Search

Image analysis quantitation of cytochrome oxidase activity, cytochemicaUy visualized in mitochondria, and the comparison between the activities of pulmonary alveolar Type I and Type II cells were performed. Normal lungs of rats were fixed with 2% glutaraldehyde for 10 min at 4°C and then small blocks were incubated for 60 min at 37°C in a medium containing 1.0 mg/ml (2.8×l0M) 3,3′-diamiDobcnzidlnc-tctrahydrochloride, 1 mg/ml cytochrome c, 0.1 mg/ml catalase, 7% sucrose and 0.1 M phosphate buffer at pH 7.4. The intermembrane space, between the outer and inner membranes, and intracristal space of Type II cell mitochondria were filled with deposits whose mean volume was 0.047 μm. Type I cell mitochondria were smaller in size than those of Type II cells, but because reaction deposits were not fully accumulated within the spaces the resulting net deposit volume was 0.00182 μm If the intermembrane spaces of Type I cell mitochondria had been thoroughly filled by the deposits, the mean volume would have been 0.00248 μm. These results indicate that stereoscopic image analysis of the reaction product is appropriate in the quantitation of cytochemicaUy visualized enzyme activity, and furthermore suggest that Type I cells may be rather deficient in cytochrome oxidase activity. [ABSTRACT FROM PUBLISHER]

Ultrastructural and cytochemical changes in rat livers after paraquat administration were Investigated. A single intraperitoneal injection of 150 mg/ kg paraquat dichioride caused primary toxicological changes in the hepatocyte mitochondria., such as decrease in the electron density of the matrices, disintegration of crlstae, disappearance of intramitocliondrial granules, and irreversible swelling. Ca-ATPase activity in those mitochondria disappeared. experiments, the activity of fixed normal livers was significantly inhibited by 10 mM paraquat. No significant effect on the plasma membrane Ca-ATPase activity was observed. These results suggest that the mitochondrial damage caused by paraquat may be related to the possible inhibition of their Ca-ATPase activity, resulting in cell injury. [ABSTRACT FROM PUBLISHER]

A high incidence of cytoplasmic pore complex similar to Maul's (, 104: 233, 1977) was found in the cultured myeloblastic leukemic cells, Ml line, clone 264-2. The pore complex was composed of one to four pores perforating a single cisterna of the endoplasmic reticulum. When the blastic M1 cells were cultured in the presence of a conditioned medium, they successfully differentiated into macrophages. This resulted in a loss of malignancy, and simultaneously the pore complex changed to the stacked annulate lamellae. The well-developed annulate lamellae were also generally found in the cytoplasm of the cloned macrophages. Mm-1 subline was derived from the M1 line by spontaneous differentiation, and the cells appeared less transplantable to animals. These results indicate that myeloblastic M1 cells possess the simple pore complex, which may develop into the annulate lamellae during cell differentiation into macrophages. [ABSTRACT FROM PUBLISHER]

Although the intracellular toxic sites of paraquat, a herbicide toxic to human bodies, have remained unclear for a long time, we recently demonstrated paraquat-induced mitochondrial injury in rat lung and liver . In the present study, cultured human lung cells (A549 adenocarcinoma cell line) which received 0.15 mM paraquat (equivalent to 40 mg/kg i.v.) showed selective mitochondrial breakage at 6–24 hr and died at 36–48 hr. These results suggest that mitochondria are the initial toxic site of paraquat as well as in contrast to the previously proposed microsome theory. [ABSTRACT FROM PUBLISHER]

The cytochrome oxidase activity of rat dorsal root ganglion cells was cytochemically and quantitatively measured. The 100 μm thick tissue slices fixed for 10 min were incubated in a DAB medium for 0, 30, 60, and 90 min at 37°C, and electron-dense deposit areas within the mitochondrial intermembrane-intracristal spaces were measured with a computer-controlled image analyzer. The activity was expressed as deposit accumulation rate filling the mitochondrial space, (1 unit corresponds to the deposit filling 100% of the mitochondrial space per hour). Three different activities of the enzyme in either large pale cells or small dark cells of sensory neurons could be distinguished, based on the quantitative analysis, as large cells with intense (0.83 units), intermediate (0.38), and weak activity (0.22), and small cells with the same degrees of activity (1.04, 0.35, and 0.11, respectively). The results indicate that accumulation rate measurements of reaction product may be useful to quantitatively present the cytochrome oxidase reactivity of mitochondria, and that the degree of enzyme activity may contribute to the identification of functional differences in sensory neurons. [ABSTRACT FROM PUBLISHER]

Cytochemically demonstrated cytochrome oxidase activities in mitochondria of rat pulmonary alveolar epithelial cells were quantitatively compared. As the standardized procedures, lungs were fixed for 10 mm at 4°C with 2% pure glutaraldehyde, and incubated for different times at 37°C in a medium containing 1.0 mg/ml 3,3'-diaminobenzidine-4HCl, 1 mg/ml cytochrome c, 0.1 mg/ml catalase, 7% sucrose, anb 0.1 M phosphate buffer, pH 7.4. Electron micrographs were then odtained, and the densities of the reaction deposits in the mitochondrial intermembrane-intracristal space were measured in an image analyzer, and were plotted in terms of time in minutes. The initial velocity (maximal rate) of the activity expressed as deposit accumulation rate (% area/60 min) filling the mitochondrial intermembrane-intracristal space of Type I, Type II and Type III cells was 40, 130 and 9.3, respectively. These results indicated that mitochondria are varied in the intensity of cytochrome oxidase activity among 3 types of pulmonary alveolar epithelial cells, and that the image-analyzing measurement of the accumulation rate of reaction product may be useful for quantitative analysis of enzyme activity, in particular, in the cells which are difficult to isolate from complex tissues. [ABSTRACT FROM PUBLISHER]

The ultrastructural localization of uterine peroxidase activity in the human endometrium was studied. Very weak peroxidase activity was present in a few surface nonciliated cells with very weak activity in the early proliferative phase; the activity was enhanced and distributed throughout the nonciliated cells during the late proliferative to secretory phase. The activity was observed in the endoplasmic reticulum and nuclear envelope but not in the secretory granules and Golgi apparatus. The surface ciliated cells and all glandular cells were entirely devoid of the activity throughout the menstrual cycle. Nucleolar channel system was transiently formed in the surface and glandular nonciliated cells in the postovulatory phase. The channels of the system in the surface cells showed intense peroxidase activity, but those in the glandular cells were negative. These results indicate that the human uterine peroxidase activity appears only in the superficial nonciliated cells and changes in its intensity depending on the menstrual cycle, and that the nucleolar channel system is closely related with the nuclear envelope showing peroxidase activity. [ABSTRACT FROM PUBLISHER]

The platinum-diaminobenzidine (Pt-DAB) reaction, which yields a black, electron-dense, insoluble, Pt-containing reaction product at the active site of cytochrome oxidase without postosmification, has been developed. This reaction permits us to quantitate the oxidase activity in an individual mitochondrion or in its constituent part by the energy dispersive X-ray analysis (EDAX), based on the platinum atoms incorporated in the product. [ABSTRACT FROM PUBLISHER]

Epoxy resin-embedded metal standards were prepared for X-ray microanalysis of cerium, nickel, and cobalt. Acetyl acetonate compounds of these metals, which were dissolved in Quetol 651 epoxy resin, up to 204 mM, were used as metal standards. Microanalysis of the standards gave straight calibration lines between X-ray counts and concentrations. These standards are useful for the quantitative microanalysis of cytochemically reacted specimens containing those metal deposits. [ABSTRACT FROM PUBLISHER]