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PubMed1993

Recombination nodules and axial equalization in the ZW pairs of the Peking duck and the guinea fowl.

The meiotic Z and W chromosomes of the Guinea fowl (Numida meleagris) and the Peking duck (Anas platyrhynchos) show features similar to those described previously for 3 species of Phasianidae: chicken, Japanese quail and bobwhite quail. Axial equalization in the gonosomes of the Peking duck involves lengthening of the W axis along with shortening of the Z axis. A single recombination nodule is constantly found in the synaptic, terminal region of the ZW pairs stained with phosphotungstic acid. The localization of this nodule is significantly different in these two species, but the length of synaptonemal complex harboring the nodules is not significantly different. Based on the phylogenetic distance between the observed species, the present data support the hypothesis that the presence of a pseudoautosomal region is a general property of ZW pairs in carinate birds.

Animals↗
PubMed1989

Evidence of repetitive patterns of chromatin distribution in cell nuclei of rat liver.

Samples of rat livers were fixed in glutaraldehyde, contrasted en bloc with phosphotungstic acid, embedded in an epoxy resin and serially sectioned. The study of three-dimensional models of 20 complete nuclei shows that all of them share some general features: they have more than one nucleolus (2-4), an irregular layer of compact chromatin adjacent to the nuclear membrane and well-delimited clumps of chromatin both in the nuclear sap and surrounding the nucleoli. A space of 8 sections containing the central nucleolus and a lateral one was studied in detail. In this space, 8 clumps of compact chromatin were found in 17 nuclei and 9 clumps in the other 3 nuclei. No other number of clumps was found in those zones. In all the nuclei studied the compact chromatin surrounding the central nucleolus contacts the nuclear envelope. This contact takes place in a region almost diametrically opposed to the lateral nucleolus in 13 nuclei. In 7 nuclei, these structures were at angles between 50 and 125 degrees. These results support the existence of nonrandom repetitive patterns of chromatin distribution in liver cells.

Animals↗
PubMed1991

Effects of low-power gallium aluminium arsenide diode laser irradiation on the development of synapses in the neonatal rat hippocampus.

The effect of low-power gallium aluminium arsenide diode laser irradiation on the development of synapses was studied in the radiatum layer and the lacunosum-molecular layer of field CA3 of the neonatal rat hippocampus. Neonatal rats were irradiated with a low-power diode laser (830 nm, 60 mW) at two points located above the hippocampi, for 15 s, respectively, twice per day from birth (day 1) to day 5. The mean body weights of the laser-irradiated animals were found to be lower than those of the control animals, the deficit at day 20 being 22.6%. Moreover, the density of synaptic junctions stained by ethanolic phosphotungstic acid per unit area of the radiatum layer and the lacunosum-molecular layer of the neonatal rat hippocampus was significantly reduced at day 20. It was suggested that the low-power diode laser irradiation affected the development of synapses in the neonatal rat brain.

Age Factors↗
PubMed1996

Ultrastructure of the organic matrix of embryonic avian bone after en bloc reaction with various electron-dense 'stains'.

The morphological distribution of noncollagenous proteins in bone matrix is poorly known, chiefly because their amount changes with the type of bone, and they are not easily recognizable under the electron microscope. The aim of this study was to obtain further ultrastructural data on these proteins and on their relationships with collagen fibrils and the calcification process. Specimens of woven bone from the upper metaphysis of the tibia of chick embryos were examined after en bloc treatment with acetone solution of KMnO4, or aqueous or alcoholic solutions of phosphotungstic acid or uranyl acetate. Untreated specimens were used as controls. Untreated, decalcified, and decalcified and stained sections were examined. As expected, the results showed that the woven bone matrix of the chick embryo consists of irregularly oriented collagen fibrils which outline wide interfibrillary spaces. These contain calcification nodules at the calcification front and are full of irregularly oriented crystals in completely calcified matrix. These spaces contain greater than expected amounts of filamentous or granular organic structures (probably corresponding to noncollagenous proteins), which are in a close relationship with needle- and filament-like crystals. The collagen fibrils are calcified to a lower degree than the interfibrillary spaces, and often appear uncalcified, or decalcified, when the interfibrillary spaces do not. This suggests that either the former calcify after the latter, or the respective mineral phases differ in their degree of solubility, so that the decalcification of collagen fibrils precedes that of interfibrillary spaces. The results confirm that woven bone contains an amount of interfibrillar noncollagenous material greater than lamellar bone, and show that woven bone calcification occurs not only in and on collagen fibrils, but chiefly between them, in relationship with noncollagenous filamentous material.

