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PubMed2000

Ultrastructural and lectin-histochemical differences between the scolex/strobila and bladder teguments of the Taenia taeniaeformis strobilocercus.

The strobilocercus stage of the cat tapeworm Taenia taeniaeformis is surrounded by a single syncytial sheet of cytoplasm called the tegument. The outer membrane of the tegument covers both the scolex/strobila (S/S) and the bladder portions of the strobilocercus, but only the S/S region is resistant to intestinal digestion. It has been suggested that the glycocalyx, the surface-exposed glycoconjugates of the outer membrane, may serve to insulate underlying surface membrane components from digestion. In this study, we used lectin binding to test the hypothesis that the glycocalyx of the S/S is different from that of the bladder and that this may serve as the resistance mechanism of the S/S to digestion. Biotin-labeled lectins and an avidin-glucose oxidase detection system were applied to whole strobilocerci and to 1-microm epon-araldite plastic-embedded sections. Lectins bound to either both regions of the strobilocerci, to the S/S regions only, or did not bind at all. The restriction of some glycoconjugates to the glycocalyx of the S/S region only is consistent with our hypothesis.

Animals↗
PubMed1983

Topography and ultrastructure of LHRH- and somatostatin-containing axonal terminals in the median eminence of rats.

The topography and ultrastructure of LHRH and somatostatin nerve fibers in the median eminence in male rats were investigated with special reference to the relation between the fiber terminals and the perivascular space of the portal capillaries. Vibratome sections taken from the rostral and preinfundibular portions of the median eminence were immunostained with anti-LHRH or anti-somatostatin serum before plastic embedding for electron microscopy. In the external layer of the median eminence, the LHRH fibers did not directly contact the perivascular space, although they were in close proximity to the external surface of the median eminence. On the contrary, somatostatin fibers divided into branches in the external layer of the median eminence and terminated in direct contact with the perivascular space in the median eminence. LHRH fiber terminals contain immunoreactive granules, whereas somatostatin fibers are characterized by the presence of synaptic vesicle-like structures, vesicle ghosts and immunoreactive granules in their terminals. These topographical and ultrastructural differences might be regarded as a morphological manifestation of different mechanisms in the release of neurohormones.

Animals↗
PubMed2005

Er:YAG laser in defocused mode for scaling of periodontally involved root surfaces: an in vitro pilot study.

BACKGROUND: The Er:YAG laser may be used on periodontally involved teeth in combination with conventional periodontal therapy in order to improve the efficacy of root instrumentation. The aim of this study was to compare the effect of hand instrumentation on root surfaces of periodontally involved teeth with Er:YAG laser application. METHODS: Thirty freshly extracted, non-carious, single-rooted, periodontally diseased human teeth from adult humans with advanced periodontal disease were used in this study. The teeth were divided into three groups of 10 specimens each. Group A was treated with scaling and root planing (SRP) with curets only (control). In group B, the root surfaces were scaled with curets and then lased with an Er:YAG laser (wavelength 2.94 microm). A handpiece with a water spray was used in non-contact mode (defocused) at a distance of 1 cm from root surface. Laser parameters were set at energy of 100 to 200 mJ/pulse, with 10 Hz of frequency. In group C, the root surfaces were lased only with power settings 250 to 300 mJ/pulse and 10 Hz frequency. An epon-araldite plastic embedding technique was used for light microscopic investigation. RESULTS: Histologic findings showed significant differences between the test and control sites. In control sites, after hand instrumentation, the surface was smooth, without a cementum layer, and the dentin layer presented opened tubules. Defects on the dentin layer were also present along root surfaces. In the test sites (B, C) root surfaces revealed no thermal damage; no cracking or tissue carbonization were observed. The superficial layers of lased surfaces appeared smooth and melted without alterations. CONCLUSION: Based on these findings, it appears that it may be feasible to use the Er:YAG laser for root instrumentation without prior root planing if the proper parameters are followed.

Adult↗
PubMed2002

Processing of immunoisolated pancreatic islets: implications for histological analyses of hydrated tissue.

