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PubMed1988

Subcellular electrolyte shifts during in vitro myocardial ischemia and reperfusion.

Isolated perfused rabbit right ventricular wall was studied with electron probe microanalysis (EPMA) under three conditions: 1) control (37 degrees C, 1.2 Hz), 2) 60 min global ischemia, and 3) ischemia plus 5 min of reperfusion. After 60 min of ischemia, only one cell population was evident; the variance of intracellular electrolyte concentrations was the same as in controls. When compared with controls, there was no change in Ca concentration within any region of the cell, but mitochondria were swollen with K-rich fluid. Two cell populations were evident after 5 min of reperfusion. The severely injured cells were markedly swollen, exhibited hypercontraction bands, and had electrolyte profiles similar to extracellular fluid. The moderately injured cells were normal in appearance, still retained electrolyte gradients, but had elevated Na and Cl concentrations in all compartments. Cell Ca did not increase in the moderately injured cells, but the region of the cell containing the sarcoplasmic reticulum (SR) lost 90% of its Ca. Accompanying this loss were large increases in myofibrillar and mitochondrial Ca concentration. It appears that release of SR Ca, loss of SR Ca-accumulating capacity, and increased intracellular Na are the principal electrolyte shifts in functional cells during early reperfusion.

Animals↗

PubMed2000

In vitro effects of zirconia and alumina particles on human blood monocyte-derived macrophages: X-ray microanalysis and flow cytometric studies.

The cytocompatibility of two particulate bioceramics, zirconia and alumina, was studied using human blood monocytes driven to differentiate into mature macrophages with granulocyte macrophage-colony-stimulating factor. Changes in individual cell elemental composition, particularly sodium and potassium content, were assessed by X-ray microanalysis of ultrathin freeze-dried sections. Phagocytosis and respiratory burst of macrophages exposed to biomaterial for 7 days were analyzed under flow cytometry using uptake of fluorescent latex beads and 2'7'-dichlorofluorescien diacetate oxidation, respectively. Zirconia and alumina particles were found to decrease the intracellular potassium/sodium ratio (an index of cell vitality) significantly (p<.01) in 7-day-cultured macrophages compared to control cells cultured out of material. Phagocytosis of both ceramic particles by macrophages was followed by a concomitant decrease in cell phagocytic ability (27%) and a marked altered oxidative metabolism (>2 times reduced by zirconia and >5 times reduced by alumina). The present study clearly demonstrates that reduction of the phagocytic capacity of macrophages associated with altered oxidative metabolism caused by biomaterial particles is characterized by changes in intracellular elemental content. Thus, investigation of cellular homeostasis by electron probe microanalysis together with analysis of functional changes may improve estimation of biomaterial cytocompatibility.

Aluminum Oxide↗

PubMed1987

Effect of time at 37 degrees C on the creep and metallurgical characteristics of amalgam.

When aged at 37 degrees C for six months, dental amalgam exhibits a marked decrease in the property of creep. The objective of this study was to investigate the relationship between this decrease in creep and selected metallurgical characteristics. The formation of beta 1 (Ag-Hg), the grain size of gamma 1 (Ag-Hg), and the composition of gamma 1 were chosen for investigation. Creep was determined according to ADA Specification test No. 1, beta 1 was measured by x-ray diffraction, and gamma 1 grain size and gamma 1 composition were determined by electron-probe microanalysis. The results showed that the decrease in creep was related to beta 1 formation and not to changes in either gamma 1 grain size or gamma 1 composition.

Chemical Phenomena↗

PubMed1992

[Effect of morphological properties on drug release from biodegradable microspheres].

The morphological properties of poly(beta-hydroxybutyric acid) (PHB) or poly(L-lactic acid) microspheres loading flomoxef sodium (FMOX) were investigated with regard to FMOX release. The release profiles of FMOX from the microspheres could be divided into two types, a sustained release type and a burst one. Two representative PHB microspheres, the release profiles of which were quite different from those of FMOX, were compared in detail from a morphological point of view. The shapes of their surfaces and sections were observed by using scanning electron microscopy (SEM), and FMOX distribution was analyzed by using electron probe microanalysis. The crystallinity of polymers was further measured by powder X-ray diffratometry. There was little difference in the FMOX distribution and their microscopic properties such as sphere size, specific surface area, shape of surface and section. In contrast, water penetration into the inside of the microspheres was found to be clearly different by use of cryogenic SEM. A significant difference was also observed in the crystallinity of polymers forming the microspheres. The release of FMOX from the microspheres was affected by the crystallinity of polymers forming the microspheres, and burst phenomena occurred in case the polymer was highly crystallized. It was speculated that the crystallization of polymer induced micro voids in the microspheres which functioned as channels for water penetration.

