The present study was undertaken to provide more information on the peripheral RNA containing ring of ringshaped nucleoli (RSNo). Human lymphocytes of blood donors and patients suffering from B chronic lymphocytic leukemia mostly characterized by RSNo represented very convenient cell models for such study. According to the light microscopy the peripheral RNA ring possessed several highly condensed foci. Such regions represented accumulated dense RNA fibrillar components (DFCs) seen by the electron microscopy. In contrary, the incidence of dense granular RNA-containing components (GCs) in surrounding portions of the RNA ring was small. Thus, the structural and morphological organization of the peripheral RNA ring of RSNo apparently reflects sites of micro-segregated foci of DFCs and a small incidence of GCs. That structural organization of the peripheral RNA ring of RSNo appeared to be a prerequisite for further regressive nucleolar changes resulting in the development of micronucleoli in terminal lymphocytes.
- MeSH
- buněčné jadérko * ultrastruktura patologie MeSH
- chronická lymfatická leukemie * patologie MeSH
- dárci krve * MeSH
- lidé MeSH
- lymfocyty * patologie MeSH
- RNA MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Protein arginine methyltransferases (PRMTs) are responsible for symmetric and asymmetric methylation of arginine residues of nuclear and cytoplasmic proteins. In the nucleus, PRMTs belong to important chromatin modifying enzymes of immense functional significance that affect gene expression, splicing and DNA repair. By time-lapse microscopy we have studied the sub-cellular localization and kinetics of PRMT1 after inhibition of PRMT1 and after irradiation. Both transiently expressed and endogenous PRMT1 accumulated in cytoplasmic bodies that were located in the proximity of the cell nucleus. The shape and number of these bodies were stable in untreated cells. However, when cell nuclei were microirradiated by UV-A, the mobility of PRMT1 cytoplasmic bodies increased, size was reduced, and disappeared within approximately 20 min. The same response occurred after γ-irradiation of the whole cell population, but with delayed kinetics. Treatment with PRMT1 inhibitors induced disintegration of these PRMT1 cytoplasmic bodies and prevented formation of 53BP1 nuclear bodies (NBs) that play a role during DNA damage repair. The formation of 53BP1 NBs was not influenced by PRMT1 overexpression. Taken together, we show that PRMT1 concentrates in cytoplasmic bodies, which respond to DNA injury in the cell nucleus, and to treatment with various PRMT1 inhibitors.
- MeSH
- chromozomální proteiny, nehistonové genetika metabolismus MeSH
- cytoplazma enzymologie MeSH
- DNA vazebné proteiny genetika metabolismus MeSH
- HeLa buňky MeSH
- intracelulární signální peptidy a proteiny genetika metabolismus MeSH
- lidé MeSH
- myši MeSH
- poškození DNA * MeSH
- proteinarginin-N-methyltransferasy antagonisté a inhibitory genetika metabolismus MeSH
- represorové proteiny antagonisté a inhibitory genetika metabolismus MeSH
- ultrafialové záření * MeSH
- záření gama * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
A comparison of histochemical detection of GM1 ganglioside in cryostat sections using cholera toxin B-subunit after fixation with 4% formaldehyde and dry acetone gave tissue-dependent results. In the liver no pre-treatment showed detectable differences related to GM1 reaction products, while studies in the brain showed the superiority of acetone pre-extraction (followed by formaldehyde), which yielded sharper images compared with the diffuse, blurred staining pattern associated with formaldehyde. Therefore, the aim of our study was to define the optimal conditions for the GM1 detection using cholera toxin B-subunit. Ganglioside extractability with acetone, the ever neglected topic, was tested comparing anhydrous acetone with acetone containing admixture of water. TLC analysis of acetone extractable GM1 ganglioside from liver sections did not exceed 2% of the total GM1 ganglioside content using anhydrous acetone at -20 degrees C, and 4% at room temperature. The loss increased to 30.5% using 9:1 acetone/water. Similarly, photometric analysis of lipid sialic acid, extracted from dried liver homogenates with anhydrous acetone, showed the loss of gangliosides into acetone 3.0 +/- 0.3% only. The loss from dried brain homogenate was 9.5 +/- 1.1%. Thus, anhydrous conditions (dry tissue samples and anhydrous acetone) are crucial factors for optimal in situ ganglioside detection using acetone pre-treatment. This ensures effective physical fixation, especially in tissues rich in polar lipids (precipitation, prevention of in situ diffusion), and removal of cholesterol, which can act as a hydrophobic blocking barrier.
- MeSH
- aceton chemie MeSH
- cholerový toxin chemie MeSH
- cholesterol analýza MeSH
- G(M1) gangliosid analýza chemie MeSH
- imunohistochemie MeSH
- játra chemie cytologie MeSH
- krysa rodu rattus MeSH
- mozek cytologie MeSH
- potkani Wistar MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Fibre type determination requires a large series of differently stained muscle sections. The manual identification of individual fibres through the series is tedious and time consuming. This paper presents a software that enables (i) adjusting the position of individual fibres through a series of differently stained sections (image registration) and identification of individual fibres through the series as well as (ii) muscle fibre classification and (iii) quantitative analysis. The data output of the system is the following: numerical and areal proportions of fibre types, fibre type size and optical density (grey level) of the final reaction product in every fibre. The muscle fibre type can be determined stepwise, based on one set of stained sections while further, newly stained sections can be added to the already defined muscle fibre profile. Several advantages of the presented software application in skeletal muscle research are presented. The system is semiquantitative, flexible, and user friendly.
