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This study presents the results of investigation of the micromorphology and variability of Sisymbrium polymorphum seeds collected in 49 localities in the core range and isolated populations of Central Europe, Eastern Europe and Central Asia. In addition, we compared the ultrastructure of the seeds of S. polymorphum with that of the closely-related species S. loeselii and S. linifolium. The seeds were measured with a stereoscopic microscope and a biometric programme (ImageJ) and micromorphological studies were performed by scanning electron microscopy (SEM). The seed variability showed intraspecific stability of the ultrastructure and low variability of metric features within the studied range. Studied species showed differences in the ultrastructure, which will be valuable for diagnostic purposes. We present and describe, for the first time, the ultrastructure of S. polymorphum seeds. [ABSTRACT FROM AUTHOR]

There are many studies on the biology, ecology and infrageneric classification of the cosmopolitan orchid genus Habenaria. However, there are few data on the seed morphology of species in this genus. The aim of this study was to analyse the seed morphology of Habenaria and related taxa (111 species in total). The species of Habenaria s.l. studied are mostly characterized by a scobiform seed, but the form of individual seed coat cells varies between and among some species. In most of the species analysed, the cell walls are arranged mostly along the main axis of a seed. The size of the embryos and the volume of seeds and embryos also varied in Habenaria. The periclinal wall of the cells in the seed coat of most of the taxa examined is smooth, but there are several species with distinct ornamentation on this wall (slanting ridges, transverse ridges, reticulate ridges). The periclinal wall could be used in taxonomic delimitations, especially when combined with additional features such as seed size, shape and arrangement of the cells in the seed coat. Although many of the morphological groups in Habenaria s.l. were included in this study, no qualitative seed characters that could be helpful in establishing an alternative infrageneric division of this taxon were previously recognized. [ABSTRACT FROM AUTHOR]

The egg plays a pivotal role in the reproduction of our species. Nevertheless, its fundamental biology remains elusive. Transmission electron microscopy is traditionally used to inspect the ultrastructure of female gametes. However, two-dimensional micrographs contain only fragmentary information about the spatial organization of the complex oocyte cytoplasm. Here, we employed the Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) to explore human oocyte intracellular morphology in three dimensions (3D). Volume reconstruction of generated image stacks provided an unprecedented view of ooplasmic architecture. Organelle distribution patterns observed in nine donor oocytes, representing three maturational stages, documented structural changes underlying the process by which the egg acquires developmental competence. 3D image segmentation was performed to extract information about distinct organelle populations, and the following quantitative analysis revealed that the mitochondrion occupies ∼ 4.26% of the maturing oocyte cytoplasm. In summary, this proof-of-concept study demonstrates the potential of large volume electron microscopy to study rare samples of delicate female gametes and paves the way for applying the FIB-SEM technique in human oocyte research.

Focused-ion beam-scanning electron microscopic (FIB-SEM) tomography enables easier acquisition of a series of ultrastructural, sectional images directly from resin-embedded biological samples. In this study, to clarify the three-dimensional (3D) architecture of glomerular endothelial cells (GEnCs) in adult rats, we manually extracted GEnCs from serial FIB-SEM images and reconstructed them on an Amira reconstruction software. The luminal and basal surface structures were clearly visualized in the reconstructed GEnCs, although only the luminal surface structures could be observed by conventional SEM. The luminal surface visualized via the reconstructed GEnCs was quite similar to that observed through conventional SEM, indicating that 3D reconstruction could be performed with high accuracy. Thus, we successfully described the 3D architecture of normal GEnCs in adult rats more clearly and precisely than ever before. The GEnCs were found to consist of three major subcellular compartments, namely, the cell body, cytoplasmic ridges, and sieve plates, in addition to two associated subcellular compartments, namely, the globular protrusions and reticular porous structures. Furthermore, most individual GEnCs made up a “seamless” tubular shape, and some of them formed an autocellular junction to make up a tubular shape. FIB-SEM tomography with reconstruction is a powerful approach to better understand the 3D architecture of GEnCs. Moreover, the morphological information revealed in this study will be valuable for the 3D pathologic evaluation of GEnCs in animal and human glomerular diseases and the structural analysis of developmental processes in the glomerular capillary system.

