PERSPECTIVE
Scratching the Surface
Inroads to a Better Understanding of Airway Host Defense

Charles L. Bevins

Lerner Research Institute, The Cleveland Clinic Foundation, Cleveland, Ohio

Mammalian mucosal surfaces, including that of the respiratory tract, are truly remarkable from the perspective of host defense. Their life-sustaining physiologic roles dictate that these sites maintain intimate contact with an outside world teeming with a wide array of microbes. In the respiratory and digestive tract, exposure is exaggerated by expansive surface area, which facilitates gas exchange or nutrient absorption, respectively. Environmental exposure at these sites predicts a host vulnerability, as it provides an opportunity for microbes to establish invasive infections. Yet, given the countless encounters with microbes, there is an astonishingly low incidence of infection. Furthermore, inflammation as a defense mechanism is infrequently mobilized. This is probably advantageous, as inflammation at these sites would disrupt normal physiology. These simple observations suggest that efficient, highly vigilant, non- inflammatory antimicrobial defenses, capable of dealing with a wide spectrum of microbes, have evolved to provide lifelong protection of these vital mucosal surfaces.

Innate immunity (also termed nonclonal or natural immunity) encompasses a complex of first-line host defense elements (1). At mucosal surfaces the innate defense system employs two broad and overlapping strategies that are central to effective defense: minimizing microbial adherence and creating a hostile environment for potential pathogens. In the respiratory tract, examples of innate defenses include: (1) physical processes, such as induced turbulent flow of inspired air, coughing, clearance through beating of cilia and shedding of epithelial cells, (2) chemical barriers, such as mucus, nitric oxide, and various peptides and proteins, and (3) cellular processes, such as phagocytosis by resident macrophages (2). Through these mechanisms, innate immunity provides direct incapacitation and elimination of pathogens. Innate immunity also provides mechanisms to recognize microbial organisms as foreign. This recognition can lead to inducible responses, which can amplify and integrate host-defense pathways. Additionally, the innate host defense mechanisms interface with the acquired (also termed clonal or adaptive) immune responses mediated by lymphocytes (7, 8). However, in contrast to the lymphocyte-mediated immune system, where an effective response involves both gene rearrangements and clonal selection developed over a period of days, the innate system remains ever-ready or immediately inducible.

Huttner and Bevins have recently proposed a working model of the host defense of mucosal surfaces in which elements of the innate host-defense system effectively deal with the vast majority of encounters with microbes and prevent infection (9). The model accommodates recent discoveries that defense of mammalian wet mucosal surfaces includes inducible and constitutive expression of antimicrobial peptides (10), inorganic molecules with antimicrobial activity (15, 16), and proteins that can directly inhibit microbial survival (17). These factors, highlighted in Figure 1, coupled with barrier properties and clearance mechanisms, constitute essential elements of innate mucosal immunity. In general, the antimicrobial factors are made in situ by surface and glandular epithelial cells, but in some cases they may also be derived from resident macrophages.

View larger version (14K): [in this window] [in a new window]   Figure 1.   A model of mucosal host defenses (based on that proposed by Huttner and Bevins [9], with permission).

Only when the local defenses are overwhelmed will the secondary lines of defense be called into action. The previously mentioned model suggests that inflammation and the acquired immune response as back-up systems are available to counter persistent or invasive challenges. Microorganisms may overwhelm local defenses if they have pathogenic attributes to evade defense mechanisms or if the challenge involves an exceptional number of microbes (Table 1). In addition, inadequate mucosal host-defense responses, for even routine microbial challenges, may result from genetic deficiencies, developmental immaturity, concurrent systemic disease, or environmental exposures (e.g., toxins). If the deficit in defense capacity is of limited duration, and if the host survives the infectious challenge, the mucosal defense system will return to baseline homeostasis. However, the consequence of long-term deficits would be persistent pathophysiology, which may include chronic inflammation and/or chronic mucosal infection.

A key study that focused attention on local innate defense of the respiratory tract was published in 1996 by Smith and colleagues from the University of Iowa (18). This study investigated the mechanisms that underlie the clinical observations of local host defense defects in the respiratory mucosa of individuals with cystic fibrosis (CF). Their findings indicate that defective function of CF transmembrane conductance regulator protein (CFTR) leads to an alteration in airway surface liquid (ASL) composition, which in turn, renders key antimicrobial factors in ASL less active. Specifically, an elevated NaCl concentration in the ASL may compromise activity of the key antimicrobial factor(s), permitting chronic bacterial colonization and airway infections characteristic of this disease. Since this initial report, attention has been turned toward defining more precisely the composition of the ASL and toward identifying the key antimicrobial factors in human airway surface fluid in order to better understand the lack of host defense at the CF respiratory mucosa (19).

