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Skin Pharmacology and Physiology 2018In the mid-1950s and 1960s, transmission electron microscopes became widely available, leading to many studies of the ultrastructure of various tissues including the... (Review)
Review
In the mid-1950s and 1960s, transmission electron microscopes became widely available, leading to many studies of the ultrastructure of various tissues including the epidermis. Most of these studies involved tissue fixation with formaldehyde and postfixation with osmium tetroxide. A few studies employed freeze-fracture electron microscopy. One set of these studies identified a small organelle variously called lamellar granules (LGs), lamellar bodies, membrane-coating granules, cementsomes, and Odland bodies. LGs are round to ovoid in shape, with a diameter of about 200 nm. They have a bounding membrane surrounding a stack of internal lipid lamellae. These small organelles are first seen in the spinous layer and accumulate with differentiation in the granular layer. In the uppermost granular cells, the bounding membrane of the LG fuses into the cell plasma membrane, and the internal contents are extruded into the intercellular space. The initially extruded contents of the LG then rearrange to form the intercellular lamellae of the stratum corneum. In this context, LGs serve as the precursor to the permeability barrier of the skin. Various studies have provided evidence that they are derived from the Golgi apparatus, specifically the trans-Golgi. Isolated LGs contain phosphoglycerides, sphingomyelin, and glucosylceramides. The most unusual lipid component is a linoleate-containing glucosylceramide comprising 30- to 34-carbon ω-hydroxy-acids. Isolated granules also contain acid hydrolases including glucocerebrosidase, sphingomyelinase, and phospholipase A. They also contain proteases and antimicrobial peptides. Defective LGs have been associated with a number of skin diseases including ichthyotic conditions and defective barrier function. Recently, studies employing cryo-transmission electron microscopy have called into question the validity of observations on LGs with more conventional electron microscopic techniques. These studies suggest a continuity of the membrane structure from the Golgi through the intercellular lamellae of the stratum corneum.
Topics: Animals; Epidermis; Glucosylceramides; Golgi Apparatus; Humans; Microscopy, Electron, Transmission; Skin Diseases
PubMed: 30110701
DOI: 10.1159/000491757 -
Basic & Clinical Pharmacology &... Nov 2020Osmium tetroxide is a strong oxidizing agent used in electron microscopy. Eye exposure may cause severe burns, and after inhalation or ingestion damage to the...
Osmium tetroxide is a strong oxidizing agent used in electron microscopy. Eye exposure may cause severe burns, and after inhalation or ingestion damage to the respiratory or gastrointestinal tract occurs. Exposure to osmium and its compounds is extremely rare. We present a case of a 32-year-old female stained by 9 mL of 2% osmium tetroxide in acetone during an accident in the laboratory, with rare dermal and ocular findings. Due to lack of data in toxicological databases and the absence of antidote, the therapy was symptomatic. Osmium was detected in serum 19 hours later (0.22 μg/L) and in urine during the 15-hour collection (three samples-7.05, 1.65 and 8.45 μg/L). In blood serum on admission, after 1 and 2 days after exposure, the levels of iron (28.2, 39.8 and 50.5 μmol/L; reference range 5.8-34.5 μmol/L) and transferrin receptor/ferritine were elevated. To our knowledge, this is the first paper documenting a significant absorption from the skin and potentially from the eye conjunctiva, based on serum and urine analysis. The relationship between increased iron in blood and exposure has not been described yet, and the mechanism remains unknown. The patient is being followed up for the unknown long-term effects.
Topics: Adult; Eye; Female; Humans; Osmium Tetroxide; Skin
PubMed: 32524772
DOI: 10.1111/bcpt.13450 -
Internal Medicine (Tokyo, Japan) 2023A 32-year-old man was admitted for the evaluation of proteinuria (5.69 g/day). A light microscopic examination showed markedly dilated glomerular capillary loops with...