The excretory system produces urine by ultrafiltration via a filtration epithelium. Podocytes are widely found as filtration epithelial cells in eucoelomates. In some animal taxa, including insects and crustaceans, nephrocytes serve to separate toxic substances from the body fluid, in addition to podocytes. Drosophila nephrocytes have been recently utilized as a model system to study podocyte function and disease. However, functionality and cellular architecture are strikingly different between Drosophila nephrocytes and eucoelomate podocytes, and the phylogenetic relationship between these cells remains enigmatic. In this study, using focused-ion beam-scanning electron microscopy (FIB-SEM) tomography, we revealed three-dimensional architecture of decapod nephrocytes with unprecedented accuracy—they filled an enormous gap, which can be called "missing link," in the evolutionary diversity of podocytes and nephrocytes. Thus, we concluded that nephrocytes are part of the spectrum of filtration epithelial diversity in animal phylogeny. [ABSTRACT FROM AUTHOR]

Impatiens capensis is an annual plant native to eastern North America that is currently spreading across Europe. In Poland, due to this plant's rapid spread in the secondary range and high competitiveness in relation to native species, it is considered a locally invasive species. The microstructure of seeds is an important tool for solving various taxonomic problems and also provides data useful for determining the impact of various environmental factors on the phenotypic variability of species. This issue is particularly important in regard to invasive species which occupy a wide range of habitats in the invaded range. There are few reports on seed size and thus far no descriptions of the seed ultrastructure of I. capensis in the analyzed literature. We present new data on the seed morphology of I. capensis growing in different habitats and conditions in the secondary range of the species. The studied populations differed significantly in each of the investigated traits (seed length, width, circumference, area, roundness, and mass). Our findings showed that anthropogenic disturbances in habitats and some soil parameters (presence of carbonates, potassium, loose sand, and moisture) were statistically significant with various seed sizes and morphology in the studied populations of I. capensis. Moreover, our studies showed maximum seed length (5.74 mm) and width (3.21 mm) exceeding those values given in the available literature. For the first time, we also provide a detailed SEM study of the ultrastructure of the seed coat of I. capensis. There are two types of epidermal cells on the seeds: (a) between the ribs (elongated with straight anticlinal walls, slightly concave outer periclinal walls, and micropapillate secondary sculpture on the edges with anticyclic walls), and (b) on the ribs (isodiametric cells with straight anticlinal walls and concave outer periclinal walls). Unlike the variability of size and weight of seeds, the coat ornamentation has turned out to be a steady feature within the studied secondary range of I. capensis. [ABSTRACT FROM AUTHOR]

Elastic fibres constitute an important component of the extracellular matrix and currently are the subject of intensive study in order to elucidate their assembly, function and involvement in cell–matrix interactions and disease. However, few studies to date have investigated the 3D architecture of the elastic fibre system in bulk tissue. We describe a protocol for preparation of tissue samples, including primary fixation and backscatter electron contrast-enhancement steps, through dehydration into stable resin-embedded blocks for volume electron microscopy. The use of low molecular weight tannic acid and alcoholic lead staining are critical stages in this procedure. Block preparation by ultramicrotomy and evaporative metal coating prior to microscopical examination are also described. We present images acquired from serial block face scanning electron microscopy of cornea and aorta showing target structures clearly differentiated from cells and other matrix components. The processing method imparts high contrast to fibrillin-containing elastic fibres, thus facilitating their segmentation and rendering into 3D reconstructions by image analysis software from large serial image datasets.

