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Frontiers in Cardiovascular Medicine 2021Currently, an ultrastructural analysis of cardiovascular tissues is significantly complicated. Routine histopathological examinations and immunohistochemical staining...
Currently, an ultrastructural analysis of cardiovascular tissues is significantly complicated. Routine histopathological examinations and immunohistochemical staining suffer from a relatively low resolution of light microscopy, whereas the fluorescence imaging of plaques and bioprosthetic heart valves yields considerable background noise from the convoluted extracellular matrix that often results in a low signal-to-noise ratio. Besides, the sectioning of calcified or stent-expanded blood vessels or mineralised heart valves leads to a critical loss of their integrity, demanding other methods to be developed. Here, we designed a conceptually novel approach that combines conventional formalin fixation, sequential incubation in heavy metal solutions (osmium tetroxide, uranyl acetate or lanthanides, and lead citrate), and the embedding of the whole specimen into epoxy resin to retain its integrity while accessing the region of interest by grinding and polishing. Upon carbon sputtering, the sample is visualised by means of backscattered scanning electron microscopy. The technique fully preserves calcified and stent-expanded tissues, permits a detailed analysis of vascular and valvular composition and architecture, enables discrimination between multiple cell types (including endothelial cells, vascular smooth muscle cells, fibroblasts, adipocytes, mast cells, foam cells, foreign-body giant cells, canonical macrophages, neutrophils, and lymphocytes) and microvascular identities (arterioles, venules, and capillaries), and gives a technical possibility for quantitating the number, area, and density of the blood vessels. Hence, we suggest that our approach is capable of providing a pathophysiological insight into cardiovascular disease development. The protocol does not require specific expertise and can be employed in virtually any laboratory that has a scanning electron microscope.
PubMed: 34760942
DOI: 10.3389/fcvm.2021.739549 -
Folia Morphologica 2022Cerebral white matter consists mainly of axons surrounded by myelin sheaths, which are grouped to form association, commissural, and projection fasciculi. The aim of our...
Axonal quantification of the white matter association fasciculi in cerebral hemispheres of cow (Bos taurus), pig (Sus scrofa domesticus) and rabbit (Oryctolagus cuniculus).
BACKGROUND
Cerebral white matter consists mainly of axons surrounded by myelin sheaths, which are grouped to form association, commissural, and projection fasciculi. The aim of our work was to quantify and compare under the microscope the axons of the white matter association fasciculi in the cerebral hemispheres of cow (Bos taurus), pig (Sus scrofa domesticus) and rabbit (Oryctolagus cuniculus) indirectly by identification of their myelin sheaths.
MATERIALS AND METHODS
The samples were taken from 30 cerebral hemispheres: 10 cow, 10 pig and 10 rabbit (15 right and 15 left). They were obtained following a protocol based on the Talairach-Tournoux coordinate system for human and primate brains. The slides were stained with Luxol Fast Blue, observed by optical microscopy, and photographed at 600×. Samples were also prepared for observation in scanning transmission electron microscopy with osmium tetroxide. The myelin sheaths/axons were counted with the ImageJ software.
RESULTS
Statistically significant differences in the number of myelin sheaths per 410 μm² were found in the inferior and superior longitudinal fasciculi between the left and right hemispheres of cows, with predominance of the right hemisphere; and in the inferior occipitofrontal fasciculus of the rabbit with predominance of the left hemisphere.
CONCLUSIONS
The use of animal models for experiments in the cerebral fasciculi, especially pig, could give us a greater understanding of the behaviour of demyelinating and neurodegenerative diseases in humans.
Topics: Swine; Animals; Cattle; Female; Rabbits; Humans; White Matter; Sus scrofa; Myelin Sheath; Axons; Cerebrum
PubMed: 34750803
DOI: 10.5603/FM.a2021.0116 -
Annals of Medicine and Surgery (2012) Oct 2021Electron microscopy is a powerful tool to study biological samples at higher magnification. The higher magnifications achieved by the electron microscopes are helpful to...