Animals↗
PubMed1980

Molecular sieve in renal glomerular and tubular basement membranes as revealed by electron microscopy.

Bovine glomerular basement membrane (GBM) was isolated and purified according to a modification of Spiro's method. Rat and bovine tubular basement membranes (TBM) were isolated and purified by sonic disruption or by the method of Carlson et al. Electron microscopic studies on the ultrastructure of GBM and TBM were performed after negative staining with 1% phosphotungstic acid solution, pH 7.3. When negatively stained, GBM and TBM were seen as fragments varying in size. The surface of the membranes showed a characteristic felt-like or spongy appearance. At higher magnification, GBM and TBM showed a fine meshwork composed of strands and pores which three-dimensionally resembled a crystal lattice. Pores were fairly uniform in size and shape. They were round, oval or polygonal in shape. Some of the pores were elongated to form short straight or bent channels. Strands were also uniform in diameter and surrounded a pore or channel. For an average of 50 pores, the long dimension was 3.1 +/- 0.6 nm and the short dimension 2.5 +/- 0.3 nm in bovine GBM, 3.8 +/- 1.2 and 2.5 +/- 0.7 nm in bovine TBM, and 4.9 +/- 1.5 and 2.8 +/- 0.6 nm in rat TBM, respectively. The strand was 1.8 +/- 0.3 nm in diameter in bovine GBM, 2.5 +/- 0.6 nm in bovine TBM and 3.7 +/- 0.7 nm in rat TBM for an average of 50 strands. The diameters of the pores were less than or close to the short axis of an albumin molecule. It was concluded that renal GBM and TBM were molecular sieves composed of pores and strands.

Animals↗
PubMed1994

Fibrin content of carotid thrombi alters the production of embolic stroke in the rat.

BACKGROUND AND PURPOSE: Mechanical denudation of the endothelium of the carotid artery in animals produces a nonocclusive thrombus, but the brains of these animals have not been examined for the presence of embolic stroke. METHODS: The endothelium of the right carotid artery of 16 Wistar rats was denuded using a balloon catheter. Phosphotungstic acid hematoxylin (PTAH) staining and scanning electron micrographs of the nonocclusive thrombi in the carotid arteries were compared with those produced by photochemical methods, and brains were examined for infarcts. RESULTS: Although nonocclusive thrombi were present in the carotid arteries of 4 of 4 rats killed at 4 hours and in 8 of 12 killed at 24 hours, neither cerebral infarcts nor emboli were seen in the 14 brains evaluated by light microscopy. PTAH demonstrated a high fibrin content in the thrombus produced by the endothelial denudation, with almost no fibrin seen in photochemically induced thrombi. Scanning electron microscopy confirmed dense networks of fibrin in the thrombi produced by balloon denudation. CONCLUSIONS: The composition of a nonocclusive thrombus may determine the embolic potential of this thrombus. A low fibrin content in a nonocclusive platelet thrombus may enhance the embolic potential. This suggests that platelet inhibition may also be indicated in patients with carotid artery disease who are being treated with anticoagulant.

Animals↗
PubMed2000

A new model of cerebral microthrombosis in rats and the neuroprotective effect of a Rho-kinase inhibitor.