Biodegradation, Environmental↗

PubMed1982

Changes in Schwann cells and vessels in lead neuropathy.

Transmission electron microscopy (TEM) of peripheral nerve in rats receiving 6% lead carbonate for 4-10 weeks provided evidence of a specific Schwann cell injury, associated with demyelination. Intranuclear inclusions in Schwann cells appeared within 2 weeks of administration of a lead-containing diet. Swelling of Schwann cells and disintegration of their cytoplasm was evident at 4 weeks. Distinctive electron-dense inclusions appeared in both Schwann and endothelial cells during the period of intoxication and were ultrastructurally identical to pathognomonic inclusions of lead poisoning seen in renal tubular epithelial cells. Scanning microscopy (SEM) with electron-probe microanalysis was used to identify the lead-containing deposits. In addition to Schwann cell changes, vessels revealed endothelial cell injury and alteread permeability to macromolecules. Since morphologic changes of Schwann cells precede the development of altered vascular permeability and endoneurial edema, it appears that lead gains access to the endoneurium prior to the development of altered vascular permeability, suggesting that edema and altered endoneurial fluid pressure are epiphenomena that supervene after demyelination occurs. Remyelination, Schwann cell proliferation and formation of onion bulbs are manifestations of persistent toxic injury to myelin-sustaining cells, resulting in chronic demyelination.

Animals↗

PubMed1983

The distribution of intracellular ions in the avian salt gland.

To investigate the mechanism of salt secretion in the avian salt gland, we used quantitative electron probe microanalysis to measure the intracellular elemental concentrations in dry cryosections of unspecialized and partially specialized secretory epithelial cells from fresh water- and salt water-adapted ducklings, respectively. In conjunction with this, human and duckling erythrocytes were also analyzed, since these provided the experimental basis for using in situ erythrocytes as standards for determining the local water content of epithelia from the analysis of dried cryosections. The microprobe results from both types of erythrocytes compared favorably with chemical determinations of elemental concentrations. The nucleated avian erythrocytes, whose wet-weight elemental concentrations were determined by a compartmental analysis that required neither a peripheral standard nor a measure of the local mass, revealed a marked accumulation of P and K in the nucleus (388 and 190 mmol/kg wet wt, respectively) relative to the cytoplasm (67 and 85 mmol/kg wet wt). In both developmental states of the epithelial cells, the nucleus and apical cytoplasm had essentially similar and unremarkable concentrations of Na (76 and 83 mmol/kg dry wt, respectively, in the adapted cells vs. 72 and 81 mmol/kg dry wt in the control cells) and K (602 and 423 mmol/kg dry wt vs. 451 and 442 mmol/kg dry wt). Chloride, however, which was in general rather high, was significantly depressed in the apical cytoplasm of adapted cells only (164 and 124 mmol/kg dry wt in the nucleus and cytoplasm, respectively, of adapted cells (P less than 0.05) vs. 138 and 157 mmol/kg dry wt for control cells (P less than 0.05). Cation concentrations (Na + K) were elevated approximately 15% in the basal regions of adapted cells as compared with apical cytoplasm. When tissue water variations are accounted for, the results suggest that: (a) an active, energy-requiring process is responsible for chloride accumulation in this cell; (b) the apical membrane is a regulatory site for secretion; and (c) there are regional distinctions in the distribution of ions and water, particularly in the salt water-adapted cell. These conclusions are consistent with active chloride transport as the basis for salt secretion in this tissue.

Animals↗

PubMed1978

Cytochemistry and x-ray microprobe analysis of the midgut of Tomocerus minor lubbock (insecta, collembola) with special reference to the physiological significance of the mineral concretions.