Nephrocytes are similar in structure to podocytes and play a role in the isolation of toxic substances from hemolymph in insects. Drosophila melanogaster nephrocytes have recently been used to study podocyte function and disease. However, the three-dimensional ultrastructure of nephrocytes is not clearly understood because their surrounding basement membrane makes it difficult to observe using conventional scanning electron microscopy. We reconstructed the three-dimensional ultrastructure of Drosophila pericardial nephrocytes using serial focused-ion beam/scanning electron microscopy (FIB/SEM) images. The basal surfaces were occupied by foot processes and slit-like spaces between them. The slit-like spaces corresponded to the podocyte filtration slits and were formed by longitudinal infolding/invagination of the basal plasma membrane. The basal surface between the slit-like spaces became the foot processes, which ran almost linearly, and had a "washboard-like" appearance. Both ends of the foot processes were usually anastomosed to neighboring foot processes and thus free ends were rarely observed. We demonstrated that FIB/SEM is a powerful tool to better understand the three-dimensional architecture of nephrocytes. [ABSTRACT FROM AUTHOR]

The first electron microscope was constructed in 1931. Several decades later, techniques were developed to allow the first ultrastructural analysis of microglia by transmission electron microscopy (EM). In the 50 years that followed, important roles of microglia have been identified, specifically due to the ultrastructural resolution currently available only with EM. In particular, the addition of electron-dense staining using immunohistochemical EM methods has allowed the identification of microglial cell bodies, as well as processes, which are difficult to recognize in EM, and to uncover their complex interactions with neurons and synapses. The ability to recognize neuronal, astrocytic, and oligodendrocytic compartments in the neuropil without any staining is another invaluable advantage of EM over light microscopy for studying intimate cell–cell contacts. The technique has been essential in defining microglial interactions with neurons and synapses, thus providing, among other discoveries, important insights into their roles in synaptic stripping and pruning via phagocytosis of extraneous synapses. Recent technological advances in EM including serial block-face imaging and focused-ion beam scanning EM have also facilitated automated acquisition of large tissue volumes required to reconstruct neuronal circuits in 3D at nanometer-resolution. These cutting-edge techniques which are now becoming increasingly available will further revolutionize the study of microglia across stages of the lifespan, brain regions, and contexts of health and disease. In this mini-review, we will focus on defining the distinctive ultrastructural features of microglia and the unique insights into their function that were provided by EM.

The first electron microscope was constructed in 1931. Several decades later, techniques were developed to allow the first ultrastructural analysis of microglia by transmission electron microscopy (EM). In the 50 years that followed, important roles of microglia have been identified, specifically due to the ultrastructural resolution currently available only with EM. In particular, the addition of electron-dense staining using immunohistochemical EM methods has allowed the identification of microglial cell bodies, as well as processes, which are difficult to recognize in EM, and to uncover their complex interactions with neurons and synapses. The ability to recognize neuronal, astrocytic, and oligodendrocytic compartments in the neuropil without any staining is another invaluable advantage of EM over light microscopy for studying intimate cell--cell contacts. The technique has been essential in defining microglial interactions with neurons and synapses, thus providing, among other discoveries, important insights into their roles in synaptic stripping and pruning via phagocytosis of extraneous synapses. Recent technological advances in EM including serial block-face imaging and focused-ion beam scanning EM have also facilitated automated acquisition of large tissue volumes required to reconstruct neuronal circuits in 3D at nanometer-resolution. These cutting-edge techniques which are now becoming increasingly available will further revolutionize the study of microglia across stages of the lifespan, brain regions, and contexts of health and disease. In this mini-review, we will focus on defining the distinctive ultrastructural features of microglia and the unique insights into their function that were provided by EM. [ABSTRACT FROM AUTHOR]

This study presents the results of investigation of the micromorphology and variability of Sisymbriumpolymorphum seeds collected in 49 localities in the core range and isolated populations of Central Europe, Eastern Europe and Central Asia. In addition, we compared the ultrastructure of the seeds of S. polymorphum with that of the closely-related species S.loeselii and S.linifolium. The seeds were measured with a stereoscopic microscope and a biometric programme (ImageJ) and micromorphological studies were performed by scanning electron microscopy (SEM). The seed variability showed intraspecific stability of the ultrastructure and low variability of metric features within the studied range. Studied species showed differences in the ultrastructure, which will be valuable for diagnostic purposes. We present and describe, for the first time, the ultrastructure of S.polymorphum seeds., (Agnieszka Rewicz, Weronika Torbicz, Liudmyla Zavialova, Oksana Kucher, Myroslav V. Shevera, Tomasz Rewicz, Marcin Kiedrzyński, Anna Bomanowska.)

