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We determined the Na/K x-ray intensity ratio in frozen sections of myocardial tissues obtained at autopsy from patients who died from various causes, using electron probe analysis. We have been able to distinguish between the ischemically injured and normal cells. The method is simple, fast, and dependable even when the duration of ischemia is only 30 minutes.
We have investigated the effect of orexin A on the intracellular free calcium concentration ([Ca2+]i) in primary cultured cortical neurons and explored the exact mechanisms of orexin A-evoked changes of [Ca2+]i. In the present study, changes of [Ca2+]i induced by orexin A in primary cultured cortical neurons were first detected by confocal laser scanning microscopy using Ca2+-sensitive dye fluo-4 as a novel calcium fluorescent probe. Our results showed that 1-0.1 microM orexin A induced the increase in [Ca2+]i in cortical neurons. The increase in [Ca2+]i by acute application of orexin A occurred in a dose-dependent manner. Orexin A-induced increase in [Ca2+]i was not observed under the condition of Ca2+-free Dulbecco's modified Eagle's medium. Pretreatment on the cells with 1 microM thapsigargin did not block orexin A-evoked response. These findings first illuminated the fact that orexin A-induced increase in [Ca2+]i may be mainly from extracellular calcium influx in cortical neurons.
Hydroxyapatite particles and poly(L-lactide) composites for internal fixation of bone fractures have been developed based on the hypothesis that incorporation of hydroxyapatite particles in a poly(L-lactide) matrix might enhance bone bonding. This study evaluated the bone bonding ability of these biodegradable composites. Two types of hydroxyapatite and poly(L-lactide) composite were used in this study: calcined hydroxyapatite/poly(L-lactide) and uncalcined hydroxyapatite/poly(L-lactide). Rectangular plates (2 x 10 x 15 mm) of each composite or poly(L-lactide) were implanted into the metaphysis of the tibiae of 33 male rabbits, and the failure load was measured by conducting a detaching test 8, 16, and 25 weeks after implantation. The failure loads of calcined hydroxyapatite/poly(L-lactide), uncalcined hydroxyapatite/poly(L-lactide), and poly(L-lactide), respectively, were 13.60, 13.95, and 0.46 N at 8 weeks; 29.84, 24.09, and 2.86 N at 16 weeks; and 25.50, 29.67, and 2.43 N at 25 weeks. Histologic observation revealed that the composites formed direct contact with the bone. The results in this study indicate that the composites improved the strength of the interface between bone and plate. This improved interfacial strength lead to a substantial decrease in the frequency of implant loosening in the treatment of fractured bones by internal fixation.
PURPOSE: The aim of this study was to evaluate the ultrastructure of acute calcification observed in alkali-injured rabbit corneas treated with synthetic inhibitor of metalloproteinases (SIMP). METHODS: A 12-mm alkali burn was made on the right corneas of 16 rabbits. Eight eyes were treated with topical phosphate-buffered SIMP, and eight control eyes were given the buffer only. Six corneas (five in the SIMP group and one in the control group) developed chalky white opacification in the central area of the cornea within 6-30 days. The corneas were excised for the following examinations: (a) light microscopy, (b) energy-dispersive x-ray analysis, (c) electron microscopy, and (d) high-angle x-ray diffraction. RESULTS: The results confirmed that there was stromal calcification in these corneas, as shown by positive van Kossa staining, needle-shaped calcium crystal aggregates, and increased calcium in the opaque areas of the corneas. Proteoglycans in the corneal stroma of both the SIMP group and the control group significantly decreased after alkali burns. Very few proteoglycan filaments could be identified in the calcified areas of the stroma. X-ray diffraction reflections with a Bragg spacing of 3.44 +/- 0.04 A were observed from all the calcified corneas treated with SIMP. The intermolecular spacings of collagen fibrils in the calcified stroma (1.24 +/- 0.34 nm) were significantly lower than those in the noncalcified stroma (1.64 +/- 0.04 nm) and the control corneas (1.64 +/- 0.02 nm). The results indicate that the collagen molecules within the fibrils of the calcified stroma are packed more closely than normal. CONCLUSION: The comparison of the x-ray diffraction patterns from the calcified corneas and chicken bone suggests that the calcium deposits in these corneas are in a form similar to those found in bone. The higher rate and more extensive calcification in the SIMP-treated corneas may result from the inhibitory action of SIMP on debris processing, but the mechanism is unknown.
Three types of activity composition have been found in airborne hot particles that were transported long distances from the Chernobyl accident. Their characterization is based on the analysis of single particles isolated from Pinus Sylvestris needles. The average activity of the particles was 130 Bq at the time of the accident. The most common type of particle contains the radioactive species 141Ce, 144Ce, 95Zr and 95Nb; the second type includes 103Ru and 106Ru along with the previous isotopes; and the third contains 103Ru and 106Ru only. Cesium-134 and -137 were present only in very small amounts. The activity composition of the Chernobyl reactor core fuel was similar to the composition of the first and second type particle; apparently the core fuel was only partially volatilized. The main bulk composition of the particles is shown to be U. The average aerodynamic size of the identified hot particles is 10 microns. The particles are rectangular or pentagonal in shape.
