Weibel's hypothetical 3D model in 1966 provided first ultrastructural details into tubular myelin (TM), a unique, complex surfactant subtype found in the hypophase of the alveolar lining layer. Although initial descriptions by electron microscopy (EM) were already published in the 1950s, a uniform morphological differentiation from other intraalveolar surfactant subtypes is still missing and potential structure-function relationships remain enigmatic. Technical developments in volume EM methods now allow a more detailed reinvestigation. To address unanswered ultrastructural questions, we analyzed ultrathin sections of humanized SP-A1/SP-A2 co-expressing mouse as well as human lung samples by conventional transmission EM. We combined these 2D information with 3D analysis of single- and dual-axis electron tomography of serial sections for high z-resolution (in a range of a few nm) and extended volumes of up to 1 µm total z-information. This study reveals that TM constitutes a heterogeneous surfactant organization mainly comprised of distorted parallel membrane planes with local intersections, which are distributed all over the TM substructure. These intersecting membrane planes form, among other various polygons, the well-known 2D "lattice", respectively 3D quadratic tubules, which in many analyzed spots of human alveoli appear to be less abundant than also observed non-concentric 3D lamellae. The additional application of serial section electron tomography to conventional transmission EM demonstrates a high heterogeneity of TM membrane networks, which indicates dynamic transformations between its substructures. Our method provides an ideal basis for further in and ex vivo structural analyses of surfactant under various conditions at nanometer scale.
- Lettau, M.
- Timm, S.
- Dittmayer, C.
- Lopez-Rodriguez, E.
- Ochs, M.
Keywords
- 3D electron microscopy
- alveolar lining layer
- serial section electron tomography
- surfactant
- tubular myelin
