Exposure to polymers reverses inhibition of pulmonary surfactant by serum, meconium, or cholesterol in the captive bubble surfactometer

Abstract

Dysfunction of pulmonary surfactant in the lungs is associated with respiratory pathologies such as acute respiratory distress syndrome or meconium aspiration syndrome. Serum, cholesterol, and meconium have been described as inhibitory agents of surfactant's interfacial activity once these substances appear in alveolar spaces during lung injury and inflammation. The deleterious action of these agents has been only partly evaluated under physiologically relevant conditions. We have optimized a protocol to assess surfactant inhibition by serum, cholesterol, or meconium in the captive bubble surfactometer. Specific measures of surface activity before and after native surfactant was exposed to inhibitors included i), film formation, ii), readsorption of material from surface-associated reservoirs, and iii), interfacial film dynamics during compression-expansion cycling. Results show that serum creates a steric barrier that impedes surfactant reaching the interface. A mechanical perturbation of this barrier allows native surfactant to compete efficiently with serum to form a highly surface-active film. Exposure of native surfactant to cholesterol or meconium, on the other hand, modifies the compressibility of surfactant films though optimal compressibility properties recover on repetitive compression-expansion cycling. Addition of polymers like dextran or hyaluronic acid to surfactant fully reverses inhibition by serum. These polymers also prevent surfactant inhibition by cholesterol or meconium, suggesting that the protective action of polymers goes beyond the mere enhancement of interfacial adsorption as described by depletion force theories.

Figure 1

CBS model to test surfactant inhibition by serum. (a) Sequential images of a surfactant sample immediately before (t = 0) and at different times after injection underneath an air bubble at the CBS. (b) Fluorescence image of a serum sample doped with a trace of rhodamine-labeled albumin, injected underneath the bubble at the CBS.

Figure 2

Interfacial adsorption of native surfactant in the absence or presence of serum. Initial (left panel) and post-expansion (right panel) adsorption kinetics of surfactant (NS) (black dots), serum alone (white dots), NS 10 mg/ml applied underneath the bubble surface coated with a preformed serum layer (triangles), and NS 20 mg/ml applied underneath the bubble surface coated with serum (squares), all at 37°C. Data are mean ± SD after averaging data from three experiments.

Figure 3

Compression-expansion isotherms of native surfactant films in the absence or presence of serum. Q-static (upper panels) and dynamic (lower panels) compression/expansion isotherms from surfactant films formed upon injection of NS at 10 or 20 mg/mL phospholipid, in the absence or in the presence of a preformed serum layer. A representative experiment is shown here after repeating three independent experiments with each sample.

Figure 4

Interfacial adsorption of native surfactant in the absence or presence of inhibitors and polymers. (a) Minimal surface tension upon 5 min of initial (black bars) or post-expansion (gray bars) adsorption of a native surfactant sample combined or not with the indicated polymers, injected under a clean or a serum-coated bubble. (b) Minimal surface tension upon initial or post-expansion adsorption of surfactant in the absence or presence of meconium, cholesterol, and/or polymers. Data are mean ± SD after averaging three independent experiments from each sample.

Figure 5

Quasi-static compression-expansion cycling isotherms for surfactant injected under the bubble at the CBS, in the absence or in the presence of serum, meconium, or cholesterol, combined or not with HA or dextran. A representative experiment is shown in each panel, after three repetitions from each sample.

Figure 6

Dynamic compression-expansion cycling isotherms for surfactant injected under the bubble at the CBS, in the absence or in the presence of serum, meconium, or cholesterol, combined or not with HA or dextran. A representative experiment is shown in each panel, after three repetitions from each sample.