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Pulmonary surfactant stabilizes the lung by reducing surface tension at the air-water interface of the alveoli. Surfactant is present in the lung in a number of morphological forms, including tubular myelin (TM). TM is composed of unusual 40 x 40 nm square elongated proteolipid tubes. Atomic force microscopy (AFM) was performed on polymer-embedded Lowicryl and London Resin-White (LR-White) unstained thin sections. AFM was used in imaging regions of the sections where TM was detected by transmission electron microscopy (EM) of corresponding stained sections. Tapping- and contact-mode AFM imaging of the unstained sections containing TM indicated a highly heterogeneous surface topography with height variations ranging from 10 to 100 nm. In tapping-mode AFM, tubular myelin was seen as hemispherical protrusions of 30-70 nm in diameter, with vertical dimensions of 5-8 nm. In contact-mode AFM and with phase imaging using a sharper (>10 nm nominal radius) probe, square open-ended tubes which resembled typical electron micrographs of such regions were observed. The cross-hatch structures observed inside the tubes using EM were not observed using AFM, although certain multilobe structures and topographic heterogeneity were detected inside some tubes. Other regions of multilamellar bodies and some regions where such bilayer lamella appear to fuse with the tubes were found in association with TM using AFM. EM of acetone-delipidated tubes in LR-White revealed rectangular tubular cores containing cross-hatched structures, presumably protein skeletons. AFM surface topography of these regions showed hollow depressions at positions at which the protein was anticipated instead of the protrusions seen in the lipid-containing sections. Gold-labeled antibody to surfactant protein A was found associated somewhat randomly within the regions containing the protein skeletons. The topography of the gold particles was observed as sharp peaks in contact-mode AFM. This study suggests a method for unambiguous detection of three-dimensional nanotubes present in low abundance in a biological macromolecular complex. Only limited detection of proteins and lipids in surfaces of embedded tubular myelin was possible. EM and AFM imaging of such unusual biological structures may suggest unique lipid-protein associations and arrangements in three dimensions., (Copyright 1999 Academic Press.)

Surfactant protein A (SP-A) is an octadecameric hydrophilic glycoprotein and is the major protein component of pulmonary surfactant. This protein complex plays several roles in the body, such as regulation of surfactant secretion, recycling and adsorption of surfactant lipids, and non-serum-induced immune response. Many of SP-A's activities are dependent upon the presence of cations, especially calcium. Here, we have studied in vitro the effect of cations on the interaction of purified bovine SP-A with phospholipid vesicles made of dipalmitoylphosphatidylcholine and unsaturated phosphatidylcholine. We have found that SP-A octadecamers exist in an "opened-bouquet" conformation in the absence of cations and interact with lipid membranes via one or two globular headgroups. Calcium-induced structural changes in SP-A lead to the formation of a clearly identifiable stem in a "closed-bouquet" conformation. This change, in turn, seemingly results in all of SP-A's globular headgroups interacting with the lipid membrane surface and with the stem pointing away from the membrane surface. These results represent direct evidence that the headgroups of SP-A (comprising carbohydrate recognition domains), and not the stem (comprising the amino-terminus and collagen-like region), interact with lipid bilayers. Our data support models of tubular myelin in which the headgroups, not the tails, interact with the lipid walls of the lattice., (Copyright 1998 Academic Press.)

The effects of short-term ozone exposure on the lung function and surface activity of surfactant subtypes isolated from rat lung lavage were studied. Rats were exposed to 0.8 ppm ozone for 2 or 12 hr. The surface activity of surfactant was affected by ozone exposure, whereas distinct morphological changes in bronchoalveolar lavage or in the surfactant subtypes were not observed. Adsorption experiments indicated that bronchoalveolar lavage from rats exposed for 12 hr to ozone remained at lower equilibrium surface pressures than lavage from control rats. These observations suggest interference of inflammatory proteins with the surface film. Extracted surfactant, containing only lipids and surfactant proteins B and C, had a decreased adsorption rate after ozone exposure. These results suggest that the activity of one or both of the hydrophobic surfactant proteins (SP-B and SP-C) was affected by ozone.