Institute of Anatomy, University of Bern, Bern, Switzerland.
Histochem Cell Biol. 2010 Oct;134(4):319-26. doi: 10.1007/s00418-010-0736-4. Epub 2010 Sep 1.
Lamellar bodies are the storage sites for lung surfactant within type II alveolar epithelial cells. The structure-function models of lamellar bodies are based on microscopic analyses of chemically fixed tissue. Despite available alternative fixation methods that are less prone to artifacts, such as cryofixation by high-pressure freezing, the nature of the lung, being mostly air filled, makes it difficult to take advantage of these improved methods. In this paper, we propose a new approach and show for the first time the ultrastructure of intracellular lamellar bodies based on cryo-electron microscopy of vitreous sections in the range of nanometer resolution. Thus, unspoiled by chemical fixation, dehydration and contrasting agents, a close to native structure is revealed. Our approach uses perfluorocarbon to substitute the air in the alveoli. Lung tissue was subsequently high-pressure frozen, cryosectioned and observed in a cryo-electron microscope. The lamellar bodies clearly show a tight lamellar morphology. The periodicity of these lamellae was 7.3 nm. Lamellar bifurcations were observed in our cryosections. The technical approach described in this paper allows the examination of the native cellular ultrastructure of the surfactant system under near in vivo conditions, and therefore opens up prospectives for scrutinizing various theories of lamellar body biogenesis, exocytosis and recycling.
板层小体是 II 型肺泡上皮细胞内肺表面活性物质的储存场所。板层小体的结构-功能模型是基于对化学固定组织的微观分析。尽管有替代的固定方法，如高压冷冻的冷冻固定，不太容易产生伪影，但由于肺主要充满空气，因此很难利用这些改进的方法。在本文中，我们提出了一种新的方法，并首次展示了基于纳米分辨率玻璃切片的冷冻电子显微镜下细胞内板层小体的超微结构。因此，未经过化学固定、脱水和对比剂的处理，揭示了接近自然的结构。我们的方法使用全氟碳代替肺泡中的空气。随后，肺组织进行高压冷冻、冷冻切片，并在冷冻电子显微镜下观察。板层小体显示出紧密的板层形态。这些板层的周期性为 7.3nm。在我们的冷冻切片中观察到板层分支。本文描述的技术方法允许在接近体内条件下检查表面活性剂系统的天然细胞超微结构，因此为仔细研究板层小体生物发生、胞吐和再循环的各种理论开辟了前景。
