© 2003 American Thoracic Society
New Insights from an Old ModelColumbia University New York, New York The respiratory mucosa is often exposed to contamination from opportunistic pathogens such as Pseudomonas aeruginosa. Whether these organisms can establish infection in the airways is dependent on many factors such as the genetic flexibility of bacteria to adapt rapidly to a new environmental niche and to avoid innate clearance mechanisms, as well as the nature of the host response to the pathogen. Although the normal host is rarely infected by this route, bacterial contamination of damaged airways is common following viral infection or prolonged intubation, or in patients with chronic bronchitis or cystic fibrosis. The innate resistance of normal mice and even cftr-/- mice to pulmonary infection reflects the efficiency of the mucosal immune system in protecting the lung and has also limited the efficacy of animal models of infection in establishing which bacterial and host factors are critical in airway infection. It has been even more difficult to determine exactly what is responsible for P. aeruginosa infection in cystic fibrosis. Over 20 years ago, Cash and coworkers (1) developed a rat model of airway infection, using agar beads containing P. aeruginosa, to try to approximate the airway infection in cystic fibrosis. Although many useful observations have been made with this model, it is limited by the need to introduce a foreign body (agar beads), circumvention of nasopharyngeal colonization, and the requirement for high bacterial inocula, conditions that may not be physiological. Despite these limitations, in this issue of AJRCCM (pp. 1478–1482), Lesprit and coworkers (2) use the Cash model effectively to make some novel observations regarding the pathogenesis of P. aeruginosa infection in the lung. They demonstrate that, unlike wild-type P. aeruginosa, which elicits a polymorphonuclear leukocyte response, avirulent mutants of P. aeruginosa instilled directly into the lower airways evoke a mononuclear infiltrate. The immune response to P. aeruginosa lasR mutants, which lack the ability to coordinately express a number of critical virulence factors, was primarily due to recruited macrophages and was tolerated at all but the highest inocula. In contrast, rats infected with wild-type bacteria, which stimulate a florid neutrophil response and vascular congestion, succumbed to infection. By manipulating the bacterial inocula used, these investigators could demonstrate that the neutrophil response, under the appropriate conditions, serves to clear the organisms, whereas the infiltration of mononuclear cells, despite less immediate toxicity, does not. It has been shown previously that lasR mutants are defective in the production of a large number of virulence genes (3). These mutants lack expression of one of the quorum sensing systems necessary for P. aeruginosa to evade the local activity of lactoferrin (4) and to adapt to the airway milieu by forming a biofilm (5). The homoserine lactone "quorum sensor" is secreted by bacteria and activates epithelial interleukin-8 expression, a major neutrophil chemokine (6). When lasR mutants are introduced intranasally in a neonatal mouse model of infection, they cause minimal pathology because they cannot establish infection (7, 8). The direct clinical relevance of such rodent studies may not be immediately apparent, because humans with airway infection rarely become bacteremic or septic when infected with even the wild-type, more virulent organisms. By directly instilling avirulent mutants into the lung, using the Cash model, however, Lesprit and coworkers are able to demonstrate that expression of lasR-dependent virulence factors is necessary to evoke an effective neutrophilic response. Although it was not surprising that a lasR mutant did not cause invasive infection, it is of considerable interest that the organisms elicited a macrophage response, not expected of extracellular gram-negative organisms. Because the lasR mutation does not affect LPS biosynthesis there should be considerable amounts of LPS in the rat airways, shed by both wild-type and mutant bacteria. In the presence of large numbers of organisms (107 cfu/ml), however, corresponding amounts of LPS, and an intact immune system, the predominant response to the lasR mutant strain was that of mononuclear leukocytes and not neutrophils. These findings indicate that P. aeruginosa LPS, even in vivo, is not particularly immunostimulatory when present in the airway, and is insufficient to recruit neutrophils in this setting. In the absence of neutrophilic influx, there was relatively minimal pathology and significantly decreased mortality. This is in contrast to what occurs when LPS is present in the bloodstream and potently activates a systemic immune response. Several in vitro studies demonstrate that airway epithelial cells are not activated by P. aeruginosa LPS (6). The lack of pathology associated with infection by the lasR mutant indicates that it is the expression of genes regulated by lasR (not LPS) that is especially important in eliciting the critical neutrophilic response. This study also demonstrates, again, that it is the neutrophilic response that is crucial in clearing P. aeruginosa from the airway. Infections in neutropenic mice have been used for decades to study the efficacy of anti-P. aeruginosa antimicrobial agents. Although the role of neutrophils in killing P. aeruginosa is well established, particularly in cystic fibrosis, the damaging effects of neutrophil-dominated inflammation clearly contribute to the pathology of this disease. It has not been entirely clear how macrophages might perform in this setting. As shown in this report, rats that had primarily a macrophage response not only tolerated 107 cfu/ml of bacteria in their airways, they failed to limit bacterial replication. Thus, a conclusion from these studies is that mononuclear cells are not sufficient to contain a P. aeruginosa infection, even with organisms unable to form biofilms or to invade. While at first glance these authors present yet another study of a well characterized P. aeruginosa mutant in an agar bead model of infection, several novel inferences may be made from their data. These authors clearly show that the expression of lasR-dependent immunostimulatory virulence factors (not LPS) and their recruitment of a neutrophilic response are critical for the clearance of P. aeruginosa from the airways. REFERENCES
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