Histochemical and cytochemical analyses have been made on the mineral concretions within the midgut cells of Tomocerus minor. The classical histochemical methods are not specific and precise enough and have been supplemented with cytochemical techniques on ultrathin sections. The most interesting of these was the K-pyroantimonate technique combined with glutaraldehyde-osmium fixation. This technique shows the distribution of cations such as Ca++, K+, Mg++ and Na+ on the concentric layers of the concretions. Chloride ions can be detected by means of the silver lactate technique. The action of calcium chelators such as E.D.T.A. shows an important distribution of calcium ions in the concretions. The spectra obtained by electron probe microanalysis from areas of fresh, dried and carbon coated midguts as well as from carbon coated semithin or ultrathin sections reveal the presence of Ca, K, Mg, S, Cl and P principally. Other elements such as aluminium, silicon and manganese have also been detected. Iron is not always present. The chemical and X-ray analytical investigations indicate that the midgut concretions are mainly built up of calcium, potassium, magnesium and sodium phosphates, perhaps associated with chlorides and carbonates. An organic matrix formed by polysaccharides seems to join the different mineral layers. These concretions may be formed within the vesicles of rough endoplasmic reticulum. The midgut cells are highly differentiated and very active in transport. Extensive basal infoldings and apical microvilli as well as lateral membranes are a site of small cationic deposits. The possible pathway of ion transport in the cell and the physiological significance of the concretions are discussed. The principal function of these concretions seems to be the maintenance of the mineral balance and to trap foreign and excess ions.

Aluminum↗

PubMed1996

Bonding of a mica-based castable ceramic material with a tri-n-butylborane-initiated adhesive resin.

Adhesive bonding of a mica-based castable ceramic material (Olympus Castable Ceramics, OCC) was evaluated in vitro with the use of a silane primer in conjunction with an adhesive luting material. The primer contained a silane coupler and 4-methacryloxyethyl trimellitate anhydride (4-META), while the methyl methacrylate (MMA)-based luting agent was initiated with a tri-n-butylborane derivative (TBB) and contained 4-META (4-META/MMA-TBB resin). Ceramic specimens were sanded with No. 600 silicon carbide paper followed by blasting with alumina and/or etching with ammonium bifluoride. The specimens were bonded with various combinations and shear bond strengths were determined. Both priming and alumina blasting enhanced the bond between 4-META resin and OCC. Although etching with ammonium bifluoride roughened the ceramic surface, this procedure did not improve the bond strength. Electron probe microanalysis of the ceramic surface revealed a decrease in silicon and aluminium elements after etching with ammonium bifluoride.

Acid Etching, Dental↗

PubMed1983

Synthesis of polypeptide models of elastin. Synthesis and properties of a cross-linked polytetrapeptide.

Synthesis of two copolymers, H-(phi-Pro-Gly-Gly)n-Val-OMe and H-(phi'-Pro-Gly-Gly)n-Val-OMe, where phi is Val or Lys and phi' is Val or Glu is described. Cross-linking between the two copolymers is achieved by a coupling reaction between the epsilon-amino groups of the lysine containing copolymer and delta-carboxyls of the glutamic acid of the other copolymer. The cross-linking reaction was performed during a temperature elicited phase separation with flow orientation of the copolymers. Coacervation of the intermediate polymers is presented, as is the scanning electron micrograph of the insoluble cross-linked product and its calcifiability as determined with electron probe microanalysis. The purity of the key intermediates and polymers is demonstrated by the usual analytical methods. Carbon-13 magnetic resonance spectra of the intermediate monomer units are included to validate the purity of their synthesis.

Cross-Linking Reagents↗

PubMed2000

A biodegradable hybrid sponge nested with collagen microsponges.

A biodegradable hybrid sponge of poly(DL-lactic-co-glycolic acid) (PLGA) and collagen was fabricated by forming microsponges of collagen in the pores of PLGA sponge. Observation of the PLGA-collagen hybrid sponge by scanning electron microscopy (SEM) showed that microsponges of collagen with interconnected pore structures were formed in the pores of PLGA sponge. The hybrid structure further was confirmed by scanning electron microscopy-electron probe microanalysis (SEM-EPMA), and elemental nitrogen was detected in the microsponges of collagen and on the pore surfaces of PLGA, but not in cross-sections of PLGA regions. The formation of collagen microsponges was dependent on collagen concentration, the effective range of which was from 0.1 to 1.5 (w/v) %. The mechanical strength of the hybrid sponge was higher than that of either PLGA or collagen sponges, in both dry and wet states. The wettability with water was improved by hybridization with collagen, which facilitated cell seeding in the hybrid sponge. Mouse fibroblast L929 cells attached well and spread on the surfaces of the microsponges of collagen in the hybrid sponge. The distribution of cells was spatially uniform throughout the hybrid sponge. Use of the PLGA sponge as a skeleton facilitated formation of the hybrid sponge into desired shapes with high mechanical strength while collagen microsponges contributed good cell interaction and hydrophilicity.

Animals↗

PubMed1993

Elemental composition of secretory granules in pancreatic islets of Langerhans.