Routine tissue processing is usually associated with histological artifacts as a consequence of shrinkage and distortion during dehydration required for embedding. With hydrated specimens such as lung, embryonic, and tissues in hydrophilic membranes, tissue processing can induce severe artifacts that interfere with adequate microscopic evaluation. Here we present a method for embedding hydrophilic alginate-polylysine microencapsulated pancreatic tissue that combines the absence of histological artifacts with a practical tissue processing method. We found that the glycol-methacrylate (GMA)-embedding method preserved the integrity of the encapsulated tissue better than snap-freezing or paraffin embedding, but the overall quality of the hydrophilic capsules remained poor Next, we modified the GMA method by introducing gradual dehydration to investigate whether the integrity of the sectioned capsules was better maintained by a more gradual pattern of water extraction. This modification resulted in well-preserved morphological details of the hydrophilic membranes, hydrogel-cell interface, and encapsulated pancreatic tissue. Subsequent routine staining gave excellent contrast between the islet tissue and hydrophilic components, which allowed adequate quantitative histological and pathological comparisons.

Alginates↗
PubMed1996

Enhanced cytochemical detection of viral proteins and RNAs using double-sided labeling and light microscopy.

We have developed a double-sided labeling technique for detecting viral proteins or RNAs in plastic-embedded leaf tissue by immunocytochemistry or in situ hybridization, respectively, and light microscopy. The signal from the target was enhanced by double-sided labeling when compared with single-sided labeling because sections were submerged in labeling solutions with both sides accessible to antibodies or complementary RNAs. The additional label was visible during microscopic analysis. Background signal was decreased since the tissue was probed and washed under conditions where folds and creases in the tissue were minimized. This technique uses the same equipment and chemicals as for single-sided labeling, and thus adjustments for reagent expenditures are not necessary. The procedure should be applicable to animal and plant tissue.

Animals↗
PubMedDate not supplied

Correlation of audiometric data with changes in cochlear hair cell stereocilia resulting from impulse noise trauma.

In a previous experiment, after chinchillas had been exposed to impulse noise trauma, plastic-embedded surface preparations of the organ of Corti were examined with the light microscope. A consistent relationship between cochlear hair cell loss and hearing loss was not found (Hamernik et al., 1980). In the present study, four cochleas from that experiment were sectioned and examined with the transmission electron microscope to determine if their were consistent patterns of damage to the sensory cells at the ultrastructural level that would more closely correlate with the audiometric data. Alterations of the outer hair cell stereocilia were found when threshold was elevated 15 to 30 dB. The membranes of the stereocilia appeared loose and wrinkled and the stereocilia were no longer erect. In some cases, predominantly in the first row of outer hair cells, stereocilia were missing and in other cases, stereocilia were fused. Within these giant stereocilia, the rootlets of the individual stereocilia had disintegrated. Other alterations in sensory cell ultrastructure, though present, had no consistent pattern and could not be related to changes in hearing thresholds. Only the changes in the outer hair cell stereocilia appeared to correlate with hearing loss and the degree of damage was reflected in the amount of threshold elevation.

Animals↗
PubMed1989

Spontaneous papillary necrosis in the heterozygous Gunn rat.

Spontaneous papillary necrosis develops in aging heterozygous non-jaundiced Gunn rats. The lesion is situated in the subapical or mid papilla and in its earliest stages is manifest by the appearance of amorphous material in the interstitial space. This is seen in plastic-embedded sections taken from rats 6 months old. In its later stages, the accumulation of amorphous material is accompanied by loss of interstitial cells and cyst formation, but there is no associated inflammatory reaction. The largest lesions are found in the oldest rats, but even in these animals the macroscopic appearance of the papilla is normal. Similar papillary changes were not found in albino or homozygous Gunn rats, but in aging albino rats there was loss of papillary interstitial cells without accumulation of amorphous material.

Aging↗
PubMed1975

The use of an optical brightener in the study of plant structure.

An optical brightener Calcoflour White M2R New has been used to stain cell walls of higher plants. It can be used either as a vital stain for intact plants or for hand sections and plastic-embedded thin sections. Walls are brilliantly fluorescent while most cytoplasmic components are normally unstained. The brightener binds strongly to cellulose, carboxylated polysaccharides, and callose. Staining for 20 sec to 2 min in a 0.01% solution of the brightener is preferred for most purposes.

