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PubMed1983

Microscopic studies of Candida albicans and Torulopsis glabrata after in vitro treatment with bifonazole. Light and scanning electron microscopy.

1-[(4-Biphenylyl)-phenylmethyl]-1H-imidazole (bifonazole, Bay h 4502, Mycospor) preferentially inhibited yeast-hypha conversion in the dimorphic fungus Candida albicans. As observed by phase-contrast and scanning electron microscopy the morphology of C. albicans yeast cells and Torulopsis glabrata cells markedly altered after treatment with bifonazole. Whereas control cells were mostly occurring singly, the bifonazole-treated cells did not separate, resulting in the formation of chains and clusters of interconnected cells. Furthermore, in contrast with control cells bifonazole-treated cells of C. albicans and T. glabrata were almost spherical in shape. Evidence is presented by means of fluorescence microscopy that bifonazole-treated yeast cells of C. albicans frequently lacked septa and often contained irregular deposition of chitin.

Antifungal Agents↗
PubMed1992

Microvillus form of focal anchorage in human Chang liver cells rounded by antiporter activation: scanning electron microscopy profiles and evidence of traction origin.

Na+/H+ antiporter activation in human Chang liver cells produces a flat-to-round (FTR) change in cell shape with gross reduction in cell profile area. Scanning electron microscopy (SEM) vividly displays a third phenomenon, viz., the development of focal microvillus anchors. Reduction in cell profile area concomitant with the development of this microvillus form of focal anchorage is quantitated by on-line image analysis during SEM examination. The reduction in profile area is corroborated by spectrophotometric digitization in light microscopy. Transmission electron microscopy (TEM) of rounded cells shows large endocytic channels and endosomes consistent with the observation of internalization of fluoresceinated-dextrans (FDx) of a diverse range of sizes, from 4,400 to 2,000,000 molecular weight, with cell rounding. Concomitant endocytosis of this magnitude indicates massive plasma membrane internalizations which could explain the very considerable profile area reduction and suggest that the microvillus anchors are probably traction processes. Antiporter mediated rounding (AMR) provides a highly reproducible and simple model for the production of anchoring microvilli ('filopodia') whereby they can be further explored.

Carrier Proteins↗
PubMed1983

[Albumin receptors on isolated human hepatocytes. Studies by phase contrast and scanning electron microscopy].

In the present work a minibead method has been employed to study human hepatocyte receptors for albumin-coupled latex particles. The cell-latex reaction was observed in both phase contrast and scanning electron microscopy. It has been demonstrated that human hepatocytes exhibit bindings sites for different forms of glutaraldehyde-treated albumin, and the ligand has proved to be species-nonspecific. The albumin binding activity is temperature-dependent and can be only partially blocked by preincubation with free, glutaraldehyde-treated protein, unless hepatocytes are pretreated with vinblastine sulfate. The possible biological role of these receptors as regards the infection due to the hepatitis B virus (that shares common, but not identical binding sites) is also discussed.

Humans↗
PubMed1995

Improved methods for preserving macromolecular structures and visualizing them by fluorescence and scanning electron microscopy.

To determine the optimal procedures to preserve cytoskeletal and other macromolecular structures for microscopic studies we have evaluated the effects of various methods to extract cultured cells. In this report, we compare results using different fixatives, crosslinking reagents, and permeabilization methods on (1) the labeling of cells for fluorescence microscopy with phalloidin or antibody against tubulin; and (2) the morphological preservation of macromolecular structures for scanning electron microscopy. Maximal labeling of F-actin with phalloidin was obtained by fixing cells in 4% paraformaldehyde (PFA) and labeling the unextracted cells with methanolic phalloidin, whereas maximal labeling of tubulin required prefixation with either PFA or the bifunctional protein crosslinking reagent, dithiobis (succinimidylpropionate) (DSP) and extraction with ethanol or Triton in a high salt buffer. However, for both qualitative and quantitative light and electron microscopic studies of intracellular macromolecular structures, prefixation with DSP and extracting with Triton X-100 in a stabilizing buffer is the overall method of choice for both labeling and morphological studies. Although other methods provide maximal labeling or preservation of specific structures, this method provides excellent preservation of morphological structure while allowing proteins to be preserved and labeled by specific probes.

Actins↗
PubMed1990

Different phenotypes of cultured microvessel endothelial cells obtained from bovine corpus luteum. Study by light microscopy and by scanning electron microscopy (SEM).