We have characterized, by electron probe microanalysis, rapidly frozen cultured rat islets at the level of individual secretory granules. Elemental analysis of thin, dried cryosections showed that beta granules could be distinguished by high Zn, Ca, and S, whereas non-beta (mainly alpha) granules contained elevated P and Mg. Although a single granule type predominated in a particular cell, some rebel granules were found in A cells that had the compositional fingerprint of B cell granules. Zn, which was found in millimolar concentrations in B cell granules, was considered a marker for the insulin storage complex. The data indicate that non-B islet cells in the adult pancreas may produce insulin-containing organelles and that, when glucagon and insulin are coexpressed, these hormones are packaged in separate granules.

Animals↗

PubMed1987

Abnormal hepatic copper storage in a teleost fish (Morone americana).

Excessive copper storage in livers of feral white perch (Morone americana) from the Chesapeake Bay is described. Age-related, progressive accumulation of hepatic copper in levels often exceeding 1,000 micrograms/g wet weight was associated with peribiliary fibrosis and inflammation, bile duct hyperplasia, prominent, enlarged melanomacrophage centers, and disruption of hepatic architecture in older fish. Levels of zinc were mildly elevated compared to striped bass (Morone saxitilis) and adult rats. Cholangiomas were found in two perch. Rubeanic acid-stained liver had abundant copper-positive cytoplasmic granules in hepatocytes and cells of melanomacrophage centers. Subcellular fractionation showed that 90% of hepatocellular copper was in nuclei/cell debris fractions (which also contain tertiary lysosomes). Using electron probe microanalysis, high copper levels were localized in hepatocellular cytoplasmic bodies. Resolution of hepatic cytosol by gel permeation chromatography indicated that approximately 50% of the cytosolic copper in the white perch was bound to non-specific high molecular weight proteins, with the remaining 50% eluting at a peak where rat metallothionein is located. Ultrastructural examination revealed abundant lysosomes, increased size and number of peroxisomes, and increased density and numbers of mitochondrial matrix granules. This study indicates that white perch may be a model for studying effects of excessive copper accumulation and cellular mechanisms which control copper kinetics.

Animals↗

PubMed1999

The electron microscope appearance of the subchondral bone plate in the human femoral head in osteoarthritis and osteoporosis.

The subchondral bone plate supports the articular cartilage in diarthrodial joints. It has a significant mechanical function in transmitting loads from the cartilage into the underlying cancellous bone and has been implicated in the destruction of cartilage in osteoarthritis (OA) and its sparing in osteoporosis (OP), but little is known of its composition, structure or material properties. This study investigated the microscopic appearance and mineral composition of the subchondral bone plate in femoral heads from patients with OA or OP to determine how these correspond to changes in composition and stiffness found in other studies. Freeze-fractured full-depth samples of the subchondral bone plate from the femoral heads of patients with osteoarthritis, osteoporosis or a matched control group were examined using back scattered and secondary emission scanning electron microscopy. Other samples were embedded and polished and examined using back-scattered electron microscopy and electron probe microanalysis. The appearances of the samples from the normal and osteoporotic patients were very similar, with the subchondral bone plate overlayed by a layer of calcified cartilage. Osteoporotic samples presented a more uniform fracture surface and the relative thicknesses of the layers appeared to be different. In contrast, the OA bone plate appeared to be porous and have a much more textured surface. There were occasional sites of microtrabecular bone formation between the trabeculae of the underlying cancellous bone, which were not seen in the other groups, and more numerous osteoclast resorption pits. The calcified cartilage layer was almost absent and the bone plate was apparently thickened. The appearance of the osteoarthritic subchondral bone plate was, therefore, considerably different from both the normal and the osteoporotic, strongly indicative of abnormal cellular activity.

Aged↗

PubMed1983

Diffuse pulmonary injury associated with gold treatment.

A patient showing a gold-induced diffuse pulmonary reaction is described. An open lung biopsy specimen showed thickening of alveolar septa due to inflammation. The alveolar walls were lined by cuboidal epithelium. On electron microscope examination electron-dense structures were observed within lysosomes of endothelial cells of the alveolar capillaries and interstitial macrophages. Electron probe microanalysis showed that these structures contained gold. The accumulation of gold in endothelial cells and macrophages and a direct toxic effect might be an important feature in the pathogenesis of gold-induced pulmonary reactions. On the other hand the accumulation of gold may reflect exposure rather than disease.

Adult↗

PubMed1980

Microhardness of molar teeth in cattle with fluorosis.

