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PubMed1995

Histology and electron microscopy of fucosidosis of the skin. Subtle clues to diagnosis by electron microscopy.

Fucosidosis is an autosomal recessive inborn error of metabolism in which fucose-containing glycolipids, glycoproteins, and oligo- and polysaccharides accumulate in tissues as a consequence of alpha-L-fucosidase deficiency. Since the detection of this entity in 1966 several cases have been described, but until now investigations of clinically uninvolved skin have not been performed. In this study we have investigated clinically normal skin obtained from a patient with fucosidosis and his healthy sister, by light and electron microscopy, to determine whether normal skin in this condition yields clues that may have prognostic relevance. We found "empty"- appearing storage vesicles in melanocytes, endothelial cells, sweat glands, and fibroblasts in the skin.

Child, Preschool↗

PubMed1979

Observations by immunofluorescence microscopy and electron microscopy on the cytopathogenicity of Naegleria fowleri in mouse embryo-cell cultures.

The destruction of secondary mouse-embryo (ME) cells by Naegleria fowleri was studied by indirect immunofluorescence with ME-cell antiserum as a specific label to trace the fate of mammalian-cell cytoplasm. The appearance of naegleria-induced cytopathic effect in the cultures coincided with the accumulation of discrete particles containing granules of ME-cell antigen within the cytoplasm of amoebae, suggesting that the organisms ingested host-cell material. In cultures containing cytochalasin B, a non-lethal inhibitor of phagocytosis by N. fowleri trophozoites failed to acquire any granular fluorescence and were not cytopathogenic. The engulfment of mammalian-cell cytoplasm by the organisms was confirmed when thin sections of naegleria-infected ME-cell cultures were examined by electron microscopy. Amoebae were seen in the process of detaching portions of cytoplasm from whole ME cells by means of distinctive ingesting pseudopodia, and fragments of mammalian-cell cytoplasm were identified within the food vacuoles of trophozoites. There was no evidence for cytotoxic disruption of ME cells before or during engulfment of these fragments. It is concluded that N. fowleri trophozoites attack and destroy cultured ME cells by a phagocytosis-like mechanism alone, without the aid of any amoeba-associated cytotoxic or cytolytic agents. The possible significance of these findings with respect to the in-vivo pathocity of N. fowleri is discussed.

Amoeba↗

PubMed1994

Quantitative and 3-dimensional analysis of Langerhans cells in basal cell carcinoma. A comparative study using light microscopy and confocal laser scanning microscopy.

We have analysed Langerhans cells (LCs) in basal cell carcinoma (BCC) and in healthy skin in 15 patients, using three different techniques: light microscopic examination of horizontal sheets, and of 6-micron-thick vertical skin sections, and confocal laser scanning microscopy (CLSM) of 25-micron-thick vertical sections. The use of CLSM enables both a quantitative and a three-dimensional (3-D) analysis of the cells in the same tissue volume. A statistically significant reduction in the relative volume of epidermal CD1a reactivity confined to tumour areas was found with CLSM. This difference was confirmed when the number of LCs in horizontal sheets were counted. In contrast, no significant reduction in epidermal CD1a+ cells was found in thin vertical sections. This is probably due to the smaller tissue sample examined, and to variations in the number of CD1a+ cells, with less cells directly overlying the tumour nests. The ratio of CD1a-expressing cells in the epidermis/dermis was significantly reduced in BCCs, compared with healthy looking skin. Few LCs were observed in tumour nests, but they were numerous in the surrounding stroma of the dermis. Three-dimensional reconstructions of CD1a+ cells in BCC revealed striking morphological changes; they had a reduced number of dendrites, and these were often short and had few branches. The results demonstrate that CLSM is a suitable technique for quantitative and morphological analysis of CD1a-expressing cells in the skin. We suggest that the alterations in LC numbers, distribution and morphology in BCC most probably are secondary to changes in the local environment.

Adult↗

PubMed1983

A method for the detection of amyloid in human tissue embedded in epon using a combination of polarizing light microscopy and electron microscopy.

