Biomedical subjects

J A Litwin

Publications and source records attributed to J A Litwin.

At least 37 records · Page 2Linked to original sources

PubMed1995

Morphological heterogeneity of peroxisomes in cultured mouse Leydig cells.

Peroxisomes of different shapes were visualized in mouse Leydig cells from primary cultures by immunofluorescent demonstration of catalase and two peroxisomal lipid beta-oxidation enzymes, acyl-CoA oxidase and 3-ketoacyl-CoA thiolase. Apart from typical spherical profiles, rod-shaped, tubular and annular peroxisomes were also observed. The proportion between spherical and elongated forms varied from cell to cell; the latter forms seemed to occur more frequently in flattened cells spread on the glass in the course and after monolayer formation. On very rare occasions the Leydig cells contained small local networks of interconnected tubular peroxisomes, probably representing areas of peroxisomal reticulum, whose existence has been so far supported by substantial biochemical, but hardly any morphological evidence.

Acetyl-CoA C-Acetyltransferase↗

PubMed1994

Blood vessels in epiphyseal cartilage of human fetal femoral bone: a scanning electron microscopic study of corrosion casts.

Formation of intrachondral vessels (cartilage canals) in the proximal femoral epiphysis was studied in 13- to 22-week-old human fetuses using a corrosion casting technique and scanning electron microscopy. Several successive morphological stages of angiogenesis occurring inside the hyaline cartilage were distinguished. The process of cartilage vascularization starts with the formation of hairpin loops sent off from the perichondrial vascular network into the adjacent cartilage. A capillary glomerulus is then formed at the leading end, and the entire vascular unit grows in length, assuming a mushroom-like shape. Its further elongation is accompanied by a backward expansion of the capillary network which surrounds a pair of main vessels (arteriole and venule) like a manchette. The subsequent branching of such primary vascular units proceeds according to the same morphological patterns. The resulting tree-like vascular formations become interconnected via their lateral branches. This study clearly supports the invasion theory of cartilage canal formation.

Blood Vessels↗

PubMed1994

The vascular architecture of human fetal oviduct: a scanning electron microscopic study of corrosion casts.

Vascular architecture was investigated, in 18-21 week old human fetuses, for the first time with the use of corrosion casting and scanning electron microscopy, which offer high resolution and three-dimensional images of the vascular networks. The general arrangement of large vessels was similar to that described for the mature oviduct; however, the intramural vasculature of the muscular layer and mucosal folds consisted predominantly of capillary and sinusoidal networks. A characteristic feature of the fetal oviduct was a prominent sub-serosal venous plexus, which was most extensive in the isthmic segment. The relatively low degree of differentiation found in the fetal oviduct vasculature seems to reflect its functional immaturity.

Arteries↗

PubMed1994

The lung in closeview: a corrosion casting study on the vascular system of human foetal trachea.

The aim of this study was to examine the tracheal vasculature in 5 month human foetuses, and to determine whether it differs from that previously described by other authors for adult human trachea. The vascular bed was visualized using the technique of corrosion casting and examined by scanning electron microscopy. The arrangement of larger vessels: longitudinal tracheo-oesophageal arteries and veins, as well as their segmental branches running circumferentially in the intercartilaginous spaces, was similar to that observed in the trachea of adults. However, no blood sinuses reported to occur in the submucous venular plexus of the human trachea could be found in the foetuses. It is postulated that the possible functions of such sinuses acting as the capacitance system are related to the functional respiratory tract; hence, this vascular specialization is not yet developed in the foetus.

Blood Vessels↗

PubMed1994

The vascular system of human fetal long bones: a scanning electron microscope study of corrosion casts.

The vascular system of the femur and humerus was investigated in 17-24 wk human fetuses by scanning electron microscopy of corrosion casts. The number of nutrient foramina present in both bones ranged from 1 to 3 and the number of nutrient vessels associated with individual foramina also varied. The medullary arteries supplied both the bone cortex and marrow. There was no arterial supply to the shaft cortex from the periosteal side, where only capillaries were found to enter the bone. The metaphyses were supplied and drained by conspicuous vascular triads composed of an artery and 2 veins. In the marrow cavity, 2 morphologically different areas of the fine vascular network could be distinguished: diaphyseal sinusoids and metaphyseal capillaries forming a 'vascular besom' which was abruptly demarcated by the growth plate cartilage. The cortical microvascular bed was composed of capillaries and more numerous irregular sinusoids. The 2 main vascular systems, nutrient and periosteal, were interconnected not only via the network of cortical capillaries/sinusoids, but also by larger arteries and veins traversing the cortex. The features of the vascular system of human fetal long bones suggest its considerable functional flexibility and its capacity to modify blood flow patterns depending on circumstances.