The study by Travis and coworkers published in this volume (23) contributes to our understanding of antimicrobial factors in ASL in relation to the pathogenesis of CF. From a technical perspective, the investigators have developed a sensitive and quantitative assay of antimicrobial activity capable of efficiently handling large numbers of samples in microtiter plates. This assay may prove very valuable in high-throughput screening analysis. Levels of two abundant antimicrobial proteins in ASL, lysozyme and lactoferrin, were quantitated in the range of 20 to 100 µg/ml, sufficient to kill two important CF pathogens, Straphyloccus aureus and Pseudomonas aeruginosa. Interestingly, Burkholderia species are associated with severe CF infections late in the course of the disease and are relatively resistant to these proteins. A third antimicrobial peptide of ASL, secretory leukoprotease inhibitor, was less active against all three of these bacteria. For each of these proteins, in vitro antimicrobial activity was inhibited by either saline or the divalent cations Ca²⁺ and Mg²⁺, but not by nonionic osmolytes, mucin, or elastase. Inhibition depended both on the concentration of salt, and, at a single ionic strength, on the concentration of the antimicrobial factor. This suggests that partial inhibition by ionic strength could be overcome by higher levels of antimicrobial factors. Consistent with earlier results using primary cell cultures (18), when diluted and tested in vitro, ASL from CF specimens had levels of antimicrobial activity equal to those from normal controls, implying that correction of ASL alterations may restore effective host-defense activities.

Travis and her colleagues raise several issues that remain to be addressed further. First, the arsenal of antimicrobial factors in the ASL is complex and their individual, as well as collective, antimicrobial activities need to be defined better. Second, levels of the various ASL antimicrobial agents need to be quantitated in baseline states, throughout development and during various states of pathophysiology. This endeavor should include establishing biochemical pathways that regulate expression of these factors. Third, the effects that the many other components of the ASL (ions to macromolecules) have on antimicrobial activity need to be thoroughly addressed. Although challenging, the development of in vitro assays that reproduce the environment where endogenous antimicrobials naturally function would help clarify their biology. In terms of their relative contribution to host defense, further investigations are needed to elucidate more clearly the role of major antimicrobial factors in vivo.

In addition to appreciating a beautifully orchestrated biological system, defining defense mechanisms of the mammalian respiratory tract facilitate the development of therapeutics to combat airway infections in CF (Table 2). Two approaches may prove useful. One approach would entail supplementing the ASL with additional antimicrobial agents. Current therapeutics using conventional antimicrobial agents would fall into this approach, but increased therapeutic effectiveness might be achieved if future drug development directly addressed the possibility of synergistic (or inhibitory) interactions with endogenous antimicrobials and other components of the CF ASL. An alternate strategy would use antimicrobials, which are based on endogenous molecules. Innate host-defense effector molecules of human or xenobiotic origin may overcome inhibitory factors of CF ASL if supraphysiological concentrations are achieved. Xenobiotics and synthetic derivatives could be selected based on their spectrum of antimicrobial activity and relative resistance to the inhibitory milieu of the CF ASL. In the long term, therapeutics may be designed to modulate levels of endogenous antimicrobial factors when pathways regulating their expression are better elucidated. Additionally, it may be possible to generate therapeutic concentrations of peptides and proteins in situ by delivering genes that encode these molecules to surface and/or glandular epithelial cells of the airway. The second approach would be aimed at altering the composition of the ASL to enhance activity of endogenous factors. Ultimately, restoration of CFTR function to normal levels in the airway is predicted to accomplish this goal. However, in the short term, this strategic approach could involve pharmacologic agents that would alter selected aspects of ASL composition.

Finally, therapeutic interventions in very young CF patients would be aimed at preventing pathologic colonization prior to extensive airway inflammation. Here, useful strategies may be very different from those geared toward older CF patients who have a heavy burden of bacteria and pronounced inflammation. The numbers and types of bacteria, the distribution of host cells in the airway, and the many aspects of ASL composition in these two settings are significantly different. Development of new strategies for treating the life-threatening bacterial infection of the CF airway will emerge as we advance our understanding of the pathways involved in innate mucosal defense.

	Footnotes

Address correspondence to: Charles L. Bevins, M.D., Ph.D., Research Institute/NB30, The Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland OH 44195. E-mail: bevinsc{at}ccf.org

(Received in original form March 24, 1999).

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