This paper describes a method for demonstrating amyloid in human tissue embedded in epon using a combination of congo red staining and polarizing microscopy. The method involves the removal of the embedding medium from semi-thin sections prior to staining with congo red and examination in the polarizing microscope. Typical apple green birefringence was detected in areas of amyloid deposition; these areas were mapped for selective trimming and ultra-thin sections taken serially were examined in the electron microscope. The method was found to be reliable, easy to use and particularly of value as a sampling procedure prior to investigation in the electron microscope.

Amyloid↗

PubMed1984

Surface reflection interference microscopy: a new method for visualizing cytoskeletal components by light microscopy.

Surface reflection interference microscopy of detergent resistant residues of cultured cells stained with protein dyes can be used to obtain high resolution images of the cytoskeleton. We have compared the images obtained using different dyes and have examined the effect of four of these dyes on the visualization of different parts of the cytoskeleton in detail. The dependence of contrast of the images obtained on the illuminating numerical aperture and the wavelength of incident light was determined. Staining with Acid Yellow 36, Guinea Green B and Naphtol Blue Black produces images from the entire cytoskeleton and contrast in these images is relatively insensitive to changes in the incident wavelength. Coomassie Brilliant Blue R250 images, on the other hand, result primarily from reflection from the lower surface of the cytoskeleton and the contrast of these images is sensitive to changes in incident wavelength dropping abruptly in the region of the transmission peak of the stain. From the different spectral sensitivities of the reflection images obtained and from differential interference effects at low and high illuminating numerical apertures, we conclude that the reflection images obtained using the first three stains result from modulation of the reflection by interference effects. In contrast, in the case of Coomassie Brilliant Blue R250 the resulting image originates mainly from selective reflection of wavelength near the absorption range of the dye.

Animals↗

PubMed1994

Role of magnesium and calcium in alcohol-induced hypertension and strokes as probed by in vivo television microscopy, digital image microscopy, optical spectroscopy, 31P-NMR, spectroscopy and a unique magnesium ion-selective electrode.

It is not known why alcohol ingestion poses a risk for development of hypertension, stroke and sudden death. Of all drugs, which result in body depletion of magnesium (Mg), alcohol is now known to be the most notorious cause of Mg-wasting. Recent data obtained through the use of biophysical (and noninvasive) technology suggest that alcohol may induce hypertension, stroke, and sudden death via its effects on intracellular free Mg2+ ([Mg2+]i), which in turn alter cellular and subcellular bioenergetics and promote calcium ion (Ca2+) overload. Evidence is reviewed that demonstrates that the dietary intake of Mg modulates the hypertensive actions of alcohol. Experiments with intact rats indicates that chronic ethanol ingestion results in both structural and hemodynamic alterations in the microcirculation, which, in themselves, could account for increased vascular resistance. Chronic ethanol increases the reactivity of intact microvessels to vasoconstrictors and results in decreased reactivity to vasodilators. Chronic ethanol ingestion clearly results in vascular smooth muscle cells that exhibit a progressive increase in exchangeable and cellular Ca2+ concomitant with a progressive reduction in Mg content. Use of 31P-NMR spectroscopy coupled with optical-backscatter reflectance spectroscopy revealed that acute ethanol administration to rats results in dose-dependent deficits in phosphocreatine (PCr), the [PCr]/[ATP] ratio, intracellular pH (pHi), oxyhemoglobin, and the mitochondrial level of oxidized cytochrome oxidase aa3 concomitant with a rise in brain-blood volume and inorganic phosphate. Temporal studies performed in vivo, on the intact brain, indicate that [Mg2+]i is depleted before any of the bioenergetic changes. Pretreatment of animals with Mg2+ prevents ethanol from inducing stroke and prevents all of the adverse bioenergetic changes from taking place. Use of quantitative digital imaging microscopy, and mag-fura-2, on single-cultured canine cerebral vascular smooth muscle, human endothelial, and rat astrocyte cells reveals that alcohol induces rapid concentration-dependent depletion of [Mg2+]i. These cellular deficits in [Mg2+]i seem to precipitate cellular and subcellular disturbances in cytoplasmic and mitochondrial bioenergetic pathways leading to Ca2+ overload and ischemia. A role for ethanol-induced alterations in [Mg2+]i should also be considered in the well-known behavioral actions of alcohol.

