Scanning electron microscopy最新文献

Although X-ray microanalysis represents a useful tool for identifying electron dense histochemical end-products, quantitative microanalytic measurements are seriously hampered in the case of the activities of certain membrane-bound enzymes. For example, the electron histochemical methods revealing K+-dependent pNPPase activity result in a very fine, granular reaction product of lead phosphate. Therefore microanalytic, densitometric of similar evaluations of the reaction, even in semiquantitative terms are not practical by the usual procedures. This paper describes a method of X-ray microanalysis of thick sections (0.5 micron) processed for K+-pNPPase, where a sufficient amount of lead is present for X-ray microanalytic determination. The analysis is performed in the line scan mode on transversely cut membrane profiles by means of the line scan ratemeter of an EDAX System F. This yields quantitative data on the relative lead concentrations in the vicinity of the cell membrane. A method is proposed for calculation of relative enzyme activities based on the Pb-signal of the ratemeter curve and the average "noise"-level of the cytoplasm, containing also non-specifically bound lead. This method avoids the necessity of measuring the section thickness; it may be useful for a variety of purposes in the electron microscopic histochemistry of membrane-bound enzymes.

This paper presents a summary of the morphological categorization of cell death, results of two in vivo studies on the cell death induced by mild hyperthermia in rat small intestine and mouse mastocytoma, and a comparison of the cell death induced by hyperthermia, radiation and cytotoxic drugs. Two distinct forms of cell death, apoptosis and necrosis, can be recognized on morphologic grounds. Apoptosis appears to be a process of active cellular self-destruction to which a biologically meaningful role can usually be attributed, whereas necrosis is a passive degenerative phenomenon that results from irreversible cellular injury. Light and transmission electron microscopic studies showed that lower body hyperthermia (43 degrees C for 30 min) induced only apoptosis of intestinal epithelial cells, and of lymphocytes, plasma cells, and eosinophils. In the mastocytoma, hyperthermia (43 degrees C for 15 min) produced widespread tumor necrosis and also enhanced apoptosis of tumor cells. Ionizing radiation and cytotoxic drugs are also known to induce apoptosis in a variety of tissues. It is attractive to speculate that DNA damage by each agent is the common event which triggers the same process of active cellular self-destruction that characteristically effects selective cell deletion in normal tissue homeostasis.

Cryoultramicrotomy, which avoids the use of harsh fixation procedures, deleterious dehydration and plastic embedding can be combined with immunocytochemistry to determine the ultrastructural localization of cellular proteins. Our attempts to use the cryosectioning technique in combination with immunolabelling to bridge the gap between light and electron microscopic analysis of muscle morphology have enabled us to obtain new information on fibre typing at the ultrastructural level. Furthermore, we have obtained a marked improvement in the resolution of myofibrillar structures by using semithin cryosections for fluorescence microscopy. Data are also presented on correlated light and electron microscope immunocytochemistry of myocardial intermediate filaments confirming the presence of longitudinally oriented intermediate filaments of desmin in the region of the intercalated discs of mammalian cardiac myocytes, whereas elsewhere in the myocyte the bulk of intermediate filaments of desmin is concentrated in the intermyofibrillar space at the level of the Z disc.

The glomerular complexity of several species of birds and reptiles is investigated in this study by scanning electron microscopy of vascular corrosion casts. Comparing these results with those of a freshwater teleost and a mammalian species, a trend towards small, simple glomeruli of the avian type, beginning with large, well vascularized glomeruli resembling the type found in fish, can be observed in reptiles. A close correlation between glomerular size and habitat can be established comparing related species having a similar physiological mode of renal function. Entirely different from this sauropsidian evolutionary line of development is the highly complex, large differentiation of the mammalian glomerulus.