Morphological heterogeneity has not been documented for cultured endothelial cells isolated from the microvascular bed of any organ. As the corpus luteum depends on a rich microvascularization, endothelial cells were dislodged from developing corpora lutea by mechanical dissection followed either by collagenase digestion or by no digestion. Cell separation was carried out by Percoll density centrifugation. Although the yield of intact cells was higher with collagenase treatment than without, successful endothelial cell cultures were only established when cells remained untreated. Viewed by light microscopy after an average lag phase of 10 days, five different phenotypes of endothelial cells were found under similar simple culture conditions: isomorphic epithelioid, polymorphic epithelioid, spindle-shaped, round, and phase-dense phenotypes. Monolayers appeared within 2-4 weeks. After an additional period of 2-4 weeks, tubular forms with a specific pattern were noted for types 1-3, the so-called pseudotubular forms for type 4, and none for type 5. Cell types differed in their cytochemical and immunocytochemical responses. Examined by SEM, type 1 displayed a more conspicuous surface anatomy than type 2. Types 3-5 demonstrated striking cell processes that were characteristic of each type. Tubular forms of types 1 and 2 showed cell borders and a marked increase in surface specializations, whereas tubular forms of type 3 lacked detectable cell borders in the absence of a striking surface anatomy. Pseudotubular forms of type 4 developed no particular spatial organization. Thus, for the first time, morphological evidence is provided that different endothelial cell types are obtained from diverse segments of the microvascular bed.

Acid Phosphatase↗
PubMed1981

Behaviour and pathogenicity of Trichomonas vaginalis in epithelial cell cultures: a study by light and scanning electron microscopy.

The behaviour and pathogenic effects of Trichomonas vaginalis in mammalian cell cultures were studied using light microscopy and scanning electron microscopy. Six hours after inoculation of the parasites into the cell cultures about 10% of the epithelial monolayer was destroyed. The parasites adhered to the epithelial cells, developed an amoeboid morphology, and crawled over and under the monolayer of cells. These observations suggest that the adhesiveness, amoeboid morphology, and motility of T vaginalis may be important mechanisms in the injury caused to the vaginal epithelium.

Animals↗
PubMed1985

Optical and scanning electron microscopy in the single osteoclast resorption assay.

The present studies relate to the single or isolated osteoclastic resorption function assay which we introduced in 1983 to overcome objections to assays based upon measurements of calcium release from bones, in which it was never strictly controlled whether the mechanism involved the destruction of bone with the formation of classical Howship's lacunae. The method may prove to be quite popular in the near future and has already been adopted by other research groups. In previous work, we had utilised stereophotogrammetry of scanning electron micrographs to measure the depth, volume and other parameters of the individual lacunae. However, increasing experience with the method has suggested that we can await a wide range of biological variability in single cell function in any one experiment. We have therefore tested other methods from which data could be obtained more rapidly to permit a better statistical analysis, albeit with reduced accuracy, of each resorption complex. The main aim of the studies reported here was to evaluate various methods of optical and scanning electron microscopy that can be used for the visualization of osteoclasts and their associated resorption lacunae generated in vitro in slabs of dentine and bone. Optical microscopy was found to be complementary to SEM, enabling vital microscopy of unstained and stained cells. In particular, oblique illumination LM and tandem scanning reflected LM (TSRLM) proved to be of paramount value for this purpose. Fixed coated specimens could be most rapidly scanned for resorption lacunae using darkfield reflected LM or TSRLM.

Animals↗
PubMed1993

The amelocemental junction in young premolar teeth. A replica study by scanning electron microscopy.

To investigate the validity of traditional data on the frequency of various relationships of root cementum to coronal enamel at the amelocemental junction, the relationship in 50 young premolar teeth extracted on orthodontic indications was documented, using replica techniques for scanning electron microscopy (SEM), combined with polarization microscopy of thin ground sections (< 40 microns). Of 200 surfaces, 30 were excluded because of damage during specimen preparation. Replicas of the other 170 surfaces were photographed at 20x and 80x in SEM and classified as having an edge-to-edge relationship of enamel to cementum in 76% and an overlap of cementum onto the cervical enamel in 14%. What appeared to be a gap between enamel and cementum at low magnification (10%) was seen at high magnification to be a ditch or trough in the cementum: no gaps with exposed dentin were observed in SEM. Polarization microscopy of ground sections of the original specimens confirmed the SEM classification. In this material, the edge-to-edge relationship of enamel to cementum predominated: overlapping of cementum onto the enamel was less prevalent than previously reported. The methods applied in this study circumvented some major sources of artefact formation due to shrinkage during specimen preparation, which may previously have caused misinterpretation of 'gap' relationships.