Alcoholism↗

PubMed1999

Comparison of sensitivity of immunofluorescent microscopy to that of a combination of immunofluorescent microscopy and immunomagnetic separation for detection of Cryptosporidium parvum oocysts in adult bovine feces.

A direct immunofluorescence assay (DFA) (Merifluor; Meridian Diagnostics, Inc., Cincinnati, Ohio) was compared to an immunomagnetic separation (IMS) assay (Dynabeads; Dynal, Inc., Lake Success, N.Y.) coupled with immunofluorescent microscopy (Waterborne, Inc., New Orleans, La.) for their ability to detect low concentrations of Cryptosporidium parvum oocysts in adult bovine fecal material. IMS-DFA resulted in a 2-log-unit increase in sensitivity (10 oocysts/g) compared to DFA alone (1,000 oocysts/g). The higher sensitivity obtained with IMS-DFA resulted from testing 2 g of fecal material instead of the 13 to 19 mg of fecal material tested in the DFA; the increased sensitivity was not attributable to a higher percent recovery.

Animals↗

PubMed1974

Diagnostic electron microscopy of faeces. I. The viral flora of the faeces as seen by electron microscopy.

A method is described for examining viruses in faeces by direct electron microscopy using negative staining. The particles found in a group of patients with gastroenteritis and a group with other conditions are compared. Small particles in the range of sizes covering parvoviruses and enteroviruses were found about as frequently in each group. Many of these were probably bacteriophages. Many bacteriophages with tails of various sizes and lengths were found. Adenoviruses were found in five of eight patients from whom they were isolated. Reovirus- (or orbivirus-) like particles were found associated with some gastroenteritis patients.

Adenoviridae↗

PubMed1998

Concepts in imaging and microscopy: color image processing for microscopy.

Digital image analysis has long been applied to monochrome images from the light microscope. Using these techniques one can locate, measure, identify, and count the objects of interest in a microscope field of view. Since microscope images often contain color information of interest, it is now becoming more common to analyze the spectral content of images as well. Although most of the digital imaging techniques developed for monochrome microscopy work well with color images, there are a few considerations specific to the analysis of multispectral imagery. These include obtaining a properly balanced multicolor digital image, specifying the color components of the image, and compensating for imperfect separation among the color channels.

Color↗

PubMed1995

Evaluation of cell death in EBV-transformed lymphocytes using agarose gel electrophoresis, light microscopy and electron microscopy. I. Induction of classic apoptosis by the bile salt, sodium deoxycholate.

In this study, we examined the effect of different concentrations of sodium deoxycholate (NaDOC), a secondary bile salt, on an Epstein-Barr virus transformed human lymphoid cell line (NC-37). We found that NaDOC induces classic apoptosis in a dose-dependent manner at 0.1-0.4 mM doses, and necrosis at much higher concentrations (0.8-3.1 mM). This is the first demonstration that a bile salt can induce apoptosis in any cell type. The mode of cell death was determined using morphologic methods (light and electron microscopy) as the gold standard. Standard agarose gel electrophoretic techniques were applied to identify the "ladder" of DNA fragments that have been associated with apoptosis in certain cell types. Although DNA fragmentation was observed during the apoptotic death of NC-37 cells, we were not able to identify a "ladder" pattern of fragmentation. Two other types of cells, however, that previously have been reported to display a characteristic "ladder" pattern of DNA fragmentation, glucocorticoid-treated WEHI7.2 cells and isolated human neutrophils, did display the "ladder" pattern. This study emphasizes the need to examine morphology when identifying the mode of cell death induced by a new agent.

Apoptosis↗

PubMed2004

Spectral and lifetime fluorescence imaging microscopies: new modalities of multiphoton microscopy applied to tissue or cell engineering.