BACKGROUND AND PURPOSE: The aim of this study was to develop a new model of stroke based on endothelial damage and thrombotic occlusion in a perforating artery, leading to small cerebral infarcts and neurological deficits in rats. Moreover, the neuroprotective efficacy of fasudil, a rho-kinase inhibitor, was investigated in this model. METHODS: Fifty-six male Sprague-Dawley rats were used in the present study. Rats were anesthetized with sodium pentobarbital, and 100 microg of sodium laurate was injected into the left internal carotid artery on days 1 and 3. The thrombus induction and consequent of ischemic brain damage were examined by histopathological analyses and neurological deficit scoring in a posture reflex test. To investigate the neuroprotective effects of fasudil, 1 or 10 mg/kg was administered intraperitoneally 5 minutes after the first injection of sodium laurate and once daily thereafter on the following 2 days. RESULTS: One hour after the injection of sodium laurate, microscopic examination of phosphotungstic acid hematoxylin-stained sections (n=5) revealed that microthrombi containing fibrin strands obstructed the perforating arteries in the ipsilateral hemisphere. Under a transmission electron microscope (n=6), endothelial cells appeared exfoliated and the vascular lumen was obstructed by a thrombus composed of degranulated platelets, fibrin, leukocytes, and erythrocytes. No evidence of endothelial cell damage or thrombus could be found in the ipsilateral side of the pial artery (middle cerebral artery). Twenty-four hours after the second injection of sodium laurate (day 4), 13 of 15 rats (86.6%) showed mild to severe neurological deficits. Multiple small cerebral infarcts were observed in the hippocampus, cortex, and thalamus. Treatment with fasudil (1 and 10 mg/kg, n=15 each) resulted in a significant improvement in neurological deficits. Fasudil also significantly reduced the area of cerebral infarction. CONCLUSIONS: We present a new model of stroke in rats, in which the perforating arteries are selectively occluded by microthrombi. This model is useful to investigate the pathophysiology and treatment of small cerebral infarction, which is caused by perforating arterial occlusive diseases such as lacunar infarcts. Fasudil may be beneficial in the treatment of acute ischemic stroke.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1979

Ultrastructural study of the protein-polysaccharides distribution in enamel from rat incisors.

The distribution pattern of the protein-polysaccharides in the rat incisor enamel was similar whether stained for glycoproteins (phosphotungstic acid) or for proteoglycans (alcian blue, bismith nitrate). Except for the holes at the dentino-enamel junction, where these components seem to exist as amorphous gel, protein-polysaccharides are closely associated to the proteinic fibrous matrix in rods and interrods structures as in prism sheaths where they are particularly abundant.

Animals↗
PubMedDate not supplied

Eosinophilic granulated cells comprising a tumor in a Fischer rat.

A systemic tumor developed in multiple organs, including spleen, bone marrow, lymph nodes, liver, ovaries, and thoracic and lumbar vertebrae, of a female F344Du/Crj rat. The tumor was composed of round to indented mononuclear cells containing abundant large eosinophilic granules in the cytoplasm. The peripheral blood smear revealed that the large granules in the cytoplasm of the tumor cells were stained basophilic with Giemsa, suggesting a basophil or mast cell origin. However, these granules did not show metachromasia with toluidine blue and were stained blue to dark blue with phosphotungstic acid hematoxylin. Cellular morphology and characteristics in the specific stains of tumor cells suggested the development of a tumor of globule leukocytes in a F344Du/Crj rat.

Animals↗
PubMed1986

A neoplasm of globule leukocytes in a cat.

A neoplasm involving the ileo-cecal-colic junction, thymus, and tracheobronchial lymph nodes of a 7-year-old domestic cat was composed of dense sheets of round to oval mononuclear cells with oval to indented nuclei, moderate amounts of cytoplasm, and variable numbers of round eosinophilic granules. These granules are brown to black in phosphotungstic acid hematoxylin-stained sections and stain variably with the periodic acid-Schiff stain. They are 0.8 to 1.5 micron in diameter, limited by a single unit membrane, and have variable electron density. Light microscopic cellular morphology and staining characteristics as well as ultrastructural features of these cells are consistent with feline globule leukocytes. Morphologic features of the neoplastic cells in the present case are similar to those of the only other reported neoplasm of globule leukocytes which also involved the intestine of a cat.

Animals↗
PubMed1975

The coating of mouse myocardial cells. A cytochemical electron microscopical study.

The coating of mouse myocardial cells has been investigated with a variety of cytochemical methods. The coating of the surface membrane gives a positive reaction with ruthenium red, colloidal thorium, phosphotungstic acid (PTA) at low pH, silver methenamine after periodic oxidation (PA-silver technique) and with silver proteinate after periodic oxidation and thiocarbohydrazide treatment (PA-TCH-silver technique). The coating of the T system gives almost similar results. The nexuses do not react with PTA nor with the PA-silver and PA-TCH-silver techniques, but they are strongly stained with ruthenium red which reveals periodic structures in their gaps. The specificities of the colloidal thorium technique and PAT staining have been tested by chemical treatments (methylation, acetylation, saponification), enzymatic digestions (pronase, trypsin, hyaluronidase, neuraminidase) and carbohydrate extractions (with 0.1 N NaOH and 0.05 M H2SO4). These cytochemical data indicate, considering the specificity of the reactions, that the coating of the membrane surface and the T system contains polyanionic groups. A part of them, at least, would belong to a carbohydrate-containing material (glycoproteins), whereas at the level of nexuses the sugar residues would probably be absent.