Cell Wall↗
PubMed1975

A glutaraldehyde/potassium dichromate tracing method for the localization and preservation of abdominal extra-adrenal chromaffin tissues.

The present work introduces a method for the localization in situ of the abdominal paraganglia. After treating retroperitoneal tissue blocks with a near-neutral glutaraldehyde/potassium dichromate solution following routine glutaraldehyde perfusion, intra- and extraadrenal chromaffin tissues develop a pronounced brown color from the interaction of glutaraldehyde/potassium dichromate with amines. In this manner, visualization of the abdominal extra-adrenal chromaffin organs is enhanced at the same time that cellular ultrastructure is preserved. Subsequent examination of the dichromate-reacted tissues with the electron microscope confirms that they represent the amine-rich paraganglia. This method offers an effective alternative to extensive sampling of plastic-embedded blocks for localizing peripheral chromaffin tissue and has been used to define the exact distribution of abdominal paraganglia in the rabbit.

Abdomen↗
PubMed1979

Modification of a scanning transmission electron microscope specimen holder for large section viewing.

A modification of a scanning transmission electron microscope specimen holder which permits full viewing of large plastic embedded tissue sections is discussed. The method for producing one-centimeter diameter "giant" grids is explained and the procedure for sample preparation is outlined. The modification aids the microscopist in his evaluation of tissue structural relationships by providing large areas of tissue for examination and reduces significantly the time required to prepare and examine standard 1-2 mm2 electron microscopy tissue sections. Light and electron microscopic evaluations can be made on the same tissue sections.

Microscopy, Electron, Scanning↗
PubMed1981

Ruthenium red- p-phenylenediamine staining of monolayers to facilitate handling and selection of specific cells for transmission electron microscopy.

The handling of monolayers for transmission electron microscopy has presented many problems, the main one being difficulty in visualizing the monolayers after polymerization of their plastic embedment following conventional glutaraldehyde-osmium fixation. The application of ruthenium red-p-phenylenediamine during processing intensely darkened cells which could be examined and photographed either in 95% ethanol or following Spurr monolayers when re-embedding, and permitted precise localization of monolayers within flat embedding molds when trimming and thin sectioning for transmission electron microscopy. Increased color density is the combined result of more complete retention of soluble elements during initial fixation by ruthenium red and the formation of a colored reaction product between the bound ruthenium red and osmium which is further intensified by p-phenylenediamine.

Cells, Cultured↗
PubMed1982

Vascular labelling with monastral blue B.

Vascular labelling is an established technique of experimental pathology whereby leaky vessels can be identified in vivo. A suspension of a suitable colloidal pigment is injected intravenously; the pigment is then trapped in the wall of the leaky vessels. The colloidal preparation of carbon black, which has been used for many years for this purpose, is no longer commercially available. This communication introduces a substitute: Monastral blue B which gives beautiful preparations in whole mounts, is readily visible in paraffin and plastic embedded histologic sections, has a distinctive appearance in electron micrographs, and is nontoxic in the required dosage.

Animals↗
PubMed1994

A study of criteria permitting the use of plastinated specimens for light and electron microscopy.

Plastination permits the preservation of anatomical specimens in a physical state approaching that of the living condition. We studied the possibility of using silicone plastinated fragments of spleen and pancreas for optical and electron microscopy, and found that with an adequate fixation protocol, plastinated specimens can be used for both light microscopy and ultrastructural studies. Deplastination with sodium methoxide permitted production of clean sections. Artifacts produced by plastination/deplastination could be nearly eliminated by glutaraldehyde/formaldehyde fixation. The (Biodur) silicone S10 polymer is transparent and stable in an electron beam, and plastinated tissues can be contrasted or colored similar to tissues embedded in Epon 812. In addition to being very life-like, plastinated tissues are stable and easy to handle. They can also be used for electron and light microscopic studies. This technique may also allow retrospective epidemiological studies of archived pathology specimens.

Animals↗
PubMedDate not supplied

Fungal melanin detection by the use of copper sulfide-silver.

