Large nucleoli have generally been believed to be present in less differentiated and proliferating cells including the malignant ones. Such nucleoli have also been considered to be active in the biosynthetic process and major cell developmental activities. In contrast, after cytostatic treatment, apoptotic leukaemic progenitors still containing nuclei did not exhibit substantial reduction of the nucleolar size but displayed decreased nucleolar biosynthetic activity. The present study was undertaken to provide more information on the large nucleoli in spontaneously occurring apoptotic leukaemic progenitors without further differentiation. Leukaemic progenitors of established cell lineages originating from leukaemic patients represented a very convenient model for such study. Some of them exhibit morphological signs of the spontaneously occurring apoptotic process. Since such signs are expressed by nuclear and cytoplasmic morphological variability, the present study dealt with spontaneously occurring apoptotic progenitors with preserved nuclei characterized by heavy chromatin condensation and occasional fragmentation. Based of nucleolar body and nuclear maximal diameter measurements it seems to be clear that the nucleolar size in these cells was not substantially reduced, contrary to that of the nucleus. However, large nucleolar bodies in spontaneously occurring apoptotic cells were characterized by markedly reduced biosynthetic activity, as expressed by the decreased number of nucleolar transcription markers such as nucleolar fibrillar centres. In conclusion, large nucleoli may be present not only in proliferating, but also in spontaneously occurring apoptotic cells.
The appearance of heterochromatin is generally accepted as a useful tool for the evaluation of the cell state including pathology; however, information on the heterochromatin DNA condensation state expressed by the image optical density in interphase nuclear regions and mitotic chromosomes with silent genes is very limited. Since human proliferating myeloblasts are a very convenient model, they were studied in the bone marrow of leukemic patients and established cell cultures using computer assisted image densitometry at the single cell level after heterochromatin visualization by a simple but sensitive cytochemical procedure for demonstration of DNA. As was expected, a high DNA image optical density was noted in central heterochromatin regions in contrast to the nuclear periphery at the nuclear envelope. Similarly, a high nuclear DNA image optical density was also expressed in mitotic chromosomes. Thus the possibility exists that the large heterochromatin DNA condensation expressed by the large image optical density in central nuclear regions, as in mitotic chromosomes, is related to silent gene locations. The similar width of mitotic chromosomes and chromatin fibrils in the heterochromatin regions in the interphase nuclei supports that explanation. The chromatin DNA fibrils in the central heterochromatin nuclear regions of interphase cells might just represent masked silent chromosomal segments. Such a conclusion is in harmony with “classical” cytology in the first part of the last century, which suggests the chromosome continuity from the mitotic division to the interphase where each chromatin region (“Kernbezirk”) actually represents a chromosomal territory.
- MeSH
- barvení a značení MeSH
- chromatin genetika izolace a purifikace MeSH
- chromozomy genetika MeSH
- DNA nádorová genetika MeSH
- financování organizované MeSH
- heterochromatin genetika izolace a purifikace MeSH
- leukemie etiologie genetika MeSH
- lidé MeSH
- mikroskopie metody využití MeSH
- nádorové buněčné linie MeSH
- prekurzorové buňky granulocytů cytologie imunologie MeSH
- proliferace buněk MeSH
- struktury buněčného jádra genetika MeSH
- Check Tag
- lidé MeSH
The present study was undertaken to provide more information on nucleolar changes induced by a histone deacetylase inhibitor such as valproic acid in leukaemic myeloblasts at the single-cell level. For this study, RNA in nucleoli was visualized by a simple but sensitive cytochemical procedure in unfixed cytospins of short-term bone marrow cultures from patients suffering from acute myeloid leukaemia. Valproic acid in leukaemic myeloblasts markedly reduced the nucleolar size and also produced significant transformation of "active" to "resting" and "inactive" nucleoli that reflected the alteration of the nucleolar transcription in sensitive myeloblasts. On this occasion it should be added that valproic acid significantly increased the incidence of altered myeloblasts that changed to apoptotic cells or apoptotic bodies and cell ghosts. In contrast to the above-mentioned decreased nucleolar size, the nucleolar RNA concentration, expressed by computerassisted RNA image densitometry in valproic acidtreated myeloblasts, was not significantly changed. The results of the present study clearly indicated that the nucleolar size and transformation of "active" to "sleeping" or "inactive" nucleoli are convenient markers of the sensitivity and alteration of leukaemic myeloblasts produced by a histone deacetylase inhibitor, valproic acid, at the single-cell level.