A 32-year-old man was admitted for the evaluation of proteinuria (5.69 g/day). A light microscopic examination showed markedly dilated glomerular capillary loops with vacuolated areas in many glomeruli, and vacuolated areas were seen on peritubular capillaries in the tubulointerstitium. When electron microscopy specimens prepared by pre-fixation with glutaraldehyde and post-fixation with osmium tetroxide were used for oil red staining, the deposition was confirmed on the affected areas. A genetic analysis of apoE showed that the lipoprotein glomerulopathy was due to apoE-Sendai (Arg145Pro, p.R163P) heterozygosity, which was found in not only the patient but also his mother and twin brother.
Topics: Male; Humans; Adult; Apolipoproteins E; Kidney Diseases; Kidney Glomerulus; Proteinuria; Heterozygote
PubMed: 37532513
DOI: 10.2169/internalmedicine.0834-22 -
Current Eye Research Aug 2018To examine the ultrastructure of lipofuscin bodies and melanosomes in retinal epithelium of elderly rhesus monkeys and determines changes in their number and morphology...
PURPOSE
To examine the ultrastructure of lipofuscin bodies and melanosomes in retinal epithelium of elderly rhesus monkeys and determines changes in their number and morphology as a function of retinal eccentricity.
METHODS
Electron microscopy was used to describe and quantify two major organelles in elderly monkey retinal epithelium, lipofuscin bodies and melanosomes, at different retinal loci extending from the macula to the peri-macula, equator, periphery and ora serrata. Osmium tetroxide was used to distinguish lipofuscin bodies from melanosomes.
RESULTS
Lipofuscin bodies and melanosomes diminished in number with advanced age but there was an inverse relationship between these two organelles. Lipofuscin bodies were more numerous in the macula and melanosomes more numerous in the peripheral retina. Three types of lipofuscin bodies were identified: 1) smaller and tending to locate in the middle third of the epithelial cell, 2) larger, less common, and located more basally, and 3) extremely rare, melano-lipofuscin, containing a melanosome. When osmicated, all lipofuscin bodies contained electron dense materials. When osmium tetroxide was not used for fixation, the first two types of lipofuscin bodies lost their electron densities while the third type retained its electron density due to the melanosome it contained.
CONCLUSION
As previously reported for human retina, lipofuscin is most abundant in the macular and peri-macular epithelium and least abundant in the periphery, whereas melanosomes show the opposite relationship. This distribution pattern could contribute to the macula's greater vulnerability to photo-toxicity. Three types of lipofuscin bodies are found in aging monkey retinal epithelium. All types contain electron dense material, but the most prominent two types lose their densities in the absence of osmium tetroxide during fixation. Most of the electron densities in lipofuscin bodies must contain a material that binds strongly to osmium tetroxide such as polyunsaturated fatty acids.
Topics: Aging; Animals; Electron Probe Microanalysis; Female; Lipofuscin; Macaca mulatta; Male; Melanosomes; Microscopy, Electron; Models, Animal; Osmium Tetroxide; Retinal Pigment Epithelium
PubMed: 29641909
DOI: 10.1080/02713683.2018.1464194 -
Applied Microscopy May 2020Plant specimens for scanning electron microscopy (SEM) are commonly treated using standard protocols. Conventional fixatives consist of toxic chemicals such as... (Review)
Review
Plant specimens for scanning electron microscopy (SEM) are commonly treated using standard protocols. Conventional fixatives consist of toxic chemicals such as glutaraldehyde, paraformaldehyde, and osmium tetroxide. In 1996, methanol fixation was reported as a rapid alternative to the standard protocols. If specimens are immersed in methanol for 30 s or longer and critical-point dried, they appear to be comparable in preservation quality to those treated with the chemical fixatives. A modified version that consists of methanol fixation and ethanol dehydration was effective at preserving the tissue morphology and dimensions. These solvent-based fixation and dehydration protocols are regarded as rapid and simple alternatives to standard protocols for SEM of plants.
PubMed: 33580311
DOI: 10.1186/s42649-020-00028-5 -
Cytometry. Part a : the Journal of the... May 2016Mass cytometry facilitates high-dimensional, quantitative, single-cell analysis. The method for sample multiplexing in mass cytometry, called mass-tag cellular barcoding...