Silver-staining procedures were investigated for their effectiveness in identifying cell wall-based fungal melanins in live and fixed plastic embedded samples, particularly 1,8-dihydroxynaphthalene (DHN) based polyketide melanins. We developed a simple and reliable melanin-staining technique based on a silver accumulation method originally published for histological demonstration of heavy metal sulfides in mammalian tissues. Copper is bound to fungal melanin followed by formation of the copper sulfide at melanin sites in fungal cell walls, which then are amplified into vivid black stains using a silver enhancement step. The method demonstrates patterns of melanization in a range of fungal hyphae and is suitable for light and electron microscopy. Albino mutant fungi and normally nonmelanized fungi do not stain with the sulfide-silver technique. Mammalian melanocytes also were labeled by the technique, indicating its universality as a melanin probe.

Animals↗
PubMed2001

Achondrogenesis type IB: agenesis of cartilage interterritorial matrix as the link between gene defect and pathological skeletal phenotype.

Achondrogenesis type IB is a lethal osteochondrodysplasia caused by mutations in the diastrophic dysplasia sulfate transporter gene. How these mutations lead to the skeletal phenotype is not known. Histology of plastic-embedded skeletal fetal achondrogenesis type IB samples suggested that interterritorial epiphyseal cartilage matrix was selectively missing. Cartilage was organized in "chondrons" separated by cleft spaces; chondrocyte seriation, longitudinal septa, and, in turn, mineralized cartilaginous septa were absent. Agenesis of interterritorial matrix as the key histologic change was confirmed by immunohistology using specific markers of territorial and interterritorial matrix. Biglycan-enriched territorial matrix was preserved; decorin-enriched interterritorial areas were absent, although immunostaining was observed within chondrocytes. Thus, in achondrogenesis type IB: (1) a complex derangement in cartilage matrix assembly lies downstream of the deficient sulfate transporter activity; (2) the severely impaired decorin deposition participates in the changes in matrix organization with lack of development of normal interterritorial matrix; and (3) this change determines the lack of the necessary structural substrate for proper endochondral bone formation and explains the severe skeletal phenotype.

Adult↗
PubMed2003

Autopsy-based assessment of extent and type of osteomyelitis in advanced-grade sacral decubitus ulcers: a histopathologic study.

CONTEXT: Decubitus ulcers constitute a serious medical problem, often encountered in association with hospitalization or institutionalization in senior citizens' or nursing homes. Potentially life-threatening sepsis has been reported to originate not only from soft tissue infection, but also from osteomyelitis as a complication of involvement of bone tissue in decubitus ulcers. OBJECTIVE: To assess the histopathology of osseous structures involved in grade IV decubitus ulcers. DESIGN: Autopsy-based histopathologic assessment of the presence and extent of osteomyelitis on os sacrum specimens from 28 deceased individuals with grade IV sacral decubitus ulcers using an undecalcified preparation following plastic embedding (staining with Goldner, Kossa modification, toluidine blue, and Giemsa). RESULTS: The histologic findings were classified in 4 types of pathomorphologic changes: type 1, decubitus ulcer confined to soft tissue, no inflammation (n = 7); type 2, decubitus ulcer involving bone, no inflammation (n = 7); type 3, decubitus ulcer involving bone, inflammation of soft tissue, no osteomyelitis (n = 1); and type 4, decubitus ulcer involving bone, presence of osteomyelitis (n = 13). Type 4 changes are further described as follows: type 4a, chronic osteomyelitis alone (n = 6); and type 4b, chronic and acute osteomyelitic changes (n = 7). More than half of the cases (n = 15) showed no inflammatory reaction within the medullary cavity (types 1-3). In all cases with osteomyelitis, inflammation was exclusively confined to the superficial parts of the os sacrum. Chronic osteomyelitis was seen in all cases in which osteomyelitis was present. In addition, mild acute osteomyelitic changes were observed in 7 cases. Severe liquefying osteomyelitis affecting deeper layers of the os sacrum was not found. Sepsis was present in 2 cases; in one of these cases, the decubitus ulcer was considered a possible source of infection. CONCLUSIONS: Our results provide evidence that in cases of grade IV decubitus ulcers, the macroscopic aspect and clinical imaging techniques may lead to an overestimation of the extent of osseous involvement. We suggest that the investigation of bone biopsies is not necessary in a considerable proportion of cases of grade IV decubitus ulcers in patients without sepsis, as the minor osseous alterations are of little consequence when establishing a therapeutic approach.