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This investigation studies the effect of the shot-blasting treatment on the cyclic deformation behavior of a commercially pure titanium, with two microstructures: equiaxed and acicular. The fatigue tests were carried out in artificial saliva medium at 37 degrees C. Cyclic deformation tests have been carried out up to fracture, and the fatigue crack nucleation and propagation have been analyzed. The results show that the shot-blasting treatment improves the fatigue life in the microstructures studied, and that the equiaxed was better in mechanical properties than the acicular. The cause of this improvement in the mechanical properties is due to the compressive stress on the material surface for the shot-blasted specimens. Hardness tests were carried out to determine the value of these internal stresses.
The light microscopic and polarization appearances of calcium pyrophosphate dihydrate crystal deposits in tissues are reviewed. In routine sections haematoxylinophilic crystalline deposits with a feathery or brush-like pattern are typical of calcium pyrophosphate dihydrate. Short rhomboidal crystals showing positive birefringence are seen on polarization; X-ray microanalytical and infrared spectroscopic data support the specificity of these appearances. The appearances of the crystal deposits in decalcified specimens are also described. We include six cases of calcium pyrophosphate dihydrate deposition within periarticular bone; to the best of our knowledge this has not previously been described.
A cryopreparation technique for studies of ultrastructure and distribution of diffusible elements in biological tissue is described. Electron microscopical contrast and characteristic X-ray spectra are found to be poor in completely frozen-hydrated ultrathin cryosections of fresh chemically untreated tissue. Both STEM contrast and detection of characteristic X-rays are enhanced by careful freeze-drying in the microscope. Although the ultrastructure is affected by ice crystals, intracellular compartments can be identified by STEM without staining and studied by X-ray microanalysis.
Three dry etching techniques (Ar+ ion beam, O2+ ion beam, O2 radiofrequency electrodeless discharge) were compared with respect to preferential etching and damage to the ultrastructure of glutaraldehyde-fixed Epon-embedded frog skeletal muscle sections. SEM and TEM studies were performed on both unstained and stained (osmium tetroxide, uranyl acetate) sections. Etching effects were observed to differ for the various ion beam or plasma etching techniques. Whereas selective retention of electron dense structures (e.g. Z lines, nuclear heterochromatin) was observed for oxygen plasma etching, preferential etching of these components was observed using O2+ ion beam bombardment. Selectively etched Z lines and etch-resistant nucleoli were observed for both reactive (O2+) and inert (Ar+) ion beam sputtering after sufficiently high ion doses. The above suggest that selective etching under keV ion beam irradiation is related more to physical sputtering processes (momentum transfer) than to the chemical reactivity of the incident ion. Heavy metal post-fixation and staining had no qualitative effect on the nature of the selective etching phenomena. The above findings are significant in that they potentially influence both electron and ion microprobe measurements of etched biological specimens.
Titanium alloys of 10 wt%-72 wt% Cu, 10 wt%-80 wt% Co and 20 wt%-84 wt% Ni were investigated. Ingots were fabricated in a vacuum/argon tungsten arc furnace. The surfaces of the alloys were examined by optical microscopy and SEM/EDS, and the Knoop hardness values of the alloys were measured. The corrosion resistance of the alloys was determined by a potentiodynamic polarization technique in buffered Ringer's solution. When a threshold composition of 30 wt% alloy was reached, a large decrease in corrosion resistance was found to occur. Knoop hardness measurements showed that similar hardness values of approximately 300 KHN can be obtained in all systems with lower alloy content. These values are similar to those obtained with a commercial dental titanium alloy.
The radiopacity of 12 VL-cured composite resins was determined with reference to an aluminum step-wedge. Two anterior composites were radiolucent while two anterior and one anterior/posterior composites exhibited the radiopacity equal to, or slightly greater than, that of human enamel. Three posterior and one inlay composites possessed the radiopacity equivalent to, or in tiny excess of, that of human enamel. Three posterior composites had the radiopacity, fairly exceeding that of human enamel. Chemical analyses of the filler particles were carried out with SEM/EDX. It became evident that radiopaque fillers contained at least one radiopaque oxide component such as BaO, ZrO2 and Yb2O3 with varying concentrations. In general, the radiopacity of the composite resin was linearly proportional to the amount of the radiopaque oxide in the filler. It was suggested that ZrO2 was radiopacifier equivalent to, or even stronger than, BaO.