- MeSH
- hybridizace in situ MeSH
- imunohistochemie MeSH
- kosterní svalová vlákna cytologie klasifikace MeSH
- krysa rodu rattus MeSH
- lidé MeSH
- musculus masseter cytologie MeSH
- myosiny genetika metabolismus MeSH
- počítačové zpracování obrazu metody MeSH
- protein - isoformy genetika metabolismus MeSH
- reprodukovatelnost výsledků MeSH
- software MeSH
- těžké řetězce myosinu genetika metabolismus MeSH
- uživatelské rozhraní počítače MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- zvířata MeSH
- Publikační typ
- práce podpořená grantem MeSH
The number of nucleoli in a cell and nucleolar area vary according to the cell. We compared nucleoli in mammalian circulating lymphocytes and insect circulating haemocytes. An increased nucleolar coefficient correlated with a lowered nucleoli size. The smaller nucleolar size in mammalian lymphocytes indicates a lower proteosynthetic cellular activity in both mammalian lymphocytes and insect haemocytes. Moreover, in insect haemocytes, the smaller size of the nucleoli may reflect a lowered potential to transform into another cell type.
- MeSH
- buněčné jadérko ultrastruktura MeSH
- cizopasní červi MeSH
- dospělí MeSH
- hemocyty ultrastruktura MeSH
- hmyz MeSH
- krysa rodu rattus MeSH
- lidé středního věku MeSH
- lidé MeSH
- lymfocyty ultrastruktura MeSH
- mladiství MeSH
- myši MeSH
- zvířata MeSH
- Check Tag
- dospělí MeSH
- krysa rodu rattus MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- srovnávací studie MeSH
The present study was designed to provide more information on nucleoli in apoptotic cells,which were represented in the present study by cultured leukemic myeloblasts (Kasumi-1 cells). The apoptotic process in these cells was produced by trichostatin A (TSA) that is a histone deacetylase inhibitor with strong cytostatic effects. The selected TSA concentration added to cultures facilitated to study apoptotic and not-apoptotic cells in one and the same specimen. The nucleolar diameter and density were determined using computer assisted measurement and densitometry in specimens stained for RNA. In comparison with not-apoptotic cells, in apoptotic cells, nucleolar mean diameter did not change significantly and nucleolar RNA density was also not apparently different. On the other hand, the cytoplasmic RNA density in apoptotic cells was markedly reduced. Thus it seemed to be possible that the transcribed RNA remained "frozen"within the nucleolus but its transport to the cytoplasm decreased or stopped. However, the possibility of the RNA degradation in the cytoplasm of apoptotic cells based on the present study cannot be eliminated. At this occasion it should be added that AgNORs reflecting nucleolar biosynthetic and cell proliferation activity in apoptotic cells decreased in number or disappeared. The presented results also indicated that large nucleoli intensely stained for RNA need not be necessarily related to the high nucleolar biosynthetic or cell proliferation activity and may be also present in apoptotic cells responding to the cytostatic treatment.
- MeSH
- antigeny jaderné analýza ultrastruktura MeSH
- apoptóza MeSH
- buněčné jadérko chemie účinky léků ultrastruktura MeSH
- chronická myeloidní leukemie patologie MeSH
- kyseliny hydroxamové farmakologie MeSH
- lidé MeSH
- nádorové buněčné linie MeSH
- prekurzorové buňky granulocytů účinky léků ultrastruktura MeSH
- RNA analýza MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Mean diameter of nucleolar bodies (nucleoli without the perinucleolar chromatin) per cell was studied in human leukemic myeloblasts represented by K 562 and Kasumi 1 cell lines which originated from chronic and acute myeloid leukaemia. The measurement of mean diameter of nucleolar bodies in specimens stained for RNA was very simple. Such approach eliminated the variability of the perinucleolar chromatin discontinuous shell which might influence the measured nucleolar size as suggested by earlier studies. Ageing of K 562 myeloblasts produced a significant decrease of cells in S+G2 phase of the cell cycle accompanied by a significant reduction of mean diameter of nucleolar bodies (MDNoBs) per cell. In contrast, treatment of Kasumi 1 myeloblasts with histone deacetylase inhibitor - Trichostatin A - produced a large incidence of resistant cells in S+G2 phase which were characterised by a large increase of MDNoBs. Thus, MDNoBs in leukemic myeloblasts might be a helpful tool to estimate the incidence of cells in the S+G2 phase at the single cell level in smear preparations when the number of cells is very small.
- MeSH
- akutní erytroblastická leukemie genetika patologie MeSH
- antigeny jaderné MeSH
- buněčné jadérko genetika patologie MeSH
- buňky K562 MeSH
- chronická myeloidní leukemie genetika patologie MeSH
- financování organizované MeSH
- G2 fáze fyziologie MeSH
- jaderné proteiny MeSH
- lidé MeSH
- organizátor jadérka patologie MeSH
- počet buněk MeSH
- prekurzorové buňky granulocytů patologie MeSH
- proliferace buněk MeSH
- RNA nádorová analýza MeSH
- S fáze fyziologie MeSH
- Check Tag
- lidé MeSH
- MeSH
- apoptóza účinky záření MeSH
- buněčné jadérko chemie účinky léků ultrastruktura MeSH
- finanční podpora výzkumu jako téma MeSH
- fotochemoterapie MeSH
- fotosenzibilizující látky farmakologie MeSH
- kyselina aminolevulová farmakologie MeSH
- leukemie farmakoterapie patologie MeSH
- lidé MeSH
- obrovské buňky účinky léků ultrastruktura MeSH
- Check Tag
- lidé MeSH