Aged↗
PubMed1999

Effects of single polychlorinated biphenyls on the morphology of cultured rat tubuli seminiferi.

The effects of single polychlorinated biphenyls (PCB) on isolated tubuli seminiferi of the rat were studied. Freshly isolated rat tubuli seminiferi were prepared according to their transillumination pattern, i.e. dark or pale. Tubuli seminiferi with the dark pattern included stages II to VIII and tubuli with the pale pattern represented stages IX to XIV and stage I of the seminiferous cycle. Afterwards, tubuli seminiferi were exposed to single polychlorinated biphenyls for 5 or 24 h in vitro. PCB 126, PCB 77, and PCB 118 were used in final parts per billion (p.p.b.) concentrations as determined by quantitative PCB analysis. Eventually, the specimens were plastic embedded, cut into semithin sections, stained, and morphology was evaluated by light microscopy. Single PCB congeners induced morphological alterations in cultivated rat tubuli seminiferi in a time- and stage-dependent manner. Effects comprised loosened intercellular contacts between germ cells and Sertoli cells as well as cellular fragmentation in the layer of round spermatids. Early spermiogenesis seems particularly susceptible to single PCB congeners in concentration of background magnitude. The target cell has still to be discovered.

Animals↗
PubMed1999

The relative extent and propensity of CD34+ vs. CD34- cells to undergo apoptosis in myelodysplastic marrows.

The paradox of peripheral cytopenias despite cellular bone marrow (BM) observed in myelodysplastic syndromes (MDS) has been associated with excessive intramedullary apoptosis of hematopoietic cells. Since MDS is regarded as a stem cell disorder, the present studies were undertaken to examine the relative susceptibility and propensity of early progenitor CD34+ cells to undergo apoptosis as compared to more maturing/matured CD34- cells. Five serial studies were performed on 4 independent groups of 36 newly diagnosed MDS patients. First, in 2 separate groups of 16 and 8 patients each, measurement of the extent of apoptosis in CD34+ and CD34- fractions of the BM aspirate mononuclear cells was carried out using independent biparametric flow cytometry methods, CD34 labeling/terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) (n = 16), and CD34 labeling/reduced uptake of nucleic acid staining dye LDS751 (n = 8). The difference in the median degrees of apoptosis in CD34+ vs. CD34- cells was not statistically significant by either technique (P = 0.583 and P = 0.674 for TUNEL and LDS751, respectively). In the next group of 4 MDS patients, a double-labeling was performed on plastic embedded marrow biopsy sections, to detect CD34 antigen with specific monoclonal antibody and apoptosis by in situ end labeling (ISEL) of fragmented DNA. Despite high overall apoptosis (56.2% +/- 18.4%), only an occasional CD34+ cell was found to be simultaneously labeled with ISEL. Finally, in the last group of 8 MDS patients, CD34+ cells were separated from CD34- cells on affinity column and cultured in serum containing medium for 4 hours. At 0- and 4-hour time points, ISEL was carried out to label apoptotic cells. In addition, a fluorometric assay was employed to estimate the activity of a proapoptotic enzyme, Caspase 3. Both the net increase in % ISEL labeled cells (apoptotic index or AI) and Caspase-3 activity were significantly lower in CD34+ cells as compared to CD34- cells (AI, 0.87% +/- 0.5% vs. 3.97% +/- 1.4%, n = 6, P = 0.028 and Caspase-3 Units/mg protein, 46.9 +/- 25.0 vs. 71.7 +/- 23.03, n = 5, P = 0.042, respectively). We conclude that when estimated in a total population of mononuclear cells, CD34+ cells and CD34- cells show comparable degrees of apoptosis. However, once separated the CD34+ fraction demonstrates lower propensity to undergo apoptosis, thereby suggesting the CD34- fraction as being a possible source for proapoptotic signaling.

Aged↗