Spectral and multiphoton imaging is the preferred approach for non-invasive study allowing deeper penetration to image molecular processes in living cells. But currently available fluorescence microscopic techniques based on fluorescence intensity, such as confocal or multiphoton excitation, cannot provide detailed quantitative information about the dynamic of complex cellular structure (molecular interaction). Due to the variation of the probe concentration, photostability, cross-talking, its effects cannot be distinguished in simple intensity images. Therefore, Time Resolved fluorescence image is required to investigate molecular interactions in biological systems. Fluorescence lifetimes are generally absolute, sensitive to environment, independent of the concentration of the probe and allow the use of probes with overlapping spectra but that not have the same fluorescence lifetime. In this work, we present the possibilities that are opened up by Fluorescence Lifetime Imaging Microscopy, firstly to collect images based on fluorescence lifetime contrast of GFP variants used as a reporter of gene expression in chondrocytes and secondly, to measure molecular proximity in erythrocyte (glycophorin/membrane) by Fluorescence Resonance Energy Transfer (FLIM-FRET).

Erythrocytes↗

PubMed1988

[A case of colonic polypoid ganglioneuromatosis--a light microscopy, electron microscopy and immunohistochemical study].

A 68-year-old man with colonic polypoid ganglioneuromatosis is reported. He had manifested no signs and symptoms of von Recklinghausen's disease and/or a multiple endocrine neoplasia syndrome. A single polyp was found by a barium enema and an endoscopic polypectomy was performed. Light microscopy showed the propria mucosa of the polyp was composed of a proliferation of spindle cells with a fibrillar cytoplasma and large cells with a round large nucleus, a prominent nucleolus, and vast cytoplasm. Some crypts were slightly dilated. Random biopsied specimens from the colonic normal mucosa showed focal proliferation of only the spindle cells. Electron microscopically, the spindle cells consisted of Schwann cells forming neurites, and large cells that coincided with the features of ganglion cells. Immunohistochemically, the localization of neuron specific enolase was demonstrated in the cytoplasm of these cells. These findings led to a diagnosis of colonic polypoid ganglioneuromatosis, and the pertinent literature is reviewed.

Aged↗

PubMed1987

[Stromal tumor of the testis with Leydig cell components. Light and electron microscopy analysis and fluorescence microscopy demonstration of F-actin].

A testicular gonadal stromal tumor with Leydig cells and mesenchymal cells resembling to Sertoli cells is presented. At the ultrastructural level most of the tumor cells could only be classified as endocrine differentiated cells. Because of their uncharacteristic organelles no Sertoli cells could be demonstrated by means of electron microscopy. The labelling of fibrillary actin underlines the cellular heterogeneity and the different cellular functional state of the tumor cells. These findings are interpreted as an evidence of a multidirectional differentiation potential of the gonadal stroma mesenchyme. These observations support the classification of these neoplasia as gonadal stromal tumors particularly in light of the clearly visible differentiation. Problems of evaluating the malignant potential of gonadal stromal tumors are also discussed.

Actin Cytoskeleton↗

PubMed1986

[Scanning electron microscopy and light microscopy studies on the retina of chick embryos in cell culture].

Histological and scanning electron microscopy studies (SEM) suggest a classification of neural elements into five main types: receptor cells, bipolar cells, horizontal, amacrine, and ganglion cells in dissociated cell cultures of the retina from 7 to 10 days old chick embryos. By use of SEM the development of the receptor cell inner segments was observed. At the same time many protrusions were noticed at the receptor cell surface. Specific synaptic contacts between the axons of the receptor cells and the dendrites of the bipolar neurons, as well as unspecific contacts between the pericarya of the receptor cells and other dendrites were demonstrated. The bipolar neurons showed smooth cell surfaces, however, the horizontal cells appeared rough on the cell surfaces. Müller cells were always found adjacent to the photoreceptor cells.

Animals↗

PubMed1987

Immunogold labeling of human leukocytes for scanning electron microscopy and light microscopy: quantitative aspects of the methodology.