Arteries↗

PubMed1994

Microcirculation of human fetal tooth buds: a SEM study of corrosion casts.

The microcirculation of tooth buds at the bell stage obtained from 5-month-old human fetuses was studied using corrosion casting and scanning electron microscopy. Each tooth bud has two independent vascular networks: one of the enamel organ and one of the dental papilla. Both systems are supplied by vertical branches of the inferior alveolar artery. The vascular bed of the enamel organ consists of capillaries relatively uniform in shape, forming a moderately dense network with irregular meshes. In contrast, the vasculature of dental papilla is extremely dense and its vessels show a sinusoidal character and signs of a vivid angiogenesis. The cast surfaces of capillaries in both vascular systems show the presence of tiny blebs probably representing extravasations of the casting medium through endothelial fenestrations.

Corrosion Casting↗

PubMed1993

Hyaloid vessels of the human fetal eye. A scanning electron microscopic study of corrosion casts.

OBJECTIVE: Microscopic investigation of the hyaloid vascular system in 5-month-old human fetuses. METHODS: Corrosion casting and light and scanning electron microscopy. RESULTS: The hyaloid artery ramifies into a tuft of vasa hyaloidea propria, which communicates with the posterior portion of the tunica vasculosa lentis, characterized by a network of anastomosing vessels. They further pass to the lateral portion of the tunica, acquiring a nonanastomosing palisadelike array and drain into the vessels of the ciliary processes or, after bending over the edge of the developing iris, drain into the outer choriocapillaris. The tunica vasculosa lentis vessels also communicate with the pupillary membrane, a system of vascular arcades arranged in several interconnected tiers, supplied by the terminal branches of the long posterior ciliary arteries. In tunica vasculosa lentis, arterioles seem to pass directly into veins, without forming a capillary bed. CONCLUSIONS: At the investigated developmental stage, the fully developed hyaloid system enters its subsequent involution, and the vessels nourishing the vitreous have already involuted. The system is generally similar to that observed in other mammals.

Arteries↗

PubMed1993

A simple method for demonstration of eosinophils in frozen sections.

Tissue eosinophils can be easily and specifically demonstrated in frozen sections after substrateless incubation with 3,3'-diaminobenzidine (DAB) at neutral pH. Under such conditions, other cells possessing endogenous peroxidatic activity (neutrophils, macrophages) do not stain, whereas staining of mitochondria-rich cells due to cytochrome oxidase activity can be avoided by a short prefixation of sections in glutaraldehyde-containing fixatives. The method can be useful in cases when a rapid screening of tissue eosinophils is required.

3,3'-Diaminobenzidine↗

PubMed1993

Microvascular system of the human fetal inner ear: a scanning electron microscopic study of corrosion casts.

The vascular system of the inner ear was investigated in 18-21 weeks old human fetuses, using the corrosion cast technique in scanning electron microscopy. At that developmental stage, vascularization of the cochlea and semicircular canals shows a pattern very similar to that described for adults. The most important differences which can be regarded as fetal features include: (1) denser limbus vessels, (2) the marginal vessels of the spiral lamina appearing as irregular network which shows a less clear arcade-like arrangement, (3) some radiating arterioles of the spiral lamina and marginal vessels possessing connections with the vascular system of the external wall, and (4) a dense, sinusoidal network of draining venules at scala tympani. These features apparently disappear during the final remodelling of the inner ear microvasculature in the last trimester.

Blood Vessels↗

PubMed1991

Endogenous cytochrome c and cytochrome oxidase in rat and mouse kidney: light microscopic histochemical study.

Activity of cytochrome c oxidase and the level of endogenous cytochrome c were investigated light microscopically in adult rat and mouse kidney by incubating unfixed frozen sections with diaminobenzidine (DAB) in the absence or presence of exogenous cytochrome c. The results suggest that DAB staining intensity mainly reflects the local density of mitochondria and only occasionally visualizes the differences in cytochrome oxidase activity and/or endogenous cytochrome c content. Most intense reaction was observed in proximal and distal tubules both in rat and mouse. Finer differentiation of reactivity in particular nephron segments and interspecies differences between rat and mouse kidney are also described.

3,3'-Diaminobenzidine↗

PubMed1990

Different levels of reactive endogenous cytochrome c in mitochondria rich cells revealed by diaminobenzidine staining.

Light microscopic examination of rat and mouse tissues incubated in a medium containing 3,3'-diaminobenzidine (DAB) and catalase revealed that cells known to possess abundant mitochondria (hepatocytes, cardiomyocytes, renal proximal and distal tubular cells, parietal cells of gastric mucosa, and retinal photoreceptor cells) were stained with different intensity: from moderate (parietal cells, cardiomyocytes, renal distal tubular cells) to weak (hepatocytes, renal proximal tubular cells) or even negative (photoreceptors). When exogenous cytochrome c was added to the incubation medium, all these cells displayed quite uniform, strong staining, indicating a comparable activity of cytochrome oxidase. Since DAB is oxidized directly by cytochrome c which in turn undergoes reoxidation by cytochrome oxidase, the observed differences of staining intensity in the absence of exogenous cytochrome c are postulated to result from different content of reactive endogenous cytochrome c in mitochondria of the investigated cells.

