LS-2-P-1475 Artistic compositions by surfactant in axolotl lung alveoli

Lung surfactant is a mixture of dipalmitoylphosphatidylcholine and specific proteins (SP-A, -B, -C, and -D) that covers the interior lung alveoli epithelium; it is synthesized by type II alveolar cells and packaged in membrane-bound lamellar inclusion bodies. By lowering interfacial tension, lung surfactant insures a negligible work of breathing and uniform lung inflation [1]. This paper reports the fine structure of surfactant in the lung of axolotlt Ambystoma mexicanum, revealing amazing pictures that evoke artistic compositions but that also show elaborate organizational patterns.

8 male and female adult axolotls (~ 2 years old) were killed after anesthesia in tricaine methanesulphonate. Lung portions were removed, immediately diced into 1-mm pieces, doubly fixed with glutaraldehyde (2.5%)-osmium (1%) at pH 7.4, dehydrated in ethanol and embedded in Epon. Thin sections were collected on 200-mesh grids, doubly stained with uranyl acetate and lead citrate, and examined in Philips EM300, at 60 KV accelerating voltage and 50 μm objective aperture.

Three predominant morphological types were observed.

Type I inclusions or tubular myelin (average size 3.7 x 1.7 μm) showed square to rectangular grid patterns, strikingly resembling a fishing net organization (Fig. 1). The lattice appeared to be formed by membranous elements of ~ 6 nm diameter, crossing in two directions perpendicular to each other. Each unit (square or rectangle) shared its walls (membranous elements) with four neighbors, had an internal perimeter of ~ 17 x 27 nm, and measured ~ 27 x 36 nm for center to center spacing of its walls. A centrally located dense line of 2 nm diameter with a fuzzy contour crossed the middle section of each unit longitudinally. High resolution revealed a globular structure of the membranous elements (Fig. 1, inset, arrows).

Type II inclusions consisted of arrays of osmiophilic lamellar bodies displaying concentric whorls of lamellae reminding of spider webs. However, the overall appearance of such inclusions resembled representations of an owl’s head (Fig. 2 A, B) ranging from 1 x 0.8 to 3.1 x 2.9 μm.

Type III consisted of osmiophile-dense granules of average size 4.7 x 1.8 μm (Fig. 3).

Recent research indicated that lung surfactants have micro-nano scale organizations or domains that play a critical role in functioning of membrane proteins, phase segregation and liquid ordering of soft material in two dimensions [2]. The structures reported here revealed micro scale as well as micro-nano scale organization (Fig. 1, inset) in axolotl lung surfactant.

References

[1] Braun, A., et al. (2007) Biophys J. 93:123-139.

[2] Nag, K. et al. (2007) Modern Research and Educational Topics in Microscopy (Mendez-Vilas A. and Diaz, J., eds), Badajoz: Formatex; 483-490.