When cell surface antigens are labeled with the colloidal gold marker, back-scattered electron images (BEI) reveal all the gold particles and, therefore, permit total counts. Secondary electron images (SEI) show only a small percentage of the gold particles and are inadequate for quantitative evaluation. For determination of the cellular labeling index, a time-consuming method implies the screening of 100 cells by scanning electron microscopy, at a magnification of approximately 12,000 to 15,000x, with continuous SE/BE shifts. A much more efficient method is to transfer the SEM sample or its equivalent under the light microscope and to count the total number of gold labeled cells in the epi-polarization mode. The total cell count can be evaluated under UV light, taking advantage of the autofluorescence of the glutaraldehyde fixed cells.

Antigens, Surface↗

PubMed1983

Microcirculation of the pancreas: a correlative study of intravital microscopy with scanning electron microscopy of vascular corrosion casts.

The microcirculation of the pancreas in anesthetized rats was examined by intravital microscopy. Scanning electron microscopic observation of vascular corrosion casts of the same parts of the pancreas as viewed light microscopically in vivo, allowed detailed analysis of microcirculation in particular vessels. Blood vessels with flowing blood cells were clearly observed under incident ultraviolet illumination following systemic injection of an intravascular fluorescent tracer. The islets of Langerhans were easily identified by their brighter appearance than the exocrine portions under incident ultraviolet illumination. Blood leaving the islets passed through the insulo-acinar portal vessels into the capillary network around the acini. The periductular plexus received blood from the periacinar capillary network via either venules or capillaries as well as directly from the inter- or intralobular arteries. The periductular plexus also received blood from the islets located close to the ductules. The results obtained show blood in the pancreas flowing in a manner in which it carries insular secretions in high concentrations through short and direct vascular routes from the islets to the pancreatic exocrine tissues, including their duct system.

Animals↗

PubMedDate not supplied

Giant cell carcinoma of the lung: comparative studies of the same cancer cells by light microscopy and scanning electron microscopy.

Surface ultrastructures of giant cell carcinoma cells were studied utilizing a technique developed by the authors which permits observation of the same cells by both light and scanning electron microscopy. Although some general similarity was shown to exist between giant cell carcinoma and squamous cell carcinoma cells, the following distinguishing features of giant cell carcinoma cells were demonstrated: (1) absence of keratinization, (2) full, bulging cellular edges and (3) presence of deep invaginations or small hollows. The features which distinguished giant cell carcinoma from adenocarcinoma cells were: (1) presence of many flattened cells, (2) folded cell surface, (3) absence of tight clusters of cancer cells and (4) few microvilli. Furthermore, the characteristic granularity observed in small cell carcinoma was not observed in giant cell carcinoma cells. Characteristically, cells of giant cell carcinoma are extremely large and flattened, engulf numerous leukocytes and have a bulging cellular edge.

Biopsy, Needle↗

PubMed1982

A comparative study on application of Nd-YAG laser radiation and electrocautery in canine gastric mucosa--magnified observations by dissecting microscopy and scanning electron microscopy, and histological examinations.

The mucosal change, the depth of injury and the healing process in the canine gastric wall after Nd-YAG laser (YAG) exposure and electrocautery were studied comparatively by magnified observations. The results were as follows: 1. Mucosal changes caused by the YAG laser beam were more localized within the exposure area than those by electric current. 2. The depth of tissue damage to the canine gastric wall by YAG laser exposure was easy to control when compared to that caused by electrocautery. 3. The healing time of the ulcer caused by the YAG laser was shorter than that caused by high frequency current. 4. In the present study on the mucosal changes due to laser exposure and electrocautery using endoscopy, dissecting microscopy and SEM, it was found that these examinations appear to be helpful as experimental procedures to clarify the detailed appearance of the mucosal surface. It was suggested that endoscopic application of the YAG laser to the gastrointestinal tract was safe and effective when compared with other procedures which have been applied endoscopically for diagnosis and treatment.

Animals↗