Phase transitions of the pulmonary surfactant film at the perfluorocarbon-water interface

Abstract

Pulmonary surfactant is a lipid-protein complex that forms a thin film at the air-water surface of the lungs. This surfactant film defines the elastic recoil and respiratory mechanics of the lungs. One generally accepted rationale of using oxygenated perfluorocarbon (PFC) as a respiratory medium in liquid ventilation is to take advantage of its low surface tensions (14-18 mN/m), which was believed to make PFC an ideal replacement of the exogenous surfactant. Compared with the extensive studies of the phospholipid phase behavior of the pulmonary surfactant film at the air-water surface, its phase behavior at the PFC-water interface is essentially unknown. Here, we reported the first detailed biophysical study of phospholipid phase transitions in two animal-derived natural pulmonary surfactant films, Infasurf and Survanta, at the PFC-water interface using constrained drop surfactometry. Constrained drop surfactometry allows in situ Langmuir-Blodgett transfer from the PFC-water interface, thus permitting direct visualization of lipid polymorphism in pulmonary surfactant films using atomic force microscopy. Our data suggested that regardless of its low surface tension, the PFC cannot be used as a replacement of pulmonary surfactant in liquid ventilation where the air-water surface of the lungs is replaced with the PFC-water interface that features an intrinsically high interfacial tension. The pulmonary surfactant film at the PFC-water interface undergoes continuous phase transitions at surface pressures less than the equilibrium spreading pressure of 50 mN/m and a monolayer-to-multilayer transition above this critical pressure. These results provided not only novel biophysical insight into the phase behavior of natural pulmonary surfactant at the oil-water interface but also translational implications into the further development of liquid ventilation and liquid breathing techniques.

Figure 1

(A) Lipid and protein compositions (wt %) of two animal-derived pulmonary surfactants, Infasurf and Survanta. DPPC, dipalmitoyl phosphatidylcholine; PL, phospholipid; PC, phosphatidylcholine; SP, surfactant protein. (B) Schematic of constrained drop surfactometry for studying pulmonary surfactant film at the perfluorocarbon (PFC)-water interface using a water-in-oil configuration. Since water is lighter than PFC, the water droplet appears as a pendant bubble. Interfacial tension of pulmonary surfactant film-covered droplet is determined in real time photographically from the shape of the droplet using closed-loop axisymmetric drop shape analysis. The pulmonary surfactant film can be Langmuir-Blodgett transferred from the PFC-water interface to a solid substrate under controlled interfacial pressure. To see this figure in color, go online.

Figure 2

Compression isotherms of (A) Infasurf and (B) Survanta films at the PFC-water interface obtained with the constrained drop surfactometry at room temperature. Insets demonstrate the correlations between the interfacial pressure and the shape of the droplet. All compression isotherms were repeated three times to demonstrate reproducibility. Compression isotherms of the Infasurf film can be divided into three regions: region I: monolayer; region II: monolayer-to-multilayer transition; and region III: multilayer. To see this figure in color, go online.

Figure 3

Lateral structure and topography of the Infasurf film at the PFC-water interface at four characteristic interfacial pressures, i.e., 30, 40, 50, and 53 mN/m. All AFM images in the top row have the same scanning area of 20 × 20 μm. The z range is 5 nm for all monolayers and 20 nm for multilayers. AFM images in the middle row show zoomed-in images indicated by the white boxes. AFM images in the bottom row show the three-dimensional rendering of the zoomed-in images. Double-headed white arrows indicate the lateral size of domains, and single-headed black arrows indicate the height of structures. Inset of the AFM image at 50 mN/m shows the nanodomains. Inset of the AFM image at 53 mN/m shows the multilayered protrusions. To see this figure in color, go online.

Figure 4

Lateral structure and topography of the Survanta film at the PFC-water interface at five characteristic interfacial pressures, i.e., 10, 20, 30, 40, and 55 mN/m. AFM images in the top row have the same scanning area of 50 × 50 μm, except that the scanning area at 55 mN/m is 20 × 20 μm. The z range is 5 nm for all monolayers and 20 nm for multilayers. AFM images in the middle row show zoomed-in images indicated by the white boxes. AFM images in the bottom row show the three-dimensional rendering of the zoomed-in images. Double-headed white arrows indicate the lateral size of domains, and single-headed black arrows indicate the height of structures. Inset of the AFM image at 55 mN/m shows the multilayered protrusions. To see this figure in color, go online.

Figure 5

Lateral diameter (μm) of the microdomains and the surface area coverage (%) of combined microdomains and nanodomains in the (A) Infasurf and (B) Survanta monolayers. ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001. To see this figure in color, go online.