Electron microscopy is a powerful tool to study biological samples at higher magnification. The higher magnifications achieved by the electron microscopes are helpful to the researchers to study surface morphology as well as cellular morphology of the samples. The blood sample surface morphology can be visualized at higher magnification by scanning electron microscope (SEM). For the examination of the blood cells at the cellular level, transmission electron microscopes (TEM) are used. In this article, we have described the step-by-step standard protocol for the preparation of blood samples for electron microscopy. The prepared blood samples can be visualized under SEM and TEM. The obtained electron micrographs of blood cells can be used for differential diagnosis of various diseases at the cellular level.
PubMed: 34691432
DOI: 10.1016/j.amsu.2021.102895 -
Nanomaterials (Basel, Switzerland) Sep 2021Composite membranes play a very important role in the separation, concentration, and purification processes, but especially in membrane reactors and membrane...
Composite membranes play a very important role in the separation, concentration, and purification processes, but especially in membrane reactors and membrane bioreactors. The development of composite membranes has gained momentum especially through the involvement of various nanoparticles, polymeric, oxide, or metal, that have contributed to increasing their reactivity and selectivity. This paper presents the preparation and characterization of an active metal nanoparticle-support polymer type composite membrane, based on osmium nanoparticles obtained in situ on a polypropylene hollow fiber membrane. Osmium nanoparticles are generated from a solution of osmium tetroxide in butyl alcohol by reduction with molecular hydrogen in a contactor with a polypropylene membrane. The composite osmium-polypropylene hollow fiber obtained membranes (Os-PPM) were characterized from the morphological and structural points of view: scanning electron microscopy (SEM), high resolution SEM (HR-SEM), energy dispersive spectroscopy analysis (EDAX), X-ray diffraction analysis (XRD), Fourier transform Infrared (FTIR) spectroscopy, thermal gravimetric analysis, and differential scanning calorimetry (TGA, DSC). The process performance was tested in a redox process of nitrophenol and 10-undecylenic (10-undecenoic) acid, as a target substance of biological or biomedical interest, in solutions of lower aliphatic alcohols in a membrane contactor with a prepared composite membrane. The characteristics of osmium nanoparticles-polypropylene hollow fiber membranes open the way to biological and biotechnological applications. These membranes do not contaminate the working environment, operate at relatively low temperatures, provide a large contact area between reactants, allow successive oxidation and reduction operations in the same module, and help to recover the reaction mass by ultrafiltration. The results obtained show that the osmium-polypropylene composite membrane allows the reduction of nitrophenol or the oxidation of 10-undecylenic acid, the conversion depending on the concentration in the lower aliphatic alcohol, the nature of the lower aliphatic alcohol, and the oxidant or reducing flow through the membrane contactor.
PubMed: 34684968
DOI: 10.3390/nano11102526 -
Membranes Aug 2021Membranes are associated with the efficient processes of separation, concentration and purification, but a very important aspect of them is the realization of a reaction...
Membranes are associated with the efficient processes of separation, concentration and purification, but a very important aspect of them is the realization of a reaction process simultaneously with the separation process. From a practical point of view, chemical reactions have been introduced in most membrane systems: with on-liquid membranes, with inorganic membranes or with polymeric and/or composite membranes. This paper presents the obtaining of polymeric membranes containing metallic osmium obtained in situ. Cellulose acetate (CA), polysulfone (PSf) and polypropylene hollow fiber membranes (PPM) were used as support polymer membranes. The metallic osmium is obtained directly onto the considered membranes using a solution of osmium tetroxide (OsO4), dissolved in tert-butyl alcohol (t-Bu-OH) by reduction with molecular hydrogen. The composite osmium-polymer (Os-P)-obtained membranes were characterized in terms of the morphological and structural points of view: scanning electron microscopy (SEM), high-resolution SEM (HR-SEM), energy-dispersive spectroscopy analysis (EDAX), Fourier Transform Infra-Red (FTIR) spectroscopy, thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC). The process performance was tested for reduction of 5-nitrobenzimidazole to 5-aminobenzimidazole with molecular hydrogen. The paper presents the main aspects of the possible mechanism of transformation of 5-nitrobenzimidazole to 5-aminobenzimidazole with hydrogen gas in the reaction system with osmium-polymer membrane (Os-P).