The egg plays a pivotal role in the reproduction of our species. Nevertheless, its fundamental biology remains elusive. Transmission electron microscopy is traditionally used to inspect the ultrastructure of female gametes. However, two-dimensional micrographs contain only fragmentary information about the spatial organization of the complex oocyte cytoplasm. Here, we employed the Focused Ion Beam Scanning Electron Microscopy (FIB-SEM) to explore human oocyte intracellular morphology in three dimensions (3D). Volume reconstruction of generated image stacks provided an unprecedented view of ooplasmic architecture. Organelle distribution patterns observed in nine donor oocytes, representing three maturational stages, documented structural changes underlying the process by which the egg acquires developmental competence. 3D image segmentation was performed to extract information about distinct organelle populations, and the following quantitative analysis revealed that the mitochondrion occupies ∼ 4.26% of the maturing oocyte cytoplasm. In summary, this proof-of-concept study demonstrates the potential of large volume electron microscopy to study rare samples of delicate female gametes and paves the way for applying the FIB-SEM technique in human oocyte research., Competing Interests: JJ is employed by TESCAN ORSAY HOLDING, the manufacturer of the FIB-SEM microscopes used in this study. However, the study did not receive any funding from the industrial partner. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Trebichalská, Javůrek, Tatíčková, Kyjovská, Kloudová, Otevřel, Hampl and Holubcová.)

Elastic fibres constitute an important component of the extracellular matrix and currently are the subject of intensive study in order to elucidate their assembly, function and involvement in cell-matrix interactions and disease. However, few studies to date have investigated the 3D architecture of the elastic fibre system in bulk tissue. We describe a protocol for preparation of tissue samples, including primary fixation and backscatter electron contrast-enhancement steps, through dehydration into stable resin-embedded blocks for volume electron microscopy. The use of low molecular weight tannic acid and alcoholic lead staining are critical stages in this procedure. Block preparation by ultramicrotomy and evaporative metal coating prior to microscopical examination are also described. We present images acquired from serial block face scanning electron microscopy of cornea and aorta showing target structures clearly differentiated from cells and other matrix components. The processing method imparts high contrast to fibrillin-containing elastic fibres, thus facilitating their segmentation and rendering into 3D reconstructions by image analysis software from large serial image datasets.

Focused-ion beam-scanning electron microscopic (FIB-SEM) tomography enables easier acquisition of a series of ultrastructural, sectional images directly from resin-embedded biological samples. In this study, to clarify the three-dimensional (3D) architecture of glomerular endothelial cells (GEnCs) in adult rats, we manually extracted GEnCs from serial FIB-SEM images and reconstructed them on an Amira reconstruction software. The luminal and basal surface structures were clearly visualized in the reconstructed GEnCs, although only the luminal surface structures could be observed by conventional SEM. The luminal surface visualized via the reconstructed GEnCs was quite similar to that observed through conventional SEM, indicating that 3D reconstruction could be performed with high accuracy. Thus, we successfully described the 3D architecture of normal GEnCs in adult rats more clearly and precisely than ever before. The GEnCs were found to consist of three major subcellular compartments, namely, the cell body, cytoplasmic ridges, and sieve plates, in addition to two associated subcellular compartments, namely, the globular protrusions and reticular porous structures. Furthermore, most individual GEnCs made up a "seamless" tubular shape, and some of them formed an autocellular junction to make up a tubular shape. FIB-SEM tomography with reconstruction is a powerful approach to better understand the 3D architecture of GEnCs. Moreover, the morphological information revealed in this study will be valuable for the 3D pathologic evaluation of GEnCs in animal and human glomerular diseases and the structural analysis of developmental processes in the glomerular capillary system., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Kawasaki, Hosoyamada, Miyaki, Yamaguchi, Kakuta, Sakai and Ichimura.)