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The worldwide incidence, prevalence, and fatality rates from asthma are increasing despite currently available therapeutic modalities. Systemic administration of interleukin (IL)-12 has been shown to inhibit airway reactivity in murine models of asthma, but the required dosage is high and may be toxic. This study tested the hypothesis that IL-12 administered directly into the lungs is more effective in inhibiting airway reactivity than systemically administered IL-12, allowing lower doses to be used. A low dose (10 ng) of IL-12 was delivered either intratracheally (mucosal delivery) or intraperitoneally (systemic delivery) at the time of ragweed (RW) challenge in mice sensitized to RW. Basal airway resistance and airway reactivity to methacholine were measured 3 days after RW challenge. Compared to phosphate-buffered saline (PBS) challenge of RW sensitized mice, RW challenge increased basal resistance and the slope of the methacholine dose-response curve. Methacholine challenge of RW-challenged mice also induced premature respiratory failure (respiratory rate < 150/min, tidal volume < 0.15 mL) in some animals. Administration of mucosal or systemic IL-12 at the time of RW challenge decreased basal airway resistance. However, only mucosal IL-12 decreased airway reactivity and inhibited respiratory failure during methacholine challenge. These findings indicate that mucosal delivery of a low dose of IL-12 is more effective than systemic IL-12 in inhibiting airway reactivity and respiratory failure in a mouse model of asthma.

The relative efficacy of mucosal (intratracheal) and systemic (intraperitoneal) delivery of interleukin (IL)-12 was evaluated in a mouse model of allergic lung eosinophilia. Mucosal administration of IL-12 achieved 100- to 600-fold higher bronchoalveolar lavage (BAL) levels of IL-12, but 2- to 10-fold lower serum levels compared to systemic administration. Whereas both mucosal and systemic IL-12 inhibited BAL eosinophil recruitment at high doses (100-1000 ng), only mucosal IL-12 was effective at low doses (1-10 ng). Mucosal, but not systemic, administration of 1000 ng of IL-12 increased interferon (IFN)-gamma expression in BAL cells. In a model of ongoing eosinophilic inflammation, when mucosal or systemic IL-12 doses were initiated prior to peak eosinophilia, further eosinophil recruitment was inhibited. However, when IL-12 treatment was initiated after peak eosinophil recruitment occurred, recovery from eosinophilic inflammation was not facilitated. Our findings are the first to demonstrate that locally administered IL-12 inhibits eosinophil recruitment at 100-fold lower doses than systemic IL-12. The most likely mechanism of this enhanced inhibitory activity is a sustained increase in lung levels of IL-12 that augments IFN-gamma production from BAL cells. We suggest that future studies should evaluate the efficacy of low doses of nebulized IL-12 in inhibiting eosinophilic lung inflammation in asthma.

Investigators disagree on the amount of hydrogen peroxide (H2O2) released by resting and stimulated alveolar macrophages. The method commonly used to measure H2O2 release involves horseradish peroxidase (HRP)-catalyzed oxidation of scopoletin by H2O2. We describe an artifact in this method that may explain the seemingly inconsistent data reported by other investigators. Release of H2O2 and luminol-catalyzed chemiluminescence are stimulated in rat alveolar macrophages by type II HRP at concentrations normally used in the HRP-scopoletin method. The amount of H2O2 released depends upon the length of time the cells are preincubated at 37.5 degrees C and the time at which type II HRP is added. After stimulation with type II HRP, the cells do not release additional H2O2 upon exposure to zymosan particles. Myeloperoxidase, an alternative catalyst to type II HRP, does not stimulate H2O2 release and, therefore, can be used to measure H2O2 release from rat alveolar macrophages. Using myeloperoxidase, resting H2O2 release is negligible; after zymosan stimulation, 6.14 (+/- 0.87) X 10(-6) nmoles/cell X 10 min is released. In addition, more pure HRP preparations (types VI, VII, VIII, and IX) do not stimulate alveolar macrophages to release H2O2 and can be used to monitor zymosan-induced H2O2 release. As our data indicate that type II HRP stimulates H2O2 release from rat and guinea pig alveolar macrophages, it is not the catalyst of choice for this assay. In conclusion, our data explain the conflicting results found in the literature and indicate that rat alveolar macrophages release minimal amounts of H2O2 at rest and can be stimulated by zymosan.