Bicuspid↗
PubMed1991

[A method of phase-contrast, fluorescence and scanning electron microscopy of one and the same cultured cells undergoing cryogenic action].

Cells cultured on glass substrates have been investigated by phase contrast method. After freeze-thawing the same cells were stained for identification of damage by fluorescent dyes and studied by the phase contrast method, fluorescence and scanning electron microscopy. The method has revealed correlations between the damage of cell surface and the cell damage manifestation at morphological level.

Animals↗
PubMed1979

Surface characteristics of malignant human urinary bladder epithelium studied with scanning electron microscopy.

Biopsies were obtained from 28 male and seven female patients with cystoscopically and histologically confirmed urinary bladder carcionoma. Tissue specimens for light microscopy and scanning electron microscopy (SEM) were obtained from the tumours as well as from cystoscopically tumour-free mucosa of the dome, the lateral walls and the trigone of the urinary bladder and also from the proximal urethra. Certain surface structural changes, which seem to represent malignancy, present in different regions of the urinary bladder were detected by SEM, although not visible by light microscopy.

Aged↗
PubMed2006

Quantitative and reliable in vitro method combining scanning electron microscopy and image analysis for the screening of osteotropic modulators.

The increased generation and up-regulated activity of bone resorbing cells (osteoclasts) play a part in the impairment of bone remodeling in many bone diseases. Numerous drugs (bisphosphonates, calcitonin, selective estrogen receptor modulators) have been proposed to inhibit this increased osteoclastic activity. In this report, we describe a pit resorption assay quantified by scanning electron microscopy coupled with image analysis. Total rabbit bone cells with large numbers of osteoclasts were cultured on dentin slices. The whole surface of the dentin slice was scanned and both the number of resorption pits and the total resorbed surface area were measured. Resorption pits appeared at 48 h and increased gradually up to 96 h. Despite the observation of a strong correlation between the total resorption area and the number of pits, we suggest that area measurement is the most relevant marker for osteoclastic activity. Osteotropic factors stimulating or inhibiting osteoclastic activity were used to test the variations in resorption activity as measured with our method. This reproducible and sensitive quantitative method is a valuable tool for screening for osteoclastic inhibitors and, more generally, for investigating bone modulators.

Animals↗
PubMed2000

Scanning electron microscopy and fluorescent in situ hybridization of experimental Brachyspira (Serpulina) pilosicoli infection in growing pigs.

Two groups of six 8-week-old pigs were challenged with 1x10(9) cfu Brachyspira (Serpulina) pilosicoli or Serpulina intermedia daily for 3 consecutive days to study the pathology of porcine colonic spirochetosis by scanning electron microscopy (SEM) and fluorescent in situ hybridization (FISH) with oligonucleotide probes targeting ribosomal RNA specific for B. pilosicoli and the genus Brachyspira/Serpulina. Six pigs served as noninoculated controls. The animals were euthanatized successively between postinoculation days 14 and 24. B. pilosicoli was reisolated in feces from all of the inoculated pigs; however, only two pigs developed transient watery diarrhea. S. intermedia was reisolated from four of the inoculated pigs, but clinical signs were not observed. Gross examination of the B. pilosicoli-infected pigs revealed dilated large intestines with a hyperemic mucosa, whereas the large intestines of the S. intermedia-inoculated pigs and the control pigs appeared normal. SEM examination of B. pilosicoli-infected pigs revealed degenerated epithelial cells and spirochetal colonization of the colonic mucosa in four pigs. By FISH, B. pilosicoli cells were found colonizing and invading the surface epithelium and the crypts in all the pigs. Spirochetal crypt colonization markedly exceeded the occurrence of spirochetes on the mucosal surface. SEM examination of S. intermedia-inoculated pigs revealed no abnormalities, and Serpulina cells were detected only sporadically in the otherwise normal-appearing mucosa of four pigs by FISH. The results provide further evidence that B. pilosicoli is associated with colitis in pigs, although the gross lesions are mild. The spirochete is capable of colonizing the large intestine, inducing mucosal damage, invasion of the crypt and surface epithelium, and focal infiltration of the lamina propria. In addition, the study shows the applicability of FISH for specific identification of B. pilosicoli in formalin-fixed tissue.

