Diminished lipoxin biosynthesis in severe asthma

Abstract

Rationale and objectives: Severe asthma is characterized by increased airway inflammation that persists despite therapy with corticosteroids. It is not, however, merely an exaggeration of the eosinophilic inflammation that characterizes mild to moderate asthma; rather, severe asthma presents unique features. Although arachidonic acid metabolism is well appreciated to regulate airway inflammation and reactivity, alterations in the biosynthetic capacity for both pro- and antiinflammatory eicosanoids in severe asthma have not been determined.

Methods: Patients with severe asthma were identified according to National Heart, Lung, and Blood Institute Severe Asthma Research Program criteria. Samples of whole blood from individuals with severe or moderate asthma were assayed for biosynthesis of lipoxygenase-derived eicosanoids.

Measurements and main results: The counterregulatory mediator lipoxin A4 was detectable in low picogram amounts, using a novel fluorescence-based detection system. In activated whole blood, mean lipoxin A4 levels were decreased in severe compared with moderate asthma (0.4 [SD 0.4] ng/ml vs. 1.8 [SD 0.8] ng/ml, p=0.001). In sharp contrast, mean levels of prophlogistic cysteinyl leukotrienes were increased in samples from severe compared with moderate asthma (112.5 [SD 53.7] pg/ml vs. 64.4 [SD 24.8] pg/ml, p=0.03). Basal circulating levels of lipoxin A4 were also decreased in severe relative to moderate asthma. The marked imbalance in lipoxygenase-derived eicosanoid biosynthesis correlated with the degree of airflow obstruction.

Conclusions: Mechanisms underlying airway responses in severe asthma include underproduction of lipoxins. This is the first report of a defect in lipoxin biosynthesis in severe asthma, and suggests an alternative therapeutic strategy that emphasizes natural counterregulatory pathways in the airways.

Figure 1.

Fluorescence-based detection of lipoxin A₄ (LXA₄). A new HPLC system, coupled online to a 325-nm helium–cadmium laser to enhance fluorescence, enabled detection of LXA₄ in the low picogram range and (inset) the assay was validated with authentic LXA₄ for quantitation over 3 log orders of concentration (mean ± SEM, r² = 0.9987 with a 5-pg limit of detection). The structure of LXA₄ is shown to demonstrate the unique conjugated tetraene that is responsible for its endogenous fluorescence. Results are representative of n ⩾ 3 experiments.

Figure 2.

LXA₄ and leukotriene (LT) generation in activated whole blood from individuals with moderate and severe asthma. Samples of whole blood were obtained from healthy individuals and from subjects with clinically characterized asthma and activated (30 minutes at 37°C) with A23187. After extraction, materials were analyzed by HPLC-LIF for LXA₄ determination (a), by HPLC-PDA for LTB₄ determination (b), and by ELISA for CysLT determination (b). Results are expressed as means ± SEM (n = 9) for LXA₄ and the change in LT biosynthesis in subjects with asthma relative to the mean value in nonasthmatic volunteers (2.3 ng of LTB₄/ml, 25.5 pg of CysLTs/ml, n = 4). N.D. = not detected. *p < 0.05 for subjects with moderate asthma versus nonasthmatic subjects and **p < 0.05 for subjects with severe compared with moderate asthma.

Figure 3.

Relationship between lipid mediator generation and airflow obstruction. The mean value for (a) LXA₄ (circles) and (b) CysLT (triangles) biosynthetic capacity for each subject with asthma was compared with their FEV₁ (percent-predicted values). Values for patients with severe asthma are solid, and values for patients with moderate asthma are open.

Figure 4.

Relationship between lipoxygenase-derived eicosanoids in whole blood and airflow obstruction. The value for the ratio of LXA₄ to CysLTs (squares) in nonstimulated whole blood for each subject with asthma was compared with their FEV₁ (percent-predicted values). Values for patients with severe asthma are solid, and values for patients with moderate asthma are open.