Animals↗
PubMed1977

Polyethyleneimine as tracer particle for (immuno) electron microscopy.

Polyethyleneimine (PEI) is proposed as a tracer for use in electron microscopical investigations. Relative small molecules are available (molecular weight 600-60,000). PEI is soluble in water; it is not visible in the electron microscope without further treatment, but can easily be detected as a particle by contrastting it with phosphotungstic acid or OsO4. Using PEI of a molecular weight of 40,000, particles of 10 nm diameter can be produced. The strong cationic character of PEI results in electrostatical binding to anionic sites. Hence perfusion and immersion of tissues with PEI of various molecular weights offers possibilities to either study the location of anionic sites or pathways of transport. Anionic sites could be demonstrated in the normal and pathologic glomerular basement membrane. Work on the use of PEI as a marker particle in immunoelectronmicroscopy is in progress.

Animals↗
PubMed1983

Localization of antibody binding sites in ultrathin sections of unembedded frog retinal tissue.

Much ultrastructural detail is retained in tissue fixed only with aldehydes and subsequently air-dried after suspension in a polyvinyl acetate emulsion. The latter provides an external support only, but permits ultrathin sectioning; thus, an exposure of intracellular contents for potential immunocytochemical reactions is achieved. Sections of unembedded frog retina so prepared have been studied with success. The tissue was incubated first with a rabbit antiserum prepared against gradient purified bovine rod outer segments. Following incubation, reacted sites were labeled with ferritin-conjugated goat anti-rabbit IgG and stained with phosphotungstic acid. Intense labeling of the rod outer segments was clearly achieved, whereas the cone outer segments were without label. Other parts of the retina, including the ellipsoid region of both rods and cones, were also without significant label. These regions provided an intrinsic control for the specificity of the antiserum and established the validity of the general technique.

Animals↗
PubMed1986

Negative-staining autoradiography: a new technique for ultracryotomy utilizing an interposed film.

A new radiocytochemical technique is reported for ultrastructural localization of diffusible substances, using negatively stained ultra-cryostat sections. A sheet of film interposed between the cryostat section and the emulsion layer has rendered negative-staining autoradiography (NSA) practical. The rationale of NSA is that the film completely shields the section from all moisture-producing autoradiographic processes, so that phosphotungstic acid (PTA) can stain the section either before or after autoradiography (ARG), without the possibility of ultrastructural damage by alkaline solutions, interference between PTA and photoprocessing compounds, and superimposed images of a gelatin layer stained with PTA. As a model to demonstrate the newly developed procedure of NSA, rat brains were labeled with [125I]-triiodothyronine, fixed with tannic fixative, immersed in a cryoprotectant, frozen in liquefied propane, and cryostat sectioned. The resulting higher yield of radioactivity (85%) on the section was confirmed by a radiation counter. The retention rate was approximately 20% greater than that of conventional sections. Developed silver grains were found on synaptic vesicles and mitochondria in the polymorphic layer of the dentate gyrus. In this report we will also discuss the problems associated with cryostat sectioning of fresh tissues, the concept of ARG resolution, the distribution pattern of developed silver grains, and the possible applications of NSA.

Animals↗
PubMed2003

Plasma distribution of apoA-IV in patients with coronary artery disease and healthy controls.

Recent studies showed lower apolipoprotein A-IV (apoA-IV) plasma concentrations in patients with coronary artery disease (CAD). The actual distribution of the antiatherogenic apoA-IV in human plasma, however, is discussed controversially and it was never investigated in CAD patients. We therefore developed a gentle technique to separate the various apoA-IV-containing plasma fractions. Using a combination of precipitation of all lipoproteins with 40% phosphotungstic acid and 4 M MgCl2, as well as immunoprecipitation of all apoA-I-containing particles with an anti-apoA-I antibody, we obtained three fractions of apoA-IV: lipid-free apoA-IV (about 4% of total apoA-IV), apoA-IV associated with apoA-I (LpA-I:A-IV, 12%), and apoA-I-unbound but lipoprotein-containing apoA-IV (LpA-IV, 84%). We compared these three apoA-IV fractions between 52 patients with a history of CAD and 52 age- and sex-matched healthy controls. Patients had significantly lower apoA-IV levels when compared to controls (10.28 +/- 3.67 mg/dl vs. 11.85 +/- 2.82 mg/dl, P = 0.029), but no major differences for the three plasma apoA-IV fractions. We conclude that our gentle separation method reveals a different distribution of apoA-IV than in many earlier studies. No major differences exist in the apoA-IV plasma distribution pattern between CAD patients and controls. Therefore, the antiatherogenic effect of apoA-IV has to be explained by other functional properties of apoA-IV (e.g., the antioxidative characteristics).

