EDITORIAL FOCUS
Nitric oxide production by human macrophages: there's NO doubt about it

Ferric C. Fang1 and Andrés Vazquez-Torres2

1 Departments of Laboratory Medicine and Microbiology, University of Washington School of Medicine, Seattle, Washington 98195-7242; and 2 Department of Microbiology, University of Colorado Health Sciences Center, Denver, Colorado 80262


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THE DISCOVERY OF NITRIC OXIDE (NO) production from L-arginine by NO synthases has revolutionized our understanding of many aspects of human physiology. Since the seminal studies of Furchgott and Zawadzki (7), tremendous progress has been made in understanding the regulation of vascular tone by NO. Novel pharmacological agents based on the circulatory actions of NO have already found widespread clinical application. However, this contrasts with the role of NO in resistance to infection, where clinical applications have as yet been largely unrealized. The groundbreaking studies of Hibbs et al. (8), demonstrating NO-dependent actions of murine macrophages, have proven more difficult to extrapolate to humans.

The inability to demonstrate NO production by human macrophages initially posed a stumbling block. Humans unquestionably produce dramatically elevated quantities of NO during infection and other inflammatory conditions (4, 24). Nevertheless, in vitro treatment of human peripheral blood mononuclear cells with lipopolysaccharide and interferon-gamma fails to elicit NO production (15), even though these stimuli are highly effective in provoking murine macrophages to generate copious quantities of NO (19). This led some investigators to question whether human macrophages are capable of NO generation (15).

The answer to this question has emerged incrementally from more than 100 studies over the past decade. It is now indisputable that human macrophages produce NO. NO synthase mRNA and protein have been repeatedly demonstrated in activated human macrophages in a variety of settings (10, 14, 25), along with biochemical evidence of NO production (23). Inducible NO synthase (iNOS) protein has been convincingly shown within infiltrating tissue macrophages of infected patients (1). Importantly, NO-dependent biological actions of human macrophages have also been documented (13). Recent observations correlating iNOS promoter polymorphisms with macrophage NO production and resistance to infection (11) lend further support to an important role of NO in human innate immunity. The key to demonstrating NO in human macrophages is to allow stimulation to occur in vivo. Although NO generation can be readily demonstrated in macrophages from patients with inflammatory conditions such as tuberculosis (12), rheumatoid arthritis (18), or malaria (2), in vitro conditions to stimulate NO production by peripheral blood mononuclear cells of healthy subjects remain incompletely characterized.

In this issue (Ref. 9, see p. L944), Hickman-Davis et al. make an important addition to the list of conditions found to promote NO production by human macrophages. Alveolar macrophages from some healthy lung transplant patients undergoing routine surveillance bronchoalveolar lavage, but not macrophages from normal volunteers, were found to spontaneously generate significant quantities of NO, which were further enhanced following stimulation with surfactant protein A (SP-A) or live Klebsiella pneumoniae. Notably, the SP-A-treated cells exerted antimicrobial activity against Klebsiella that could be abrogated by inhibition of NO production. This suggests that the transplanted lung represents a stimulatory environment for host defenses that promote resistance to bacterial infection and provides further evidence that collectins such as SP-A promote innate immunity in diverse ways. The demonstration of local NO synthesis in the setting of lung transplantation corroborates earlier studies showing iNOS mRNA in bronchoalveolar lavage fluid from lung allograft recipients (16). Although bacterial pneumonia is common in the initial period immediately following lung transplantation (3), alveolar macrophage activation may help to limit the occurrence of pneumonia thereafter.

The attempts of Hickman-Davis et al. (9) to elucidate the in vivo signals that precondition alveolar macrophages from certain transplanted patients to synthesize NO in response to SP-A, the mechanism by which this collectin augments phagocyte function and the role of NO production in the antibacterial activity of human phagocytes are less conclusive. Klebsiella alone elicits as much NO production as SP-A, indicating that NO is contributory but not sufficient to account for antimicrobial actions of SP-A. Similarly, SP-A-induced rises in intracellular Ca2+ appear necessary for these actions but occur in macrophages from both transplant patients and normal controls. Hickman-Davis and colleagues show that a combination of NO and reactive oxygen species produced by a xanthine/xanthine oxidase system can rapidly kill Klebsiella in vitro. However, it is unlikely that this explains the weak and predominantly bacteriostatic effects of NO observed in alveolar macrophages in vivo, and the investigators do not provide evidence for a role of reactive oxygen species production by the macrophages. A bacteriostatic effect of macrophage-derived NO would actually be consistent with NO-dependent antimicrobial actions observed in murine cells (22). The ability of NO to reversibly inhibit essential cellular processes by nitrosylation of thiols and metals provides a mechanistic rationale for bacteriostasis (17). In addition to direct antimicrobial activity, it is conceivable that NO-dependent actions of alveolar macrophages relate to pleiotropic gene regulatory (5, 17) effects of NO. In support of this hypothesis, the protective actions of NO in a murine model of pulmonary Klebsiella infection correlate with its role as a modulator of the expression of proinflammatory mediators (20).

The ability of NO to participate in diverse regulatory and cytotoxic actions (6) creates a continuing challenge to elucidating its precise molecular mechanisms in complex biological systems like the transplanted lung. Nevertheless, the observations of Hickman-Davis et al. (9) reinforce the message that NO is an important mediator in human innate immunity. Future therapeutic strategies for infectious diseases may be designed to enhance NO production at sites of infection or inhibit its deleterious collateral effects (21). Although it has taken some time, researchers studying human macrophages are finally learning to take NO for an answer.


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Address for reprint requests and other correspondence: F. C. Fang, 1959 NE Pacific St., Seattle, WA 98195-7242 (E-mail: fcfang{at}washington.edu).

10.1152/ajplung.00017.2002


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Am J Physiol Lung Cell Mol Physiol 282(5):L941-L943
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