Animals↗
PubMed1981

The application of scanning electron microscopy to the study of the cytoskeleton of cells in culture.

This paper reviews the methods available for studying the structural organization of the cytoskeleton of cells in culture. These are transmission electron microscopy of whole cells, immunofluorescence, and detergents extraction procedures. Published methods for detergent extraction are critically reviewed with special emphasis on those factors that are most relevant for the preparation of cytoskeletons for observation by scanning electron microscopy. These include the type of detergent, the use of chemical crosslinking reagents or specially formulated buffers to stabilize the cytoskeleton during extraction, the osmotic and salt composition of the extraction medium, the use of cool sputter coaters, and the application of minimal amounts of metal during coating. Finally, examples are given of the types of information about the three dimensional organization of the cytoskeleton that can be obtained by SEM.

Animals↗
PubMedDate not supplied

Scanning electron microscopy as a diagnostic procedure in giant cell arteritis.

Forty-five consecutive patients (32 women and 13 men) underwent biopsy of the temporal artery because of suspected giant cell arteritis. Their ages ranged from 38 to 84 years, mean 68.1 years. Five patients (11.1%) four of them women, were found to be affected by the disease. Their ages ranged from 54 to 80 years, mean 69 years. Clinical and laboratory findings included elevated erythrocyte sedimentation rate, prolonged fever, continuous headache, sudden onset of unilateral blindness, intermittent mandibular claudication, severe anemia and myalgia. None of these, whether present in isolation or in various combinations, were of significant diagnostic value. All biopsies were examined both by light microscopy and by scanning electron microscopy. The former examination took about 5-7 days to complete, and the latter about 3 hours. Light microscopy studies showed that 46.6% of the arterial biopsies were normal, 42.3% were arteriosclerotic and 11.1% (5 specimens) were characteristic of giant cell arteritis. Scanning electron microscopy revealed that the biopsies obtained from all five patients found to have temporal arteritis displayed the "occlusive" pattern: the three-laminar appearance of the artery was markedly distorted or lost, the internal elastic lamina was barely detectable, and the densely hypertrophied media and intima filled the arterial lumen, virtually obliterating it. We conclude that scanning electron microscopy is a quick and accurate procedure for diagnosis of temporal arteritis and that positive findings may be taken as an indication for immediate steroid treatment.

Adult↗
PubMed1988

[Periosteal bone resorption in the area of the metaphysis of growing bone as a precursor of epiphyseal injuries. A polarization optical and scanning electron microscopy study].

Macerated epiphyses of the growing infant bone exhibit an irregular surface which can already be observed at the macroscopic level. Polarizing microscopy demonstrates extended zones of bone resorption on the cortical surface, deductable from numerous lacunae of Howship. Ensuing experimentally induced epiphysiolyses, cortical bone fragments adhering to the inner surface of the periosteum are demonstrable by scanning electron microscopy. Having been torn out of the bone together with the periosteum, these cortical fragments leave corresponding defects on the bone surface. Due to the remodelling of the bone, involving the readjustment of the shape of the extremity, the attachment of the periosteum is relatively poor in the metaphyseal region of the growing bone. The influence of pathological forces can therefore easily cause a detaching of the periosteum in this region. The latter results in a significant weakening of the epiphyseal fastening in the zone between the epiphyseal plate and the metaphysis. The patterns of injury in the region of the growth plate are therefore essentially determined by the varying attachment of the periosteum to the metaphysis.

Adolescent↗
PubMedDate not supplied

Scanning electron microscopy of normal cells, dyskaryotic cells and malignant cells exfoliated from the uterine cervix.

The same exfoliated benign, dyskaryotic and malignant cervical cells were observed by light microscopy and scanning electron microscopy in order to investigate the reflection of the course of malignancy in the surface ultrastructure. The surface ultrastructural features (microridges and microvilli) of normal squamous cells and metaplastic cells showed regularities, with an approximately even thickness, length and distribution of microvilli. In dyskaryotic cells of the superficial type, the microridgelike structure was retained, and some regularity of the microvilli remained. However, in dyskaryotic cells of the deep-layer type, the microvillous surface ultrastructure had a pronounced irregularity and diversity. In malignant cells, there was unevenness in the projections and in the thickness of the cytoplasm, and these cells presented, as a whole, a bizarre appearance. The surface ultrastructure of malignant cells thus reflected a strong atypia of the cytoplasm.

Carcinoma in Situ↗