Cattle were fed forage containing fluoride at a yearly average of 40 mg of fluoride/kg of forage for 5 or 6 years from the time they were 4 months of age. A significant (P < 0.05) negative correlation was observed between the average microhardness of the 3rd premolar and 3rd molar enamel and the fluorosis score of the 2nd incisor. The microhardness of fluorotic outer molar enamel was only 41% of the microhardness values of enamel from control teeth not exposed to long-term fluoride, and electron probe microanalysis indicated increased fluoride concentrations in the molar coronal cementum, enamel, and dentin. Increased incisor fluorosis scores were diagnostic of softer molar enamel. When severe, such changes may have a detrimental effect on the proper mastication and subsequent nutrition of cattle with fluorosis.

Animals↗

PubMed1975

Subcellular localization of calcium in the mouse hypophysis. I. Calcium distribution in the adeno- and neurohypophysis under normal conditions.

Application of the K-pyroantimonate technique combined with glutaraldehydeosmium fixation results in a reproducible intracellular distribution of mineral precipitates in the mouse hypophysis. Control experiments--with chelators and electron probe microanalysis--reveal that these precipitates consist mainly of calcium. Regularly present in the mitochondria, Ca also seems to be stored in the Golgi apparatus of the glandular cells and in the axoplasmic reticulum and the "synaptic" vesicles of the neurosecretory fibres. These structures thus appear able to control intracytoplasmic calcium movements. These observations agree with physiological data showing the existence of an intracellular Ca pool that can be mobilized by specific stimulation. The presence of diffuse precipitates in the pituicytes, together with the existence of gap junctions between them, suggest that these cells regulate the ionic environment of the neurosecretory nerve fibres; in this way, they too might participate in neurohypophysial hormonal release.

Animals↗

PubMed1997

Cellular hypertrophy and calcification of embryonal carcinoma-derived chondrogenic cell line ATDC5 in vitro.

During the process of endochondral bone formation, proliferating chondrocytes give rise to hypertrophic cells, which then deposit a mineralized matrix to form calcified cartilage prior to replacement by bone. Previously, we reported that a clonal cell line, ATDC5, undergoes efficient chondrogenic differentiation through a cellular condensation stage. Here we report that the differentiated ATDC5 cells became hypertrophic at the center of cartilage nodules, when the cells ceased to grow. Formation of hypertrophic chondrocytes took place in association with type X collagen gene expression and a dramatic elevation of alkaline phosphate (ALPase) activity. After 5 weeks of culture, mineralization of the culture could be discerned as Alizarin red-positive spots, which spread throughout the nodules even in the absence of beta-glycerophosphate. Electron microscopy and electron probe microanalysis revealed that calcification was first initiated at matrix vesicles in the territorial matrix and that it advanced progressively along the collagen fibers in a manner similar to that which occurs in vivo. The infrared spectrum of the mineralized nodules indicated two absorption doublets around 1030 cm-1 and 600 cm-1, which are characteristic of apatitic mineral. Calcifying cultures of ATDC5 cells retained responsiveness to parathyroid hormone (PTH): PTH markedly inhibited elevation of ALPase activity and calcification in the culture in a dose-dependent manner. Thus, we demonstrated that ATDC5 cells keep track of the multistep differentiation process encompassing the stages from mesenchymal condensation to calcification in vitro. ATDC5 cells provide an excellent model to study the molecular mechanism underlying regulation of cartilage differentiation during endochondral bone formation.

Alkaline Phosphatase↗

PubMed1992

Enamel fluoride uptake from an experimental fluoride-releasing orthodontic adhesive.

The purpose of this study was to evaluate quantitatively the fluoride uptake by enamel from an experimental visible light-cured orthodontic adhesive (VP-862) based on YbF3 filler. Sixteen contralateral premolars were extracted from 10 orthodontic patients and were classified in four groups (A, B, C, D) of four buccal surfaces each. Standardized enamel areas located on these surfaces were acid etched and were subjected to the following adhesive treatments: (A) VP-862; (B) Heliosit Orthodontic; (C) Heliobond + VP 862; and (D) Heliobond + Heliosit Orthodontic. Groups B and D were used as a reference. After 9 months in vivo, the teeth were extracted and cross-sectioned, and the enamel-adhesive interfaces were studied by combined wavelength-energy dispersive electron probe microanalysis. According to the results, the cumulative fluoride uptake by enamel from the experimental adhesive was not statistically different from the fluoride detected in the reference groups. No effect of the liquid resin, Heliobond, on the fluoride uptake gradients of enamel could be differentiated.

Acrylates↗