The ultrastructure of bone marrow of rats was studied 24 h after exchange-transfusion with solutions of starch or modified hemoglobin to a hematocrit of 10-15. Blood smears of the transfused rats had 17-20% reticulocytes as compared to 5-6% for sham operated controls. In the transfused rats marrow macrophages had numerous heterolysosomes apparently containing the starch or hemoglobin from the transfused solutions. Endothelial cells and reticular cells also possessed a few heterolysosomes thought to contain starch or hemoglobin. Reticular cells of the transfused rats contained numerous glycogen particles scattered throughout the cytoplasm or arranged in large masses. Synthesis of glycogen may indicate a metabolic change in reticular cells in response to tissue hypoxia induced by the exchange-transfusion procedure.

The ciliated epithelium of the rabbit trachea was irradiated with daily fractions of 2 Gy to an accumulated dose of 20 Gy (TD: 2, 6, 10, 16, or 20 Gy). Fifteen to forty-five minutes before start of the first irradiation (treatment day 1), 5 mg cis-DDP was given by intraperitoneal injection to each rabbit. Examination was made 1 - 10 days after each fractionation schedule, when specimens were taken for investigations. Scanning electron microscope investigations showed a gradual development of ciliary damage from blebs on the cilia to swollen tips, broken and bent cilia and finally an epithelial lining with areas free from cilia with a surface covered with microvilli-like structures. SEM also showed cell loss, and remnants of dead cells on the surface together with detritus. By transmission electron microscope ciliary damage, cell death and cell loss of the ciliated cell layer as well as exfoliation of portions of goblet-like cells on the surface could be confirmed. The irradiated ciliated epithelium and the untreated control epithelium in each animal showed no difference in this respect. Thus no enhancement of the effects of radiation could be observed. The development of ultrastructural damage may be due to a cytotoxic effect of the drug on the ciliated epithelium. However, 19 days after the start of cis - DDP injection, a hyperplasia of the basal cell layer was observed, which indicates that the observed cytotoxicity of the drug is reversible and a normalisation occurs during the last days of observation in this study.

Aging changes of aortic valves are thought to underlie the mechanism of calcification, which leads to calcific aortic stenosis in humans. The study of calcification in the aging valvular connective tissue has been hindered by the lack of a suitable animal model. In search of the model, canine aortic valves demonstrated age changes including calcification remarkably similar to those in humans. The mechanism of calcification was studied in the aortic valves of aged Beagles by electron microscopy. Fibroblasts in the canine aortic valves showed the most prominent age changes. The cells accumulated numerous residual bodies and appeared to disintegrate. The resultant membranous cellular degradation products which sequestered in the extracellular space were the nidi of calcification. It appeared that the membrane of cell debris played an important role in calcification. Canine aortic valve is an ideal model for the study of calcification in relation to aging of the valvular connective tissue.

The free surface of the endodermal epithelial lining of the rat visceral yolk sac was examined by scanning electron microscopy at three stages of pregnancy, viz., 12, 17 and 22 days (birth typically occurring on the 23rd day). Additionally, by ultrasonic vibration of tissues that had been subjected to prolonged osmium fixation, the epithelium was removed and such microdissected membranes similarly were examined. With increasing gestational age the free surface of the epithelium underwent a relative increase in absorptive area by three mechanisms: formation of increasingly complex villous projections of the visceral yolk sac as a whole, a doming of the individual epithelial cells, and an increase in length and number of microvilli for each such cell. The resultant increase in surface area, however, could not be correlated with the onset of receptor mediated endocytosis at the latest stage. The basement membrane underlying the lining epithelium of the yolk sac easily was revealed by ultrasonication of well-fixed tissues; it was most fragile at 12 days at which time the procedure apparently either removed the lamina lucida of the basal lamina, exposing a fibrillar component of the lamina densa, or removed the entire basal laminar component of the basement membrane, exposing an underlying reticular lamina. At 17 days the basement membrane at the cores of denuded villi showed the negative impressions of the epithelial cells that had been removed; and at 22 days, when the membrane is known to be permeable to protein transfer, it was smooth, featureless, and appeared most durable.