Mass cytometry facilitates high-dimensional, quantitative, single-cell analysis. The method for sample multiplexing in mass cytometry, called mass-tag cellular barcoding (MCB), relies on the covalent reaction of bifunctional metal chelators with intracellular proteins. Here, we describe the use of osmium and ruthenium tetroxides (OsO4 and RuO4 ) that bind covalently with fatty acids in the cellular membranes and aromatic amino acids in proteins. Both OsO4 and RuO4 rapidly reacted and allowed for MCB with live cells, crosslinked cells, and permeabilized cells. Given the covalent nature of the labeling reaction, isotope leaching was not observed. OsO4 and RuO4 were used in a 20-sample barcoding protocol together with palladium isotopes. As mass channels occupied by osmium and ruthenium are not used for antibody detection the number of masses effectively utilized in a single experiment is expanded. OsO4 and RuO4 can therefore be used as MCB reagents for a wide range of mass cytometry workflows. © 2016 International Society for Advancement of Cytometry.
Topics: Amino Acids; Antibodies, Monoclonal; Antigens, CD; Cell Line, Tumor; Chelating Agents; Cytophotometry; Fatty Acids; Heterocyclic Compounds, 1-Ring; Humans; Jurkat Cells; Leukocytes, Mononuclear; Mass Spectrometry; Osmium Tetroxide; Palladium; Ruthenium Compounds; Single-Cell Analysis; Staining and Labeling
PubMed: 27018769
DOI: 10.1002/cyto.a.22848 -
Frontiers in Cell and Developmental... 2022Volume electron microscopy, a powerful approach to generate large three-dimensional cell and tissue volumes at electron microscopy resolutions, is rapidly becoming a...
Volume electron microscopy, a powerful approach to generate large three-dimensional cell and tissue volumes at electron microscopy resolutions, is rapidly becoming a routine tool for understanding fundamental and applied biological questions. One of the enabling factors for its adoption has been the development of conventional fixation protocols with improved heavy metal staining. However, freeze-substitution with organic solvent-based fixation and staining has not realized the same level of benefit. Here, we report a straightforward approach including osmium tetroxide, acetone and up to 3% water substitution fluid (compatible with traditional or fast freeze-substitution protocols), warm-up and transition from organic solvent to aqueous 2% osmium tetroxide. Once fully hydrated, samples were processed in aqueous based potassium ferrocyanide, thiocarbohydrazide, osmium tetroxide, uranyl acetate and lead acetate before resin infiltration and polymerization. We observed a consistent and substantial increase in heavy metal staining across diverse and difficult-to-fix test organisms and tissue types, including plant tissues (), nematode () and yeast (). Our approach opens new possibilities to combine the benefits of cryo-preservation with enhanced contrast for volume electron microscopy in diverse organisms.
PubMed: 36003147
DOI: 10.3389/fcell.2022.933376 -
Membranes Dec 2021The recovery of osmium from residual osmium tetroxide (OsO) is a necessity imposed by its high toxicity, but also by the technical-economic value of metallic osmium. An...
The recovery of osmium from residual osmium tetroxide (OsO) is a necessity imposed by its high toxicity, but also by the technical-economic value of metallic osmium. An elegant and extremely useful method is the recovery of osmium as a membrane catalytic material, in the form of nanoparticles obtained on a polymeric support. The subject of the present study is the realization of a composite membrane in which the polymeric matrix is the polypropylene hollow fiber, and the active component consists of the osmium nanoparticles obtained by reducing an alcoholic solution of osmium tetroxides directly on the polymeric support. The method of reducing osmium tetroxide on the polymeric support is based on the use of 10-undecenoic acid (10-undecylenic acid) (UDA) as a reducing agent. The osmium tetroxide was solubilized in -butanol and the reducing agent, 10-undecenoic acid (UDA), in -propanol, -butanol or -decanol solution. The membranes containing osmium nanoparticles (Os-NP) were characterized morphologically by the following: scanning electron microscopy (SEM), high-resolution SEM (HR-SEM), structurally: energy-dispersive spectroscopy analysis (EDAX), Fourier transform infrared (FTIR) spectroscopy. In terms of process performance, thermal gravimetric analysis was performed by differential scanning calorimetry (TGA, DSC) and in a redox reaction of an organic marker, -nitrophenol (PNP) to -aminophenol (PAP). The catalytic reduction reaction with sodium tetraborate solution of PNP to PAP yielded a constant catalytic rate between 2.04 × 10 mmol s and 8.05 × 10 mmol s.