PubMed: 34436396
DOI: 10.3390/membranes11080633 -
Journal of Anatomy Jan 2022Brachial plexus injury (BPI) occurs when the brachial plexus is compressed, stretched, or avulsed. Although rodents are commonly used to study BPI, these models poorly...
Brachial plexus injury (BPI) occurs when the brachial plexus is compressed, stretched, or avulsed. Although rodents are commonly used to study BPI, these models poorly mimic human BPI due to the discrepancy in size. The objective of this study was to compare the brachial plexus between human and Wisconsin Miniature Swine (WMS ), which are approximately the weight of an average human (68-91 kg), to determine if swine would be a suitable model for studying BPI mechanisms and treatments. To analyze the gross anatomy, WMS brachial plexuses were dissected both anteriorly and posteriorly. For histological analysis, sections from various nerves of human and WMS brachial plexuses were fixed in 2.5% glutaraldehyde, and postfixed with 2% osmium tetroxide. Subsequently paraffin sections were counter-stained with Masson's Trichrome. Gross anatomy revealed that the separation into three trunks and three cords is significantly less developed in the swine than in human. In swine, it takes the form of upper, middle, and lower systems with ventral and dorsal components. Histological evaluation of selected nerves revealed differences in nerve trunk diameters and the number of myelinated axons in the two species. The WMS had significantly fewer myelinated axons than humans in median (p = 0.0049), ulnar (p = 0.0002), and musculocutaneous nerves (p = 0.0454). The higher number of myelinated axons in these nerves for humans is expected because there is a high demand of fine motor and sensory functions in the human hand. Due to the stronger shoulder girdle muscles in WMS, the WMS suprascapular and axillary nerves were larger than in human. Overall, the WMS brachial plexus is similar in size and origin to human making them a very good model to study BPI. Future studies analyzing the effects of BPI in WMS should be conducted.
Topics: Animals; Brachial Plexus; Hand; Humans; Shoulder; Swine; Swine, Miniature; Upper Extremity
PubMed: 34355792
DOI: 10.1111/joa.13525 -
Angewandte Chemie (International Ed. in... Sep 2021This communication reports experimental and theoretical evidences of σ-hole interactions in adducts between nitrogen or oxygen nucleophiles and tetroxides of osmium or...
This communication reports experimental and theoretical evidences of σ-hole interactions in adducts between nitrogen or oxygen nucleophiles and tetroxides of osmium or other group 8 elements. Cocrystals between pyridine or pyridine N-oxide derivatives and osmium tetroxide are characterized through various techniques and rationalized as σ-hole interactions using DFT calculations and several other computational tools. We propose the term "osme bond" (OmB, Om=Fe, Ru, Os, (Hs)) for naming the noncovalent interactions wherein group 8 elements have the role of the electrophile. The word osme is the transcription of ὀσμή, the ancient Greek word for smell that was used to name the heaviest group 8 element in relation to the smoky odor of its tetroxide.
PubMed: 34260810
DOI: 10.1002/anie.202107978 -
International Journal For Parasitology Sep 2021Parasitic infections can be challenging to study because two dimensional light and electron microscopy are often limited in visualising complex and inaccessible...