Histopathological relationship between orcein positive hepatocellular material (ophm) reported by Sipponen and human lipofuscin (hlp) was investigated. Ophm was found to be very similar to hlp of the liver as well as of heart muscle cell morphologically and histochemically, and it seems to be quite rational to consider that ophm may be a substance which belongs to the lysosome series comprising hlp and prelipofuscin. Ophm has been discussed often from a view point of copper contents, which was considered to be one of its specific characters, but it has become clear that hlp can also possess copper according to our investigation and to some reports of other researchers. This fact throws a doubt on the significance of the specific relation between ophm and copper, as well as the usability of orcein stain for detecting copper in histological specimens. Victoria blue stain is quite useful for the detection of hlp including ophm, since it gives a blue color for it with a good contrast against red colored cytoplasm and nucleus. However, this can not be considered as a method for specific stain of copper either.
X-ray microprobe analysis was used to determine concentrations (millimoles of element per kilogram dry weight) of Na, P, Cl, K, and Ca in cellular compartments of frozen, unfixed sections of rat sciatic and tibial nerves and dorsal root ganglion (DRG). Five compartments were examined in peripheral nerve (axoplasm, mitochondria, myelin, extraaxonal space, and Schwann cell cytoplasm), and four were analyzed in DRG nerve cell bodies (cytoplasm, mitochondria, nucleus, and nucleolus). Each morphological compartment exhibited characteristic concentrations of elements. The extraaxonal space contained high concentrations of Na, Cl, and Ca, whereas intraaxonal compartments exhibited lower concentrations of these elements but relatively high K contents. Nerve axoplasm and axonal mitochondria had similar elemental profiles, and both compartments displayed proximodistal gradients of decreasing levels of K, Cl, and, to some extent, Na. Myelin had a selectively high P concentration with low levels of other elements. The elemental concentrations of Schwann cell cytoplasm and DRG were similar, but both were different from that of axoplasm, in that K and Cl were markedly lower whereas P was higher. DRG cell nuclei contained substantially higher K levels than cytoplasm. The subcellular distribution of elements was clearly shown by color-coded images generated by computer-directed digital x-ray imaging. The results of this study demonstrate characteristic elemental distributions for each anatomical compartment, which doubtless reflect nerve cell structure and function.
Using analytical electron microscopy, we directly tested the hypothesis that transition metals and sulfur are the cause of chlorhexidine-induced tooth stain. Plaque scraped from the teeth of water-treated individuals or chlorhexidine-treated "non-stainers" was non- or lightly-stained and contained low concentrations of sulfur and transition metals. Tooth scrapings from heavily-staining chlorhexidine-treated individuals consisted of distinct unstained and stained regions. The stained regions were organic but were in close proximity to mineralized areas. Enhanced levels of sulfur and transition metals, particularly iron, were found in stained regions, whereas unstained regions contained low sulfur and metal levels similar to the water-treated or non-staining individuals. Excluding decreased mineralization, the major elemental change in heavily-stained plaque was an increase in sulfur, and to a lesser extent, iron and other transition metals. Sulphur and iron levels were directly correlated. Following chlorhexidine treatment with iron supplementation, staining was enhanced, the Fe/S ratio increased, and sulfur and iron remained correlated. These data support the hypothesis that transition metals and sulfur are the cause of chlorhexidine-induced tooth stain. The data are consistent with chlorhexidine treatment altering the incorporation into plaque of a natural sulfur-containing organic component of saliva or bacteria. This natural component appears to readily interact with transition metals, particularly iron, producing stained material.
The analysis of particles in tissue samples, especially in lung tissue samples, has been of great use diagnostically and in the search for more precise information on the etiology of occupational/environmental diseases. There is a strong need for an interdisciplinary approach involving patients, epidemiologists, clinicians, pathologists, toxicologists, and microscopists if advancement in the recognition, diagnosis, and prevention of disease is to proceed.
The AEM is a powerful tool in biological research, capable of providing information simply not available by other means. The use of a field emission STEM for this application can lead to a significant improvement in spatial resolution in most cases now allowed by the quality of the specimen preparation but perhaps ultimately limited by the effects of radiation damage. Increased elemental sensitivity is at least possible in selected cases with electron energy-loss spectrometry, but fundamental aspects of ELS will probably confine its role to that of a limited complement to EDS. The considerable margin for improvement in sensitivity of the basic analytical technique means that the search for technological improvement will continue. Fortunately, however, current technology can also continue to answer important biological questions.
Biominerals are widely exploited to harden or stiffen tissues in living organisms, with calcium-, silicon-, and iron-based minerals being most common. In notable contrast, the jaws of the marine bloodworm Glycera dibranchiata contain the copper-based biomineral atacamite [Cu2(OH)3Cl]. Polycrystalline fibers are oriented with the outer contour of the jaw. Using nanoindentation, we show that the mineral has a structural role and enhances hardness and stiffness. Despite the low degree of mineralization, bloodworm jaws exhibit an extraordinary resistance to abrasion, significantly exceeding that of vertebrate dentin and approaching that of tooth enamel.