PubMed: 35054577
DOI: 10.3390/membranes12010051 -
Methods in Enzymology 2014Adipocytes reside in discrete, well-defined depots throughout the body. In addition to mature adipocytes, white adipose tissue depots are composed of many cell types,...
Adipocytes reside in discrete, well-defined depots throughout the body. In addition to mature adipocytes, white adipose tissue depots are composed of many cell types, including macrophages, endothelial cells, fibroblasts, and stromal cells, which together are referred to as the stromal vascular fraction (SVF). The SVF also contains adipocyte progenitors that give rise to mature adipocytes in those depots. Marrow adipose tissue (MAT) or marrow fat has long been known to be present in bone marrow (BM) but its origin, development, and function remain largely unknown. Clinically, increased MAT is associated with age, metabolic diseases, drug treatment, and marrow recovery in children receiving radiation and chemotherapy. In contrast to the other depots, MAT is unevenly distributed in the BM of long bones. Conventional quantitation relies on sectioning of the bone to overcome issues with distribution but is time-consuming, resource intensive, inconsistent between laboratories and may be unreliable as it may miss changes in MAT volume. Thus, the inability to quantitate MAT in a rapid, systematic, and reproducible manner has hampered a full understanding of its development and function. In this chapter, we describe a new technique that couples histochemical staining of lipid using osmium tetroxide with microcomputerized tomography to visualize and quantitate MAT within the medullary canal in three dimensions. Imaging of osmium staining provides a high-resolution map of existing and developing MAT in the BM. Because this method is simple, reproducible, and quantitative, we expect it will become a useful tool for the precise characterization of MAT.
Topics: Adipogenesis; Adipose Tissue, White; Bone Marrow; Cell Differentiation; Humans; Osmium Tetroxide; Staining and Labeling; Stromal Cells; X-Ray Microtomography
PubMed: 24480344
DOI: 10.1016/B978-0-12-411619-1.00007-0 -
Microscopy (Oxford, England) Aug 2022Cultured Lithospermum erythrorhizon cells were fixed with a new fixation method to visualize the metabolism of shikonin derivatives, the lipophilic naphthoquinone...
Cultured Lithospermum erythrorhizon cells were fixed with a new fixation method to visualize the metabolism of shikonin derivatives, the lipophilic naphthoquinone pigments in Boraginaceae. The new fixation method combined glutaraldehyde containing malachite green, imidazole-osmium and p-phenylenediamine treatments, and cells were then observed with a transmission electron microscope. The method prevented the extraction of lipids, including shikonin derivatives, and improved the visualization of subcellular structures, especially the membrane system, when compared with that of conventional fixation. The improved quality of the transmission electron micrographs is because malachite green ionically binds to the plasma membrane, organelles and lipids and acts as a mordant for electron staining with osmium tetroxide. Imidazole promotes the reaction of osmium tetroxide, leading to enhanced electron staining. p-Phenylenediamine reduces osmium tetroxide bound to cellular materials and increases the electron density. This protocol requires only three additional reagents over conventional chemical fixation using glutaraldehyde and osmium tetroxide.
Topics: Glutaral; Imidazoles; Lipids; Microscopy, Electron; Microscopy, Electron, Transmission; Osmium Tetroxide; Plant Cells
PubMed: 35388424
DOI: 10.1093/jmicro/dfac018