Parasitic infections can be challenging to study because two dimensional light and electron microscopy are often limited in visualising complex and inaccessible attachment sites. Exemplifying this, Trichuris spp. inhabit a tunnel of epithelial cells within the host caecum and colon. A significant global burden of this infection persists, partly because available anthelminthics lack efficacy, although the mechanisms underlying this remain unknown. Consequently, there is a need to pioneer new approaches to better characterize the parasite niche within the host and investigate how variation in its morphology and integrity may contribute to resistance to therapeutic intervention. To address these aims, we exploited three-dimensional X-ray micro-computed tomography (microCT) to image the mouse whipworm, Trichuris muris, in caeca of wild-type C57BL/6 and SCID mice ex vivo. Using osmium tetroxide staining to effectively enhance the contrast of worms, we found that a subset exhibited preferential positioning towards the bases of the intestinal crypts. Moreover, in one rare event, we demonstrated whipworm traversal of the lamina propria. This morphological variability contradicts widely accepted conclusions from conventional microscopy of the parasite niche, showing Trichuris in close contact with the host proliferative and immune compartments that may facilitate immunomodulation. Furthermore, by using a skeletonization-based approach we demonstrate considerable variation in tunnel length and integrity. The qualitative and quantitative observations provide a new morphological point of reference for future in vitro study of host-Trichuris interactions, and highlight the potential of microCT to characterise enigmatic host-parasite interactions more accurately.
Topics: Animals; Mice; Mice, Inbred C57BL; Mice, SCID; Mucous Membrane; Trichuriasis; Trichuris; X-Ray Microtomography
PubMed: 34216623
DOI: 10.1016/j.ijpara.2021.04.006 -
Molecular Brain Jun 2021Immunogold labeling allows localization of proteins at the electron microscopy (EM) level of resolution, and quantification of signals. The present paper summarizes...
Immunogold labeling allows localization of proteins at the electron microscopy (EM) level of resolution, and quantification of signals. The present paper summarizes methodological issues and experiences gained from studies on the distribution of synaptic and other neuron-specific proteins in cell cultures and brain tissues via a pre-embedding method. An optimal protocol includes careful determination of a fixation condition for any particular antibody, a well-planned tissue processing procedure, and a strict evaluation of the credibility of the labeling. Here, tips and caveats on different steps of the sample preparation protocol are illustrated with examples. A good starting condition for EM-compatible fixation and permeabilization is 4% paraformaldehyde in PBS for 30 min at room temperature, followed by 30 min incubation with 0.1% saponin. An optimal condition can then be readjusted for each particular antibody. Each lot of the secondary antibody (conjugated with a 1.4 nm small gold particle) needs to be evaluated against known standards for labeling efficiency. Silver enhancement is required to make the small gold visible, and quality of the silver-enhanced signals can be affected by subsequent steps of osmium tetroxide treatment, uranyl acetate en bloc staining, and by detergent or ethanol used to clean the diamond knife for cutting thin sections. Most importantly, verification of signals requires understanding of the protein of interest in order to validate for correct localization of antibodies at expected epitopes on particular organelles, and quantification of signals needs to take into consideration the penetration gradient of reagents and clumping of secondary antibodies.
Topics: Animals; Brain; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cell Membrane; Cell Membrane Permeability; Cells, Cultured; Chromogranin A; Hippocampus; Membrane Proteins; Mice; Microscopy, Electron; Neurons; Rats; Staining and Labeling; Tissue Embedding; Tissue Fixation
PubMed: 34082785
DOI: 10.1186/s13041-021-00799-2 -
The Journal of Organic Chemistry Jun 2021-Dihydroxylation of trinor-18α-olean-17(22)-ene with osmium tetroxide led to diol . Its cleavage with lead tetraacetate gave tetracyclic ketoaldehyde . By comparison,...
-Dihydroxylation of trinor-18α-olean-17(22)-ene with osmium tetroxide led to diol . Its cleavage with lead tetraacetate gave tetracyclic ketoaldehyde . By comparison, the ozonation of trinor-18α-olean-17(22)-ene in the presence of -toluenesulfonic acid gave the corresponding ketoacetal . Both products were subjected to an intramolecular aldol reaction under the acidic conditions and yielded unusual triterpenes bearing a bicyclo[4.3.1]decane fragment (). Further manipulation of the protective groups afforded compounds useful in triterpene synthesis, especially in the preparation of potentially biologically active saponins based on a tetracyclic terpene core.
Topics: Aldehydes; Oxidative Stress; Saponins; Triterpenes
PubMed: 34033473
DOI: 10.1021/acs.joc.1c00697