3,3'-Diaminobenzidine↗

PubMed1988

Immunogold localization of peroxisomal enzymes in Epon-embedded liver tissue. Enhancement of sensitivity by etching with ethanolic sodium hydroxide.

Epon-embedded biological materials exhibit well preserved ultrastructure but generally weak antigenicity. Brief etching of ultrathin Epon sections with resin solvent, ethanolic sodium hydroxide, brought about a nearly 2-fold increase in the immunogold labeling density of rat and human hepatic peroxisomes after incubation with antisera against catalase and 3 enzymes of lipid beta-oxidation: acyl-CoA oxidase, bifunctional protein (enoyl-CoA hydratase-3-hydroxyacyl-CoA dehydrogenase) and 3-ketoacyl-CoA thiolase. The etching was superior to pretreatment with an oxidant, sodium metaperiodate. Despite some deterioration of the cellular ultrastructure, the obtained enhancement of the sensitivity of the protein A-gold method may be helpful in cases when the antigenicity to be detected is strongly inhibited by epoxy resin embedding.

Acetyl-CoA C-Acyltransferase↗

PubMed1988

Detection of peroxisomes in human liver and kidney fixed with formalin and embedded in paraffin: the use of catalase and lipid beta-oxidation enzymes as immunocytochemical markers.

We describe the immunocytochemical localization of four peroxisomal enzymes by light microscopy in human liver and kidney processed routinely by formalin fixation and paraffin embedding. Monospecific antisera against catalase and three enzymes of peroxisomal lipid beta-oxidation (acyl-CoA oxidase, bifunctional protein (enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase) and 3-ketoacyl-CoA thiolase) were used in conjunction with either the indirect immunoperoxidase method or the protein A-gold technique followed by silver intensification. The sections of formalin-fixed paraffin-embedded tissue had to be deparaffinized and subjected to controlled proteolysis in order to obtain satisfactory immunostaining. Under the conditions employed, peroxisomes were distinctly visualized in liver parenchymal cells with no reaction in bile duct epithelial or sinusoidal lining cells. In the kidney, peroxisomes were confined to the proximal tubular epithelial cells with negative staining of glomeruli, distal tubules and collecting ducts. A positive immunocytochemical reaction was obtained even in paraffin blocks stored for several years. The method offers a simple approach for detection of peroxisomes and evaluation of their various enzyme proteins in material processed routinely in histopathology laboratories and should prove useful in the investigation of the role of peroxisomes in human pathology for both prospective and retrospective studies.

Catalase↗

PubMed1988

Immunocytochemical demonstration of peroxisomal enzymes in human kidney biopsies.

Peroxisomes are particularly abundant in the proximal tubules of the mammalian kidney. We describe the immunocytochemical localization of catalase and three peroxisomal lipid beta-oxidation enzymes: acyl-CoA oxidase, bifunctional protein (enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase) and 3-ketoacyl-CoA thiolase, in human renal biopsies fixed with glutaraldehyde and embedded in Epon. For light microscopy of semithin sections, satisfactory immunostaining required removal of the resin and controlled proteolytic digestion followed by the indirect immunoperoxidase technique. Brief etching of ultrathin sections with alkoxide followed by the protein A-gold method were used for electron microscopic localization of the enzymes. The immunoreactive peroxisomes were distinctly visualized in proximal tubular epithelial cells with no staining of any other cell organelles. The results establish the presence of catalase and of peroxisomal lipid beta-oxidation system proteins in human kidney. The immunocytochemical procedure described herein provides a simple approach for the investigation of peroxisomal structure and function in human renal biopsies processed for ultrastructural studies.

3-Hydroxyacyl CoA Dehydrogenases↗

PubMed1987

Immunocytochemical localization of peroxisomal enzymes in human liver biopsies.

