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Serial block-face scanning electron microscopy of erythrocytes protruding through the human placental syncytiotrophoblast.

ABSTRACT: The syncytiotrophoblast forms a continuous barrier between the maternal and fetal circulations. Here we present a serial block-face scanning electron microscopy (SBFSEM) study, based on a single image stack, showing pooling of fetal blood underneath a region of stretched syncytiotrophoblast that has become detached from the basement membrane. Erythrocytes are protruding from discrete holes in the syncytiotrophoblast suggesting that, under specific circumstances, the syncytiotrophoblast may be permeable to fetal cells. This observation represents a pathological process but it poses questions about the physical properties and permeability of the syncytiotrophoblast and may represent an early stage in the formation of fibrin deposits in areas of syncytial denudation. This study also illustrates how the 3D images generated by SBFSEM allow the interpretation of structures that could not be understood from a single histological section.

SUBMITTER: Palaiologou E

PROVIDER: S-EPMC5603844 | biostudies-literature | 2017 Oct

REPOSITORIES: biostudies-literature

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Serial block-face scanning electron microscopy of erythrocytes protruding through the human placental syncytiotrophoblast.

Palaiologou Eleni E Goggin Patricia P Chatelet David S DS Lofthouse Emma M EM Torrens Christopher C Sengers Bram G BG Cleal Jane K JK Page Anton A Lewis Rohan M RM

Journal of anatomy 20170717 4

The syncytiotrophoblast forms a continuous barrier between the maternal and fetal circulations. Here we present a serial block-face scanning electron microscopy (SBFSEM) study, based on a single image stack, showing pooling of fetal blood underneath a region of stretched syncytiotrophoblast that has become detached from the basement membrane. Erythrocytes are protruding from discrete holes in the syncytiotrophoblast suggesting that, under specific circumstances, the syncytiotrophoblast may be pe ...[more]

PMID: 28714092

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Project description:The placental microvasculature is a conduit for fetal blood allowing solute exchange between the mother and the fetus. Serial block-face scanning electron microscopy (SBF SEM) allows ultrastructure to be viewed in three dimensions and provides a new perspective on placental anatomy. This study used SBF SEM to study endothelial cells within the human placental microvasculature from uncomplicated pregnancies. Term human placental villi were aldehyde-fixed and processed for imaging by SBF SEM. Manual segmentation was carried out on a terminal villous capillary and an intermediate villous arteriole and venule. Twenty-seven SBF SEM stacks from terminal villi were analysed using stereological approaches to determine the volumes of microvascular components and the proportions of pericyte coverage. SBF SEM analysis of capillary endothelial cells revealed the presence of interendothelial protrusions (IEPs) originating from the donor cell at the endothelial junction and forming deep thin projections up to 7 μm into the adjacent endothelial cells. IEP density was estimated to be in the order of 35 million cm-3 placental tissue. Pericytes cover 15% of the fetal capillary surface area in terminal villi. In comparison, the cytotrophoblast covered 24% of the syncytiotrophoblast basal membrane. A trans-endothelial channel was observed in a region of the vasculo-syncytial capillary. Pericyte coverage was extensive in both arteriole and venule. Three-dimensional imaging of the placental microvasculature identified novel ultrastructural features and provided an insight into factors that may influence capillary permeability and placental function. We hypothesise that the IEPs may allow mechanosensing between adjacent endothelial cells to assist in the maintenance of vessel integrity. The numbers of endothelial junctions, the presence of trans-endothelial channels and the extent of pericyte coverage all provide an insight into the factors determining capillary permeability.

Project description:Serial block face scanning electron microscopy (SBF-SEM) is a relatively new technique that allows the acquisition of serially sectioned, imaged and digitally aligned ultrastructural data. There is a wealth of information that can be obtained from the resulting image stacks but this presents a new challenge for researchers - how to computationally analyse and make best use of the large datasets produced. One approach is to reconstruct structures and features of interest in 3D. However, the software programmes can appear overwhelming, time-consuming and not intuitive for those new to image analysis. There are a limited number of published articles that provide sufficient detail on how to do this type of reconstruction. Therefore, the aim of this paper is to provide a detailed step-by-step protocol, accompanied by tutorial videos, for several types of analysis programmes that can be used on raw SBF-SEM data, although there are more options available than can be covered here. To showcase the programmes, datasets of skeletal muscle from foetal and adult guinea pigs are initially used with procedures subsequently applied to guinea pig cardiac tissue and locust brain. The tissue is processed using the heavy metal protocol developed specifically for SBF-SEM. Trimmed resin blocks are placed into a Zeiss Sigma SEM incorporating the Gatan 3View and the resulting image stacks are analysed in three different programmes, Fiji, Amira and MIB, using a range of tools available for segmentation. The results from the image analysis comparison show that the analysis tools are often more suited to a particular type of structure. For example, larger structures, such as nuclei and cells, can be segmented using interpolation, which speeds up analysis; single contrast structures, such as the nucleolus, can be segmented using the contrast-based thresholding tools. Knowing the nature of the tissue and its specific structures (complexity, contrast, if there are distinct membranes, size) will help to determine the best method for reconstruction and thus maximize informative output from valuable tissue.

Project description:The cornea is the most highly innervated tissue in the body. It is generally accepted that corneal stromal nerves penetrate the epithelial basal lamina giving rise to intra-epithelial nerves. During the course of a study wherein we imaged corneal nerves in mice, we observed a novel neuronal-epithelial cell interaction whereby nerves approaching the epithelium in the cornea fused with basal epithelial cells, such that their plasma membranes were continuous and the neuronal axoplasm freely abutted the epithelial cytoplasm. In this study we sought to determine the frequency, distribution, and morphological profile of neuronal-epithelial cell fusion events within the cornea. Serial electron microscopy images were obtained from the anterior stroma in the paralimbus and central cornea of 8-10 week old C57BL/6J mice. We found evidence of a novel alternative behavior involving a neuronal-epithelial interaction whereby 42.8% of central corneal nerve bundles approaching the epithelium contain axons that fuse with basal epithelial cells. The average surface-to-volume ratio of a penetrating nerve was 3.32, while the average fusing nerve was smaller at 1.39 (p ? 0.0001). Despite this, both neuronal-epithelial cell interactions involve similarly sized discontinuities in the basal lamina. In order to verify the plasma membrane continuity between fused neurons and epithelial cells we used the lipophilic membrane tracer DiI. The majority of corneal nerves were labeled with DiI after application to the trigeminal ganglion and, consistent with our ultrastructural observations, fusion sites recognized as DiI-labeled basal epithelial cells were located at points of stromal nerve termination. These studies provide evidence that neuronal-epithelial cell fusion is a cell-cell interaction that occurs primarily in the central cornea, and fusing nerve bundles are morphologically distinct from penetrating nerve bundles. This is, to our knowledge, the first description of neuronal-epithelial cell fusion in the literature adding a new level of complexity to the current understanding of corneal innervation.

Dataset Information

Serial block-face scanning electron microscopy of erythrocytes protruding through the human placental syncytiotrophoblast.

Publications

Serial block-face scanning electron microscopy of erythrocytes protruding through the human placental syncytiotrophoblast.

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