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HIMES, MARION M.; POLLISTER, ARTHUR W.
doi: 10.1177/10.2.175pmid: N/A
Hepatic intranuclear glycogen of the larva of Rana pipiens has been studied by the periodic acid Schiff reaction (controlled by diastase digestion), by electron microscopy, and by interference and phase microscopy. The glycogen occurs sporadically as one or two masses in the central part of the nucleus. The absence of nuclear envelope and of typical non-glycogen cytoplasmic elements shows that the nuclear glycogen inclusions are not cytoplasmic areas that have been enclosed within the nucleus. The central position, difference in rate of resorption, and other evidence indicates that the nuclear glycogen has been synthesized in situ. Comparison with the nucleolus suggests that the nuclear glycogen body is formed as a result of activity at a specific chromosomal site.
NIEMI, MIKKO; HÄRKÖNEN, MATTI; KOKKO, AULIKKI
doi: 10.1177/10.2.186pmid: N/A
Both the Leydig and Sertoli cells of the rat testis have been shown to be rich in non-specific esterase, but lacking in cholinesterase and "true" lipase. No esterolytic activity has been observed in other elements of testicular tissue. The enzyme activity of the Sertoli cells is mainly localized to the long cytoplasmic processes of these cells, where the sperm heads are embedded. The distribution of esterase activity inside the seminiferous tubules undergoes cyclic variations corresponding to those seen in the morphology of the Sertoli cells.An essentially similar distribution of esterase activity was noticed when α-naphthyl acetate, naphthol-AS acetate, o-indoxyl acetate or Tweens 20 to 60 were used as substrates. Indoxyl acetate was, however, hydrolysed even in an acid medium and this activity was, furthermore, resistant to an organophosphorus inhibitor (E600). It is therefore concluded that the testicular Leydig and Sertoli cells have both A-type and B-type esterase activity, of which the former behaves much like an intracellular peptidase. Demonstration of the activity of electrophoretically separated enzyme proteins have further shown that the histochemically demonstrated non-specific esterase activity in the testis is composed of about eight different enzyme proteins, whose behavior towards inhibitors corresponds to that of tissue sections.
MANCINI, R. E.; VILAR, O.; DELLACHA, J. M.; DAVIDSON, O. W.; GOMEZ, C. J.; ALVAREZ, B.
doi: 10.1177/10.2.194pmid: N/A
Adult albino rats were injected intravenously with labelled homologous albumin, globulins and fibrinogen. Labelling was done with acid rhodamine B (C.I. No. 45100) and fluorescein isothiocyanate. The amount of fluorescent dye bound to proteins was measured spectrophotometrically. Paper electrophoretic mobility and clotting time of fibrinogen were also checked after labelling. As a control, free fluorescent dye solutions were injected into another lot of rats.Animals were killed at intervals from 10 minutes to 12 days. Blood samples were taken to determine spectrophotometrically the concentration of labelled proteins in the circulation. Paraffin or unfixed fresh frozen sections from all tissues were examined with the fluorescence microscope.Histoimmunological studies were also made using fluorescent antibody against the different rat serum fractions. Coons's technique was applied to tissue sections from normal rats or from animals previously injected with unlabelled serum fractions and killed between 3 and 12 hours later. The following principal observations were made:1) Globulins bind a larger amount of dye than albumin, while fibrinogen bound the least.2) Labelling does not change grossly the electrophoretic mobility of albumin and globulins but slightly modified the clotting time of fibrinogen.3) Time-concentration curves of albumin and globulins in the circulation are very much alike and reveal a fast and a slow component with 2.6 day time half-life for the former and 3.1 half-life for the latter.4) Histologically labelled albumin and globulins are seen first in the lumen and on the walls of all blood vessels, and up to 8 hours they appeared to accumulate widely in connective tissue structures as well as in basement membranes of dermis and the mucosae of digestive, respiratory and urogenital tracts; fluorescence does not cross the basement membranes in most organs, nor does it pass the perichondrium, periosteum, or blood capillaries in the nervous system into surrounding parenchyma. Finally, labelled proteins began to disappear from all these sites after 12 hours, but persist longer in the macrophages of liver, lymph nodes, spleen and bone marrow. Fibrinogen was not seen diffusing out from vessels to the surrounding connective tissue.5) That in animals previously injected with unlabelled serum fractions, histoimmunological techniques also showed the highest specific fluorescence in sections of connective tissue, stained with labelled antialbumin and antiglobulins; labelled antifibrinogen gave only a weak fluorescence in the same structutres.6) These results confirm and extend our previous studies on the role of connective tissue as a transient depot for the extravascular pool of serum proteins.
doi: 10.1177/10.2.204pmid: N/A
Since the removal of only one of the two nucleic acids, ribonucleic and deoxyribonucleic, is often necessary in biological work, the optimum conditions for the specific removal of either nucleic acid by the corresponding enzyme were investigated. The specific removal of nucleic acids by enzyme treatment was tested by toluidine blue staining and by quantitative study of the radioautographic reactions produced by the labelled deoxyribonucleic acid and ribonucleic acid appearing in tissues of a cytidine-H3 injected mouse.For the removal of ribonucleic acid by ribonuclease, the tissues must be fixed in freshly prepared Carnoy's solution for 24 hours, as a shorter fixation permits the loss of ribonucleic acid from control tissue sections. Ribonuclease extraction (1 mg crystallized salt-free Worthington ribonuclease per 1 ml of distilled water) is carried out for 4 hours at 40°C.For the removal of deoxyribonucleic acid by deoxyribonuclease, it was also necessary to use 24 hour fixation in freshly prepared Carnoy. A high concentration of magnesium was found to be necessary in the incubation solution (0.05 mg crystallized Worthington deoxyribonuclease per 1 ml of 0.2 M Mg-containing Gomori's Tris maleic acid buffer solution). The incubation was carried out for 24 hours at 37°C.The toluidine blue stained basophilic material and the radioautographic reactions in the tissues of the cytidine-H3 injected mouse were lost following a double treatment with ribonuclease and deoxyribonuclease or by hot trichloracetic acid extraction. Therefore all the reactions appearing over the histological tissue sections are due to the tritium incorporated into deoxyribonucleic acid and ribonucleic acid.The basophilic material from cytoplasm and nucleolus was removed specifically and completely by ribonuclease treatment. The deoxyribonuclease treatment removed nuclear basophilia exclusively. However, controls of ribonuclease or deoxyribonuclease treatment showed no loss of basophilia, except on occasion in the pancreas.The radioautographic reactions found in the mouse tissues 24 hours after cytidine-H3 injection were exclusively localized over the nuclear basophilia of some nuclei after ribonuclease treatment, but were largely over the cytoplasmic basophilia after deoxyribonuclease treatment. Quantitative radioautographic study showed that either enzyme treatment removed its specific substrate completely and exclusively without loss in control preparations.
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