disease
Beverly A. Dale and L. Page Fredericks
Natural Antimicrobial Peptides: A Barrier against Human Skin Infection
Mohamed Zaiou, Richard L. Gallo and Marissa H. Braff
Antimicrobial Peptide Resistance in Human Bacterial Pathogens
Victor Nizet
A Pediatric Perspective on Antimicrobial Proteins and Peptides: Expression, Function, and Clinical Relevance
Ofer Levy
Therapeutic Applications of Innate Immunity Peptides
Lijuan Zhang, Scott M. Harris and Timothy J. Falla