K. Smetana

Further studies on the cytochemistry of the standardized silver staining of interphase nucleoli in smear preparations of yoshida ascitic sarcoma cells in rats

SummarySilver staining procedure for the selective demonstration of nucleolar silver stained granules (SSG) and for the simultaneous demonstration of SSG and nucleolar silver stained matrix (SSM) were studied in smears of rat Yoshida sarcoma cells. The successful results of these procedures depend mainly on the quality of silver nitrate and formaldehyde. However, both chemicals can be easily standarized and stabilized disregarding their origin and batch. In standardized procedures (one-step procedure for the selective demonstration of SSG and two-steps procedure for the simultaneous demonstration of SSG and SSM) the silver is apparently bound to acidic groups of proteins of SSG and SSM. The proteins of SSG and SSM seem to be different but both belong to the group of acidic non-histone proteins. According to the results of digestion experiments a possibility also exists that the acidic proteins of SSG may be associated with DNA. The identification of SSG visualized by described standardized procedures was determined not only by cytochemical extraction tests but also by biological experiments. The latter demonstrated that the number of SSG in Yoshida sarcoma cells decreases after treatment of experimental animals with actinomycin D and therefore depends on the state of the nucleolar RNA synthesis.

Ring shaped nucleoli in liver cells of rats after treatment with Actinomycin D

SummaryThe incidence of ring shaped nucleoli was studied by means of light and electron microscopical methods in liver cells of rats injected intravenously with Actinomycin D in a dose of 150 μg per kg body weight.Ring shaped nucleoli were most frequent 20 minutes after administration of Actinomycin D. The number of fragmented nucleoli was increased 20 minutes after administration of the drug and did not decrease even after 72 hours.Fibrillar ring like inclusions within nuclei and more perichromatin granules at the periphery of nucleoli were observed after treatment with Actinomycin D.ZusammenfassungDie Wirkung von Aktinomycin D (150 μg/kg) auf die Leberzellen der Ratte wurde licht- und elektronenenmikroskopisch untersucht. 20 min nach intravenöser Aktinomycin-D-Gabe wurden ringörmige Nukleolen beobachtet. Die Zahl der fragmentierten Nukleolen war 20 min nach der Applikation gesteigert. Ihre relative Zahl änderte sich nach Aktinomycingabe nicht. Nach Behandlung der Tiere mit Aktinomycin D kann man in den Kernen der Leberzellen fibrilläre ringförmige Einschlüsse und eine größere Menge von „perichromatin-granules“ an der Peripherie der Nukleolen beobachten.

Analysis of ring-shaped nucleoli in serially sectioned human lymphocytes

SummarySerial section analysis has demonstrated that ring-shaped nucleoli of mature human lymphocytes are spherical structures consisting of a peripheral ribonucleoprotein shell that surrounds one large fibrillar center. The shell exhibits usually one or, less frequently, two openings. The fibrillar center is in contact with the nucleoplasm and perinucleolar condensed chromatin, which frequently appears as a pedicle-like structure. Several chromocenters are associated with the ring-shaped nucleolus.

Nucleolar silver-stained granules in maturing erythroid and granulocytic cells.

SummaryHuman and rabbit erythroid and granulocytic precursors in bone marrow have been investigated to provide information concerning the number of nucleolar silverstained granules (SSGs), which represent active interphasic nucleolar organizer regions (NORs). The differentiation and maturation of precursor cells of both investigated cell lines are characterized by a gradual decrease in number of nucleolar SSGs. In advanced maturation stages of erythroblasts or granulocytes, which are known to lose the capacity to divide, the number of nucleolar SSGs is smaller than the reported average or maximal values of NORs determined for human or rabbit cells. Since committed stem cells from both cell lines contain several times the number of nucleolar SSGs than the last dividing maturation and differentiation stages, the number of active parts of interphasic NORs in committed stem cells seems to be increased and might represent a stock for the later stages. In addition, the number of nucleolar SSGs appear to be a very convenient marker of nucleolar biosynthetic activity in individual differentiating and maturing blood cells. The differences between erythroid and granulocytic stem cells with respect to the number of nucleolar SSGs disappear during the course of further differentiation and maturation.

Nucleolar changes in maturing avian erythroblasts and erythrocytes

SummaryMaturing erythroblasts and erythrocytes were studied in chickens and adult hens to provide more information on the presence and frequency of various nucleolar types in these cells. Nucleoli were present at all stages of erythroblastic and erythrocytic development except in the case of a few reticulocytes and the mature erythrocytes. The number of nucleoli per cell (expressed as the nucleolar coefficient) reached a maximum at the stage of the polychromatic erythroblast. Early erythroblasts were characterized by the presence of compact nucleoli or nucleoli with nucleolonemata. Ring shaped nucleoli and micronucleoli increased in number with further maturation. Cells of the final erythroblast stage (orthochromatic erythroblasts) contained mostly micronucleoli, and micronucleoli alone were present in reticulocytes and mature erythrocytes.