Adult↗
PubMed1975

The synaptinemal complex in Rhoeo spathacea.

The synaptinemal complex in meiocytes of Rhoeo spathacea is described. Unpaired zygotene chromosomes do not exhibit well defined axial cores under the ordinary fixations of electron microscopy and appear diffuse. However, the axial core is defined by ethanolic phosphotungstic acid (PTA) although it does not respond to uranyl-EDTA-lead. Thus the core appears to contain histone but not RNA and presents a condition which is modified later in pairing when lateral elements of the synaptinemal complex respond positively to both tests. The total number of attachments of synaptinemal complexes to the nuclear envelope was determined in several nuclei from serial sections. Eleven of the twelve possible attachments were found in one nucleus. It thus seems certain that all must be so attached. In the same manner all chromosomes can be seen to have an attachment to a chromocentre. Chromocentres are often very large and compound in that two kinds of heterochromatin can be distinguished. These states of chromatin within the chromocentre are considered to be a function of the degree of condensation. Segments of synaptinemal complexes are distributed randomly through sections of pachytene nuclei and long uncoiled segments of complexes are frequently found in or near the centres of median nuclear sections. Synaptinemal complexes are also found in chromocentres. Our findings suggest that on completion of pairing, which begins distally, homologous chromosomes in Rhoeo are paired throughout their entire lengths, rather than in small terminal segments only.

Cell Division↗
PubMed1977

Ultrastructure of the adrenal cortex in frozen thin sections.

Pieces of both unfixed and fixed rat adrenal cortex were frozen and cut with a cryoultramicrotome. In unstained frozen sections of unfixed adrenals patches of heterochromatin were seen in the nucleus, while numerous round uniformly dense structures, the mitochondria, could be identified in the cytoplasm. Lipid droplets appear as small ligh round areas. Staining the sections with potassium permanganate, phosphotungstic acid, or uranyl acetate, or immersing them in glutaraldehyde, increased the contrast of cell organelles. In frozen sections of fixed material, membranes became visible in negative contrast. Mitochondria display vesicular cristae, whereas the Golgi complex, lysosomes and peroxisomes are easily recognizable. The possibility of cutting frozen sections of unfixed material with reasonable morphological preservation allows the fine-structural localization of steroids in adrenal cells after applying autoradiographic emulsion to the sections.

Adrenal Cortex↗
PubMed1978

Membranes in lupin root nodules. II. Preparation and properties of peribacteroid membranes and bacteroid envelope inner membranes from developing lupin nodules.

Peribacteroid membranes and bacteroid envelope inner membranes have been isolated from developing lupin nodules. Isolation of the peribacteroid membranes was achieved by first preparing membrane-enclosed bacteroids free from other plant organelles or membranes. The peribacteroid membranes were then released by osmotic shock and purified by centrifugation to equilibrium on sucrose gradients. The bacteroids were broken in a pressure cell and the bacteroid envelope inner membranes were isolated using sucrose gradient fractionation of the bacteroid total envelope preparation. The density of the peribacteroid membranes decreased during the period of development of N2-fixation in lupin nodules from 1.148 g/ml for nodules from 12-day plants to 1.137 g/ml for nodules from 18-day plants. The density of the bacteroid envelope inner membranes from nodules from 18-day plants was 1-153 g/ml. The identity and homogeneity of the isolated membranes was established, by comparison with membranes in intact nodules, using phosphotungstic acid and silver staining of thin sections and particle densitites on faces of freeze-fracture replicas of the membranes. Analyses for NADH oxidase and succinate dehydrogenase, spectral analyses and gel-electrophoretic analysis of proteins were also used to characterize the membrane and soluble protein fractions from the nodules. The ratio of lipid to protein was 6.1 for the peribacteroid membranes and 2.5 for the bacteroid envelope inner membranes. Leghaemoglobin was localized in the plant cytoplasm in lupin nodules and not in the peribacteroid space.

Cell Fractionation↗