The immunocytochemical localization of catalase and three enzymes of the peroxisomal lipid beta-oxidation system--acyl-CoA oxidase, the bifunctional protein enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, and 3-ketoacyl-CoA thiolase--in human liver biopsies was investigated by means of light and electron microscopy. The antisera raised against all four enzymes from rat liver cross-reacted with the corresponding proteins in homogenates of human liver as revealed by immunoblotting. For light-microscopic localization in glutaraldehyde-fixed Epon-embedded material, the removal of resin and controlled digestion with trypsin was necessary. At the ultrastructural level specific labeling for all four antigens was found by the protein A-gold technique in peroxisomes of liver parenchymal cells fixed with formaldehyde-low glutaraldehyde concentrations and embedded in Lowicryl K4M. In biopsies fixed with glutaraldehyde and embedded in Epon, treatment with metaperiodate or etching with sodium ethoxide improved the immunolabeling. After such treatment catalase showed the most intense labeling and acyl-CoA oxidase the weakest, the two other proteins exhibiting an intermediate immunoreaction. In material postfixed with osmium only catalase could be visualized in peroxisomes. The immunocytochemical investigation of peroxisomal proteins in human liver biopsies provides a simple and highly promising approach for further elucidation of the pathophysiology of peroxisomal disorders.

3-Hydroxyacyl CoA Dehydrogenases↗

PubMed1985

Light microscopic histochemistry on plastic sections.

As compared with conventional paraffin, celloidin, and frozen sections, semithin plastic sections offer a superior quality of the light microscopic image in terms of better resolution, absence of distortion and shrinkage artifacts, and suitability for calcified tissues. Application of histochemical methods to such sections often encounters, however, serious difficulties resulting from a considerably reduced reactivity of plastic-embedded biological material. Factors involved include a poor penetration of reagents into plastic embedding media due to a steric or hydrophobic hindrance, as well as a blockade of the reactive chemical groups in the sample due to interactions with fixatives and plastics. Embedding in polar (hydrophilic) plastics, such as glycol methacrylate, permits carrying out a large number of histochemical reactions, including the demonstration of enzymatic activities, directly on sections, but is less suitable for combined light/electron microscopic studies because of an imperfect ultrastructural preservation of tissues. Embedding in nonpolar epoxy resins, particularly if combined with a double aldehyde-osmium fixation, results in a high quality ultrastructure but almost fully inhibits the histochemical reactivity of the embedded material. In order to restore this reactivity, i.e. to unmask chemical groups bound by the polymerized resin, semithin epoxy sections require the removal of the embedding matrix by alkoxides prior to the histochemical procedure. Additional steps are also often necessary: treatment of osmium-fixed sections with oxidative agents, e.g., hydrogen peroxide or periodate which reoxidize the bound osmium and remove it from tissue, and a controlled proteolytic digestion, especially useful in immunocytochemical studies, which probably cleaves the bonds between the primary aldehyde fixative, and the reactive sites. This article reviews histochemical methods which have been successfully applied to plastic-embedded material. Using polar methacrylates and/or nonpolar epoxy resins as embedding media, it has been possible to demonstrate proteins and aminoacid residues, carbohydrates, lipids, nucleic acids, biogenic amines, inorganic ions, and some enzymes, although the spectrum of methods found as suitable for plastic-embedded material is far narrower than that available for paraffin or frozen sections.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1984

Peroxidase-positive endothelial cells in rat liver.

Rat liver fixed by perfusion with low glutaraldehyde concentrations was incubated in diaminobenzidine-containing medium to stain for peroxidase. Endogenous peroxidatic activity was found not only in Kupffer cells but also in the endothelial cells lining the sinusoids and central veins. The reaction product was localized in the nuclear envelope and endoplasmic reticulum. The peroxidatic activity in endothelial cells showed a concentration-dependent sensitivity to glutaraldehyde: in liver samples fixed with 0.25% glutaraldehyde, approx. 23% of the sinusoidal endothelial cells and 65% of central vein endothelium were peroxidase-positive; with 0.5% glutaraldehyde, only approx. 8% of the sinusoidal endothelial cells contained detectable amounts of the reaction product; with 1.5% glutaraldehyde all endothelial cells were consistently peroxidase-negative. No peroxidatic activity could be found in liver endothelial cells following isolation by centrifugal elutriation. Endothelial cell peroxidase may possibly be involved in defense responses of liver and/or, as a part of prostaglandin synthase system, in prostanoid production.

Animals↗

PubMed1984

Light microscopic immunocytochemical demonstration of peroxisomal enzymes in epon sections.

A procedure is described for light microscopic immunocytochemical localization of catalase and three enzymes of peroxisomal lipid beta-oxidation: acyl-CoA oxidase, enoyl-CoA hydratase and 3-ketoacyl-CoA thiolase in semi-thin sections of rat liver processed for routine electron microscopy. Satisfactory immunostaining required the removal of the epoxy resin with sodium ethoxide, controlled digestion of deplasticized sections with proteases and, in case of osmiumfixed tissue, bleaching with oxidants. Resin removal was essential for successful immunostaining, and protease treatment enhanced markedly the intensity of the reaction. This study shows that tissues processed for conventional ultrastructural studies can be used for postembedding immunocytochemical demonstration of various peroxisomal enzymes.

Acetyl-CoA C-Acyltransferase↗