Ultrastructural localization of rRNA in HeLa cells, rat liver cells andXenopus laevis oocytes by means of the monoclonal antibody-protein A-gold technique

SummaryThe postembedding localization of rRNA was investigated in ultrathin sections of HeLa cells, rat liver andXenopus laevis oocytes by means of the monoclonal antibody to rRNA and protein A-gold technique. The incidence of gold particles was highest in nucleoli and cytoplasmic areas containing ribosomes. The chromosomes were labelled less than the surrounding cytoplasm in mitotic HeLa cells. In nucleoli of HeLa cells and rat hepatocytes, the labelling of areas containing ribonucleoprotein components was greater than the labelling of fibrillar centres. In segregated nucleoli ofX. laevis oocytes, the labelling of the granular region substantially exceeded that of the fibrillar regions. The incidence of nucleoplasmic gold particles in interphasic HeLa cells was found to be slightly increased in the vicinity of nucleoli. The labelling of clusters of interchromatin granules in rat hepatocytes was not significantly different from that of the rest of the nucleophasmic interchromatin spaces.

The number of nucleolar silver-stained granules of active nucleolus organizer regions in mitotic and interphase cells of rat Yoshida and Zajdela ascitic tumors

SummaryTo provide more information on the number of silver-stained granules (SSGs) of active mitotic nucleolus organizer regions (NORs) as well as interphase nucleoli, rat Yoshida ascitic sarcoma and Zajdela ascitic hepatoma cells were studied by means of standardized one-step and two-step silver-staining procedures. The number of SSGs of mitotic active NORs was relatively constant and corresponded to the number of active NORs of animals bearing the tumors investigated. Some anaphases and telophases were “asymmetric”, i.e. chromosomal figures (future nuclei) in one and the same cell contained different number of SSGs (active NORs). The incidence of such asymetric anaphases and telophases was higher in aneuploid (hypoploid) Zajdela hepatoma than in euploid (diploid) Yoshida sarcoma cells. In the interphase, the number of SSGs was low in small or large cells with distinct chromocenters or chromosomes condensation presumably representing postmitotic and premitotic cells. In contrast, the highest number of SSGs was noted in nucleoli of large cells which were usually characterized by a fine chromatin structure.

Morphological changes in nucleoli of lymphocytes and their influencing by trimethazone, phenylbutazone, indometacin, and 6-mercaptopurine in albino rats with adjuvant-induced disease

SummaryThe incidence of diverse types of nucleoli in peripheral blood lymphocytes was established in albino rats with adjuvant-induced disease, either untreated or receiving trimethazone, phenylbutazone, indometacin, or 6-mercaptopurine. Apart from typical clinical symptoms, the adjuvant-induced disease was characterized by an increase in the number of active nucleoli in the lymphocytes present in the peripheral blood of the diseased animals. This increase was influenced by the administration of any of the drugs enumerated.

Studies on satellite nucleoli in human normal and leukemic lymphocytes.

SummaryLymphocytes from normal and leukemic patients, and phytohemagglutinin-stimulated lymphocytes were investigated by means of immunofluorescence procedures and a silver reaction for the demonstration of proteins characteristic of nucleolus organizer regions in interphasic cells to provide basic information on the presence of satellite nucleoli in these cells. The results clearly indicated that satellite nucleoli are present in a limited but constant percentage of peripheral lymphocytes. An increased percentage of lymphocytes with satellite nucleoli was found only in leukemic patients and after silver staining. In contrast, a decreased percentage of satellite nucleoli was found 24 h after stimulation with phytohemagglutinin in vitro. In leukemic patients, the discrepancy in the percentage of lymphocytes with satellite nucleoli between immunostained and silver-stained preparations may suggest that the silver reaction demonstrates the presence of an additional argyrophilic protein besides proteins B23 and C23, or altered forms of these proteins, which does not react with the specific antibodies.

Nuclear and nucleolar effect of experimental endotoxemia on rabbit hepatocytes

SummaryNuclear and nucleolar ultrastructure was studied in hepatocytes of rabbits intravenously injected with a single dose of endotoxin to provide more information on the ultrastructural changes of hepatocytes produced by the endotoxemia. The observed changes were represented by a clumping of interchromatin granules, alteration of nucleolar architecture and formation of rodlet-like intranuclear inclusions. The alteration of the nucleolar architecture was characterized by loosely arranged rope-like nucleolonemata, partial retraction of the perinucleolar chromatin and increased numbers of fibrillar centers.