- Klíčová slova
- Histone deacetylase inhibitors,
- MeSH
- akutní myeloidní leukemie MeSH
- buněčné jadérko účinky léků ultrastruktura MeSH
- financování organizované MeSH
- histondeacetylasy farmakologie MeSH
- kultivované buňky MeSH
- kyselina valproová farmakologie MeSH
- lidé MeSH
- prekurzorové buňky granulocytů cytologie účinky léků MeSH
- tvar buňky MeSH
- Check Tag
- lidé MeSH
The present study was undertaken to provide more information on the density and distribution of heterochromatin in early and advanced stages of the granulocytic, lymphocytic and erythroid development. Heterochromatin was visualized using a simple cytochemical method for the demonstration of DNA followed by computer-assisted densitometry of the digitized images. The largest heterochromatin density in early proliferating stages of all studied blood cell lineages was noted in the perinucleolar region and centrally located chromocentres. In contrast, the heterochromatin density at the nuclear membrane was significantly lower. In advanced nonproliferating stages or apoptotic cells the heterochromatin density increased and was similar in all nuclear regions, i.e. in the perinucleolar regions, chromocentres, and at the nuclear membrane. Thus, such observations indicated that the heterochromatin condensation in the perinucleolar region and chromocentres, i.e. in "gene-rich nuclear regions", of differentiating and maturing progenitors of blood cells preceded that at the nuclear periphery.
- MeSH
- apoptóza MeSH
- buněčná diferenciace MeSH
- buněčné linie MeSH
- buňky kostní dřeně cytologie MeSH
- erytroidní prekurzorové buňky cytologie MeSH
- hematopoetické kmenové buňky cytologie MeSH
- heterochromatin metabolismus MeSH
- HL-60 buňky MeSH
- intracelulární membrány metabolismus MeSH
- lidé MeSH
- prekurzorové buňky granulocytů cytologie MeSH
- proliferace buněk MeSH
- Check Tag
- lidé MeSH
OBJECTIVES: MicroRNAs (miRNAs) play key roles in a wide variety of normal and pathological cellular processes. A number of studies identified hematopoietic-specific miRNAs that are necessary for correct function of blood cells. Out of our microarray data, we chose 13 miRNAs that showed differential expression in peripheral blood cells (miR-15b, miR-16, miR-24, miR-30c, miR-106b, miR-142-3p, miR-142-5p, miR-150, miR-155, miR-181, miR-223, miR-342, and miR-451) and examined their expression in separated hematopoietic cell lineages. METHODS: Using quantitative real-time polymerase chain reaction, we measured relative expression of the miRNAs in fractions of reticulocytes, platelets, granulocytes, monocytes, B- and T-lymphocytes as well as in several hematopoietic cell lines. RESULTS: We observed that miR-16 and miR-142-3p were highly expressed in all native cell lineages, miR-451 reached the maximal expression in reticulocytes, miR-223 in platelets, granulocytes and monocytes, and miR-150 in B- and T-lymphocytes. Hierarchical clustering analysis grouped the lineage samples according to their origin based on the expression of these miRNAs. To validate discrimination power of the miRNAs, we quantified expression of the 13 miRNAs in several immortalized cell lines. Although the cell lines showed miRNA expression patterns considerably different from those of native cell lineages, clustering analysis distinguished between myeloid, lymphoid and non-hematopoietic cells. CONCLUSIONS: In conclusion, the study reports the expression levels of 13 miRNAs in particular blood cell lineages as well as immortalized cell lines. We demonstrate that the expression profiles of these miRNAs may be used for discrimination of the hematopoietic cell lineages.
- MeSH
- buněčná diferenciace fyziologie MeSH
- buňky K562 MeSH
- financování organizované MeSH
- HeLa buňky MeSH
- leukocyty cytologie metabolismus MeSH
- lidé MeSH
- lymfoidní progenitorové buňky cytologie metabolismus MeSH
- mikro RNA biosyntéza MeSH
- prekurzorové buňky granulocytů cytologie metabolismus MeSH
- regulace genové exprese fyziologie MeSH
- retikulocyty cytologie metabolismus MeSH
- sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů metody MeSH
- stanovení celkové genové exprese metody MeSH
- trombocyty cytologie metabolismus MeSH
- Check Tag
- lidé MeSH
- MeSH
- apoptóza fyziologie genetika MeSH
- buněčná smrt fyziologie genetika MeSH
- buněčné jadérko fyziologie genetika MeSH
- buňky - růstové procesy fyziologie genetika MeSH
- elektronová mikroskopie metody využití MeSH
- financování organizované MeSH
- geny rRNA fyziologie genetika MeSH
- histocytochemie metody využití MeSH
- krevní buňky cytologie fyziologie metabolismus MeSH
- leukemie genetika krev MeSH
- lidé MeSH
- prekurzorové buňky granulocytů cytologie enzymologie účinky léků MeSH
- proteosyntéza genetika imunologie účinky léků MeSH
- stárnutí buněk fyziologie genetika MeSH
- Check Tag
- lidé MeSH
